Research Report

Transcription

1 Research Report

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3 Preface The Equine Reproduction Laboratory (ERL) has been an active education, research, outreach, and clinical service unit of Colorado State University since The ERL has evolved significantly through the years and is recognized as one of the premier programs in equine reproduction in the world. The ERL is a unit within the Animal Reproduction and Biotechnology Laboratory (ARBL) and the Department of Biomedical Sciences. ERL currently is home to 4 primary faculty, 1 staff veterinarian, 3 research associates, 1 postdoctoral student, 5 graduate students, 2 veterinary residents, and 2 equine breeding farm interns. Additional faculty and staff providing key contributions to the mission of the ERL are located in the Departments of Clinical Sciences and Animal Sciences as well as the Veterinary Teaching Hospital. The ERL plays an active role in the education of undergraduate, graduate, postdoctoral, and veterinary students. A wide variety of didactic lectures, hands-on laboratories, and elective clinical rotations are available. In addition, 7 continuing education courses for horse owners and 2 continuing education courses for veterinarians have been offered annually. Research is a core commitment of the ERL faculty, staff, and graduate students. The ERL has a 40-year history of ground-breaking research in the areas of reproductive physiology, pathology, therapeutics, and management. In the past 5 years, more than 150 peer-reviewed scientific publications and research abstracts have been published by members of the unit. Innovation translated into clinical services has long been a hallmark of the ERL. More than 500 client-owned horses are examined annually at the ERL. Mare services offered range from breeding by live cover, artificial insemination with fresh, cooled or frozen semen, pregnancy examinations, embryo transfer, and assisted reproduction techniques such as oocyte transfer and intracytoplasmic sperm injection. Stallion services include breeding soundness evaluations, collection of semen for cooled-transport or cryopreservation, and behavioral training. The clinical program is also recognized for expertise in the diagnosis, treatment, and/or management of reproductive problems in mares and stallions. The goal of this publication is to outline the recent accomplishments (2007 to 2010) of the faculty, staff, and graduate students of the ERL. Dr. Patrick M. McCue Iron Rose Ranch Chair in Equine Reproduction Equine Reproduction Laboratory Senior Veterinarian in Charge of ERL Clinics Dr. Thomas R. Hansen Traubert Professor Director, Animal Reproduction and Biotechnology Laboratory The Equine Reproduction Laboratory is a program of the Animal Reproduction and Biotechnology Laboratory, Colorado State University 1

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5 Table of Contents Preface... 1 Table of Contents... 3 Mission Statement... 5 Accomplishments Strategic Plan Goals for Future Studies Personnel Faculty Collaborating Scientists Areas of Expertise of Core Personnel Staff Veterinarian Research Associates Current DVM Residents Graduate Students Postdoctoral Student Interns Synopsis of Program Overview Facilities Research Firsts Undergraduate Education PVM Teaching Equine Theriogenology Residency Program Intern Program Awards Editorial Boards Grant Proposal Review Boards Research Support Major Donors Equine Scholarship Funding Through ARBL and ERL Preservation of Equine Genetics (PEG) Program

6 Funded Research Projects Scientific Publications Chapters in Books Refereed Journal Articles Abstracts Invited Presentations Short Courses Visiting Scientists Past Graduate Students / Placement

7 Mission Statement The Equine Reproduction Laboratory is committed to pursuing excellence in education, performing innovative research, and providing outstanding clinical service to horse owners. Elements key to the Mission: 1. Excellence in education for undergraduate, graduate, postdoctoral, and professional veterinary medicine students 2. World class programs of basic and applied research in the mare and stallion 3. Dedication to horse health and clinical service 4. Dissemination of knowledge through continuing education 5

8 Accomplishments Uterine Infections Bacterial and fungal infections of the uterus are a significant problem in mares. Uterine infections can be difficult to detect and are often associated with reduced fertility. Dr. Ryan Ferris, in collaboration with Drs. Mike Lappin and Julia Veir from the Department of Clinical Sciences, has developed a rapid, quantitative PCR assay for detection of bacterial or fungal DNA in equine uterine samples. The RT-PCR assay is able to detect minute quantities of bacterial or fungal DNA in uterine swab, lavage, or biopsy samples. Timely and accurate identification of a pathogenic organism is critical for development of an optimal therapeutic program. Foal IgG Assessment Embryo Biopsy Foals are born without immune protection and receive immunoglobulins through ingestion of colostrum within the first few hours after birth. Foals that do not obtain sufficient antibodies through passive transfer are susceptible to infectious disease. We have developed a rapid immunoturbidimetric assay for quantitative measurement of IgG concentration in the serum or plasma of foals. Early detection of complete or partial failure of passive transfer can allow for supplementation with frozen colostrum or plasma and a decrease in the risk of morbidity or mortality due to infectious diseases. Evaluation of cells (blastomeres) removed from an early equine embryo can allow for detection of genetic abnormalities or determination of sex. Drs. George Seidel and Jennifer Barfield have developed a technique for aspiration of a small number of blastomeres from morula or early blastocyst stage embryos using a micromanipulator. The aspirated cells can be submitted for genetic testing and the biopsied embryos cryopreserved for later transfer into a recipient mare. Current research is involved in optimizing the biopsy and cryopreservation techniques. 6

9 Stallion Spermatozoa Over the past few years the ERL has examined new technologies for cryopreserving stallion spermatozoa, with specific interest in increasing cholesterol content of the sperm. This technique leads to higher percentages of sperm surviving cryopreservation, but the added cholesterol may lead to problems in sperm capacitation. This has led to studies involving methods to capacitate stallion sperm in vitro as well as develop techniques to evaluate sperm capacitation. This work has led to several different methods for capacitating stallion sperm in vitro, including the use of cyclodextrins, ionophores, lysophospholipids and a short-chained phospholipid (PC12). Each of these compounds induces several of the characteristics associated with sperm capacitation. In addition, a technique was developed to test in vitro fertilizing capacity of these sperm, using a heterogametic IVF system, in which stallion sperm fertilized bovine oocytes in vitro. Using this system we were able to determine the optimal treatment to capacitate stallion sperm in vitro. Unfortunately these sperm did not fertilize equine oocytes in vitro, leading us to hypothesize that the problem is not with the sperm, but with some aspect of oocyte zona-hardening. This is being tested currently. We have also developed new techniques for sperm analysis, including developing a computer program that can identify stallion sperm that exhibit a unique type of swimming motion, hyperactive motility, which is associated with sperm capacitation. In addition, the system was used to analyze the motion of sperm treated with compounds that alter the biochemistry of sperm in ways that should induce hyperactive motility. MicroRNA in Mares with Granulosa Cell Tumors Granulosa cell tumors (GCTs) are the most common ovarian tumor of the mare. Affected mares may show behavioral abnormalities, such as persistent estrus or stallion-like behavior. Diagnosis is made by a combination of clinical signs (i.e. abnormal behavior), transrectal ultrasonography (i.e. a unilaterally enlarged ovary and a small inactive contralateral ovary), and hormone analysis (i.e. elevation in plasma inhibin and testosterone concentrations). Treatment is surgical removal of the affected ovary. Unfortunately, not all GCTs are accurately identified by traditional procedures. MicroRNAs are small (20-23 nucleotide), single-stranded RNA molecules that offer potential as biomarkers in the diagnosis, monitoring, and as prognostic indicators for certain cancers. Identification of mirnas that are conserved between horses and humans offers future hope for diagnostic tests in other species as well, providing a broad forum of true translational medicine. Dr. Alicia Lindholm, a Resident in Equine Reproduction, working in conjunction with Dr. Gerrit Bouma (ARBL) and Dr. Lynne Bemis (University of Colorado, Denver, Division of Medical Oncology) has identified several mirnas that are elevated in the serum of mares with GCTs as compared to normal cycling mares. Pharmacokinetics Bacterial and fungal infections of the uterus are two of the most important causes of reduced fertility in mares. Antimicrobial protocols for treatment of equine infectious endometritis are often based on data from other species. Drs. David Scofield and Ryan Ferris have conducted a series of studies investigating the pharmacokinetics of antimicrobial medications administered orally or as 7

10 an intrauterine infusion. The goal of the studies is to determine the concentrations of drugs achieved in the serum and in endometrial tissue and compare those levels with the minimum inhibitory concentration (MIC) required to kill or prevent replication of potential uterine pathogens. Studies have been completed recently on fluconazole (oral administration) and miconazole (intrauterine administration), and a third study will be conducted in 2011 in collaboration with Dr. Jerry Black on the pharmacokinetics of a long-acting ceftiofur antibiotic (intramuscular administration). Maternal Recognition of Pregnancy Maternal recognition of pregnancy depends on endometrial responses to a conceptus, including attenuation of release of prostaglandin F2α (PGF2α), and continued production of progesterone by the corpus luteum (CL). Exactly how this occurs is unknown. Because PGF2α induces proinflammatory responses, we hypothesized that gene expression in the endometrium at the time of maternal recognition entailed an anti-inflammatory event. Mares were not inseminated (non-pregnant), or inseminated and then endometrial biopsies were recovered on day 18 post-ovulation. Endometrial gene expression profiles were determined by screening an Affymetrix equine GeneChip array containing probes specific for genes related to inflammatory processes. Microarray analysis, using Affymetrix equine GeneChip array revealed 118 genes that were up-regulated and 93 genes that were down-regulated (P 0.001) at least 1.5 fold in the endometrium of pregnant versus nonpregnant mares. Microarray results for 3 up-regulated genes homologous to TSC22D3 (a zinc finger binding transcription factor that regulates inflammatory and immunosuppressive responses), PPAPDC2 (phosphotidic acid phosphatase 2; involved with innate immune responses) and KLF6 (Krueppel-like factor; a tumor suppressive gene), and 3 down-regulated genes, ESR1 (estrogen receptor; induces PGF2α release) and genes homologous to MARCKSL1 (macrophage myristoylated alanine-rich C kinase substrate; couples protein kinase C and calmodulin signaling associated with macrophage function) EPSTI1 (epithelial-stromal interaction protein; associated with tumorogenesis and endometrial remodeling) (P<0.05) were confirmed using Real Time PCR. It is concluded that presence of the equine embryo induces differential gene expression that is associated with endometrial inflammatory processes, cellular growth and proliferation in Day 18 pregnant mares. A better understanding of these maternal responses to pregnancy may be used to improve fertility and embryo survival in mares. Morphological Services The Morphological Services at ARBL provides clinical and research support services to domestic as well as foreign clients, clinicians, and researchers. These services, encompassing a variety of species (including horses, cattle, sheep, pigs, dogs, cats, and laboratory animals), facilitate detection of pathologic conditions in the male reproductive system and/or assessment of potential fertility. Services include: Evaluation of semen for sperm morphology and ultrastructural pathology 8

11 Histopathology, immunocytochemistry, and transmission electron microscopy of a variety of tissues following necropsy or biopsy in various clinical conditions or research paradigms. This broad-based service integrating pathophysiological, toxicological, and clinical aspects of reproduction in the male has been recognized far and wide. In the last ten years, more than 400 light and electron microscopic evaluations of sperm were performed for stallions alone. Chromosomal Alignment in Equine Oocytes from Young and Old Mares Occurrences of chromosomal problems within the oocytes of older women have been well established in human medicine, although the causes are not definitively known. In the human, problems in chromosomal alignment can result in the birth of babies with genetic problems. Few foals from old mares are born with abnormal chromosomal complements; however, this is probably a contributing factor to high pregnancy losses in older mares. Drs. Carnevale and Maclellan collected oocytes from the preovulatory follicles of young and old mares. The oocytes were fixed, stained and examined with confocal microscopy in collaboration with Dr. David Albertini at The University of Kansas Medical Center to obtain detailed images. Using these images, alignment of the chromosomes in mature oocytes is being compared between young and old mares. 9

12 Cryopreservation of Equine Oocytes The first foals to be produced from cryopreserved oocytes were at CSU in the early 2000s. Since then, no additional foals have been produced; however, significant advances have occurred in methods to cryopreserve human oocytes. Dr. Lisa Maclellan (visiting scientist) worked in collaboration with Drs. Elaine Carnevale and Dr. Kim Preis (Colorado Center for Reproductive Medicine) to cryopreserve equine oocytes using vitrification (a rapid cryopreservation technique). Some of the oocytes were warmed, injected with sperm, and the early embryos were transferred into recipient mares to produce pregnancies. Additional oocytes were collected from young and old mares, vitrified, warmed, and examined using confocal microscopy. This work was done in association with Dr. Albertini s laboratory to determine the effect of cryopreservation on the oocyte. The longterm goal of this research is to develop methods to cryopreserve equine oocytes. We are continuing this research to optimize results. In the future, we hope to have an efficient system to cryopreserve oocytes from mares with valuable genetics and from ovaries salvaged after the death of a mare or endangered equid. An oocyte with poor morphology is seen in this picture. The space between the ZP and egg is large. These changes are often associated with reduced potential for the egg to develop into a normal pregnancy after sperm injection. The pipette on the left is holding the egg in place, while the sperm is injected into the egg using the pipette on the right. Changes in Oocyte Morphology Associated with Aging and Developmental Competence Until recently, relatively little was known about the morphology of the equine oocyte. However, with the development of procedures for sperm injections (ICSI) in the horse, equine oocytes are now frequently observed under a high-powered microscope. While most oocytes from young, fertile mares have a similar morphology, some oocytes from older, subfertile mares seem to have distinct abnormalities. In conjunction with our clinical Advanced Assisted Reproduction Program, the morphology of oocytes from young and old mares have been studied prior to ICSI to determine if certain changes are associated with mare age or oocyte quality. Before an oocyte is 10

13 injected with a sperm, an image of the oocytes is obtained through a high-powered microscope. The pictures are being examined using a computer program to establish indicators of oocytes that have high or low potential of producing a viable pregnancy. Use of Sex-Sorted Sperm for ICSI Methods to separate sperm bearing X (female) or Y (male) chromosomes have been developed in the horse, but obtaining sufficient numbers of sex-selected sperm for standard inseminations can be difficult. However, relatively few sperm are required for ICSI, allowing sex selection prior to sperm injection. Dr. Graham developed methods to refreeze semen that had been frozen, thawed, and sexed. The semen was then thawed and used for ICSI. Over 80% of oocytes injected with control (non-sexed) sperm resulted in embryos, while 30% of oocytes injected with sex-sorted, refrozen sperm produced embryos. Success of the sex-sorted, refrozen semen seemed to vary with the individual stallion. Dr. Carnevale s group then injected three oocytes with X-bearing sperm from the most successful stallion in the project. The resulting embryos were cultured in an incubator prior to being transferred into recipients uteri. Two recipients became pregnant and gave birth to fillies. Cell-to-Cell Communication Within the Equine Follicle Oocyte quality and competence is acquired during follicular growth and maturation; i.e., folliculogenesis. Studies have demonstrated that folliculogenesis involves extensive cell-to-cell communication between somatic cells (theca and granulosa cells) and somatic cells (granulosa cells) and oocytes. This communication is essential for oocyte growth and maturation, and as such, oocyte quality and competence. Follicular growth is characterized by increased follicular size due to increased somatic cell number and development of an antrum containing follicular fluid. The role of follicular fluid (FF) in oocyte growth and maturation is unclear. FF is a product from blood serum constituents and secretory products from theca and granulosa cells; FF helps constitute the environment for oocyte maturation. Components of FF include hormones (e.g., estrogen), members of TGFβ signaling family, and amino acids. Data from our laboratory revealed that small non-coding RNA molecules, called mirnas, are present in equine FF. MiRNAs play an important role in regulating gene expression and function and are a critical component underlying cell differentiation. We further demonstrate that FF isolated from oocytes in young and old mares contains significant differences in relative amounts of selected mirnas. Currently, our efforts focus on the presence of mirnas in cell-secreted vesicles within the follicle and their role in cell communication and follicle development. 11

14 Determining Hierarchy of KiSS Kisspeptin is a small peptide that may be the signal controlling seasonal reproduction in the horse as well as communicating signals for fetal-placental well-being. This work was initiated by Dr. Christianne Magee in collaboration with members of the ARBL and ERL. The group provided the first anatomical and biological evidence for a role for kisspeptin in the mare. Current objectives include developing a radioimmunoassay for kisspeptin and using non-terminal methods to study the ability of kisspeptin to release gonadotropin releasing hormone (GnRH) from the hypothalamus (part of the brain). GnRH causes the secretion of luteinizing and follicle stimulating hormone from the pituitary gland (Picture: Kisspeptin (red) and GnRH (green) at the connection between the hypothalamus and pituitary gland, Endocrinology 2009). 12

15 Strategic Plan The College of Veterinary Medicine and Biomedical Sciences (CVMBS) and the College of Agricultural Sciences (CAS) recently conducted strategic planning discussions for the Equine Reproduction Laboratory (ERL), which is a program within the Animal Reproduction and Biotechnology Laboratory (ARBL). Two very positive events occurred for the ERL as a consequence of such extensive strategic planning. The first is that we recently learned we can start advertising for a new faculty member to serve as the Alexander Chair in Equine Reproduction. This person will be expected to provide senior research leadership in equine reproduction. The second positive action by the CVMBS is that we received permission to also hire an additional faculty member with expertise in equine clinical reproduction services. This Clinical Sciences hire will fill the position created by Dr. McCue s recent honor of being named the Iron Rose Ranch Chair in Equine Reproduction. Being able to hire two new faculty members into the ERL and ARBL is fantastic in context of the many economic issues we are addressing at CSU. The research training of graduate students is designed to develop: 1) a philosophy of research centered on careful delineation of the hypothesis to be tested combined with critical selection and validation of methods to be used, adequate replication, thoughtful interpretation, and timely and concise publications; 2) mastering diverse research techniques that allow pursuing new problems aggressively and developing new approaches with confidence; and 3) functioning as an independent investigator as well as team researcher (Fig. 1). To accomplish these training goals, our students generally are involved with several projects and collaborate and interact with research units within the ARBL and ERL and throughout the scientific community at CSU. Because the ERL is involved with basic, applied-translational, and clinical research, and has very strong relationships with biotechnology companies and industry, our students are trained to think in broad, global terms, rather than with narrow disciplinary perspectives. These activities also assist in placement of our students into other professional training. Figure 1. ERL Collaborative Missions in Reproductive Sciences. 13 The ERL has been an active research, education, and clinical unit at Colorado State University since The ERL has grown significantly through the years and is recognized as one of the premier programs in equine reproduction in the world. The success of the ERL stems from participating and collaborating faculty from two colleges and three departments at CSU. Dr. Patrick McCue, Professor and Iron Rose Ranch Chair in Equine Reproduction, serves as the veterinarian in charge of theriogenology clinical activities at the ERL and specifically manages all general mare services. Dr. Elaine Carnevale is an Associate Professor and manages the oocyte transfer and intracytoplasmic sperm injection (ICSI) services at the ERL. Dr. Jason Bruemmer is a Professor and manages the stallion services at the ERL. Dr. Ryan Ferris is a lecturer and primarily contributes to mare services at the ERL. Other ARBL faculty contribute significantly to this program,

16 but these four primary faculty represent the core leadership at the ERL along with the ARBL-ERL Director, Thomas Hansen. Currently, the ERL is home to 4 primary faculty, 4 research associates, 1 postdoctoral student, 5 graduate students, 2 veterinary residents, and 2 equine breeding farm interns. Additional faculty and staff providing key contributions to the mission of the ERL are located in the ARBL, Veterinary Teaching Hospital, Department of Clinical Sciences, and the Department of Animal Sciences. The ERL plays an active role in the education of undergraduate, graduate, and veterinary students. In addition, 7 continuing education courses for horse owners and 2 continuing education courses for veterinarians have been offered annually. Research remains a core commitment of the ERL faculty, staff, and graduate students. The ERL has a 40-year history of ground-breaking research in the areas of reproductive physiology, pathology, therapeutics, and management. In the past 5 years, more than 150 peer-reviewed scientific publications and research abstracts have been published by members of the unit. From a clinical perspective, more than 500 client-owned horses are examined annually at the ERL. Mare services offered range from breeding by live cover, artificial insemination with fresh, cooled or frozen semen, pregnancy examinations, embryo transfer, and assisted reproduction techniques such as oocyte transfer and intracytoplasmic sperm injection. Stallion services include breeding soundness evaluations, collection of semen for immediate use, cooled-transport or cryopreservation, and behavioral training. The clinical program also is recognized for expertise in the diagnosis, treatment, and/or management of reproductive problems in mares and stallions. Innovation translated into clinical relevance is a hallmark of the ERL. Following are bullets from the ERL Research and Clinical-Outreach-Education strategic planning process. Bullets from the ERL Strategic Research Plan ERL Strategic Goals o Develop Strategic Research Faculty Hiring Plans o Strengthen Graduate Research Education Plan o Develop Strategic Funding Plans for Research o Develop Strategic Research Plan Develop Strategic Research Faculty Hiring Plans o Alexander Chair Position (new opportunity for ERL to grow and expand!) 14

17 o o Research in equine reproduction is a clear expectation New Alexander Chair will contribute to leadership in growth and direction of research in equine reproduction by ERL and ARBL faculty Strengthen Graduate Research Education Plan o Extensive document for Masters Plan A program was adopted last year and has been implemented to improve the graduate educational experience Strategic Funding for Research o Complete historical research reporting and initiate new ERL Research Foundation program called ERL-RF o Develop strategic funding plan for the following critical needs at the ERL Research Scholarships for Honors Undergraduate Research Scholarships (fellowships) for Graduate Education Facilities and Equipment Resident/PhD Fellowships Research Plan (strategic plans were developed for the following emphasis areas) o Equine Genetic Preservation o Assisted Reproductive Technologies o Fertilization and Pregnancy o Contraception o Reproductive Physiology o Reproductive Pathology Bullets from the ERL Clinical Services, Outreach, and Education Strategic Plan ART Services o Continue excellent services that are provided o Develop depth beyond one person in any given expertise o Train senior veterinary partners and new residents in key technical phases of ART Mare Services o Continue excellent mare services that are provided o Boost awareness of expertise and focus on the problem mare o Continue research in this area to strengthen mare services Stallion Services o Continue excellent stallion services o Encourage research studies in stallions for development of future clinical innovation o Expand stallion customer service role and educational opportunities for ERL faculty and residents o Provide highest standards of care and customer service for stallions 15

18 o Succeed in a strategic Alexander Chair and/or Clinical Sciences faculty hire to strengthen stallion research and services Budgetary Management and Reporting o Collaborate with other revenue services (VTH) when developing reports to enhance consistency across the College and provide more timely information to the faculty o Refine monthly reports and indicators o Improve E-thority financial reports to provide real-time accounting information and access o Complete cost versus revenue analysis for services at ERL o Employ a DVM/MBA for assistance with analysis Develop Client Survey for Feedback on Services o Survey on site, mailed or phone interview Enhance Existing Equine Sciences Undergraduate Experiences at the ERL o Formalize ERL internships for CSU Equine Sciences students Study Cooperative Education so it Becomes More Profitable o Expand one short course to on-line offering as starting point o Include lectures in Spanish as a mechanism to attract a larger audience o Reduce costs; possibly limit to fewer days with online pre-course Enhance Visibility o Advertising, attendance at meetings, Facebook, web site Hire Faculty Member to Assist with Mare and Stallion Services o DVM and Board Certification is preferred; PhD is desirable o Possibly the future primary contact for stallion services Restructure ERL Leadership o Provide faculty and personnel the opportunity to maximize their strengths o Re-establish the historical administrative structure that recognizes the ERL as a program within the ARBL o Dr. McCue, as Iron Rose Ranch Chair and senior member of the ERL faculty, will serve as primary contact and veterinarian in charge of ERL clinics, outreach, and teaching efforts; he also will be the primary point person for development and interaction with the public and press o The person hired as the Alexander Chair in Equine Reproduction will serve as an ERL Research Leader Prepared by the 2011 ERL Strategic Planning Committee: Dr. Terry Nett, CVMBS Associate Dean served as chair of this strategic planning group. Dr. Thomas Hansen, ARBL Director, served as chair of the research strategic planning group which was composed of Dr. Terry Nett (ex officio), Dr. Pat McCue (ERL Director), Dr. Elaine Carnevale, Dr. Jason Bruemmer, Dr. James Graham, and Dr. Quinton Winger. Dr. Christopher Kawcak served as chair of the ERL clinical strategic planning group which was composed of Dr. Terry Nett (ex officio), Dr. Dean Hendrickson (Director, VTH), Dr. Jerry Black (Director, Equine Sciences Program), Dr. Jason Bruemmer (Animal Sciences), Dr. Elaine Carnevale (Biomedical Sciences), and Dr. Patrick McCue (Clinical Sciences). 16

19 Goals for Future Studies Equine Genetic Preservation Introduction With the acceptance of foal registration from cooled or frozen sperm and embryos by many breed registries, interest in these technologies has increased. Although foals have been produced using all of these technologies, they are not efficient for all genetic donors or all types of gametes or embryos. For example, although cooled and frozen stallion sperm are used routinely, sperm from some stallions do not survive cooling well (and cannot be used for cooled shipped semen) and sperm from even more stallions do not survive freezing. Similarly, although small embryos (6- to 6.5-day embryos) survive freezing quite well; larger embryos, that can be easily flushed 7-8 days after insemination, do not survive freezing well. And equine oocytes, regardless of source, do not freeze well. Impact and Significance The ability to utilize frozen semen from more stallions would have a great impact on the equine industry. Improving cryosurvival rates of stallion sperm would benefit the industry, as sperm could be used to inseminate many mares; moreover, the ability to cryopreserve sperm that have been sex-sorted could add to this impact. The ability to utilize cryopreserved embryos and oocytes would affect fewer individuals, but the genetics conserved are likely to be from elite females and, therefore, could significantly affect the industry. In addition, this research would improve our understanding of cryopreservation in general, and specifically on how membrane composition, cryoprotectant interactions, and cellular osmotic tolerance affect the cryosurvival rates of cells. Goals 1. Develop new techniques for preserving stallion sperm by developing more universally tolerated cryopreservation diluents and methods to alter sperm membrane lipids. 2. Develop techniques to cryopreserve oocytes, ovarian tissue slices, and large embryos. 3. Develop techniques to cryopreserve various equine stem cell lines. Assisted Reproductive Technologies Introduction A wide range of assisted reproductive technologies are being used in the equine industry, providing more efficient use of valuable equine genetics and providing new insight into equine reproductive physiology. CSU has been a leader in the development of assisted reproductive technologies (ARTs) including artificial insemination, embryo transfer, and oocyte procedures. 17

20 Impact and Significance Impact on equine industry. The development of ART at CSU has resulted in profound changes in the equine industry from methods for artificial insemination to storing and transporting embryos, semen, and oocytes. Many of the procedures that were developed or advanced at CSU are commonly used by equine breeders. The program at CSU has trained numerous students, visitors and short course participants, who have helped to disseminate these technologies throughout the world. Impact on research. The ability to collect and manipulate equine gametes has provided an opportunity to advance our understanding of gamete physiology, fertilization, and early embryo development. The horse is one of few species in which advanced ARTs, such as intracytoplasmic sperm injection (ICSI), are used for clinical purposes, making the horse a potential animal model to advance and study the effects of ARTs in humans. Goals 1. Improve the efficiency and flexibility of ARTs, to reduce costs and increase use of the procedures in the equine industry through methods such as laparoscopic oocyte transfers, optimal sperm selection, and shipment of harvested ovaries and testes, oocytes, and ICSI-produced embryos. 2. Advance our knowledge and competency in embryo culture, including effects of culture conditions on long-term animal health. 3. Use the unique research opportunities provided by advanced ARTs to study factors affecting oocyte viability, including maternal aging. 4. Study the interaction of the equine sperm and oocyte to determine what factors prevent in vitro fertilization in the horse and develop methods to produce offspring using IVF. 5. Increase the efficiency of procedures to increase the number of oocytes or embryos for collection per estrous cycle. Fertilization and Pregnancy Introduction Although thousands of foals are born every year, it is noteworthy to mention that the actual mechanisms involved in fertilization remain a mystery. The interaction between the oocyte and the sperm are vaguely defined and most often related to appropriate timing and morphological normality. Listed below are areas not fully understood in normal fertilization. The horse world is currently faced with two dilemmas related to reproduction. The first is an ever-growing desire to maximize reproductive efficiency, and the second is a need for an effective contraception protocol. A thorough understanding of the events leading to and following the gamete interactions will potentially allow for both a more efficient mating and/or an ability to effectively contracept the horse. Impact and Significance Gametogenesis: Oocyte. Current dogma suggests that the filly is born with all of the oocytes she will ever produce, arrested in the ovarian stroma. Understanding oocyte production, their meiotic 18

21 arrest, re-activation, and maturation within the pre-ovulatory follicle are essential. These include a thorough identification of zona pellucida production and subsequent function as the failure of IVF is thought to be associated primarily with this structure. Potential Outcomes: Improved preservation of genetic material Improved in vivo maturation techniques Diagnostics related to oocyte quality Contraception Gametogenesis: Spermatogenesis. Spermatozoa production is an exciting area of research. We believe that the answers to many mysteries may be found in this process, not the least of which is true stem cell production as is occurring constantly in spermatogonial regeneration and spermatocyte generation. Additionally, we hope to focus on the processes that contribute to the metamorphosis of spermatocytes into spermatozoa and the associated maturational signals occurring along the epididymis. We have interesting preliminary data suggesting such things as direct action of GnRH at the site of the testis, epididymis, or sperm cell itself. Potential Outcomes: Germ cell production Improved sperm production Improved sperm quality Diagnostics related to sperm quality Contraception Fertilization. We will define fertilization as the actual fusion of the equine gametes. However prior to their interaction it is imperative that the following pre-fertilization processes are adequately defined: From the mare s side we are investigating sperm capacitation, acrosome reaction, oocyte/ oviductal interaction, identification, and creation of the oviductal sperm reservoir. On the male side we must better define the chemical composition and role of seminal plasma as it relates to capacitation, which allows for zona binding, acrosome reaction and zona penetration. Our studies have provided great insight into the mechanisms associated with the acrosome reaction and will continue until this rate-limiting event is thoroughly defined. Oocyte activation initiated by the acrosome-reacted sperm, which leads to the second resumption of meiosis, also is being studied. Potential Outcomes: Successful IVF Production of transgenics Contraception Maternal Recognition of Pregnancy. Immediately following successful fertilization it is imperative that the embryo be recognized by the dam so that 1) she ceases to cycle and 2) the oviductal and uterine environment support the developing pregnancy. To date, the oviductal signal has been identified but the mode, method, or signal for communication between the embryo and uterus remain unknown. We hypothesize that this signal is initiated by the embryo and results in detectable changes in gene expression, endocrine function, and metabolism of the endometrium and potentially the corpus luteum. 19

22 Potential Outcomes: Early pregnancy diagnostics Improved pregnancy rates Contraception Goals 1. Develop diagnostics to evaluate normal physiologic functions of gametes. 2. Develop diagnostics to evaluate normal physiologic functions of an embryo and developing fetus. 3. Develop therapeutics to address abnormalities in gametes and embryos. 4. Develop accurate early pregnancy diagnostics. 5. Develop efficient and effective contraceptives. Contraception Introduction There is a need for techniques for both contraception and sterilization of both sexes of horses. Impact and Significance Contraceptives that could be applied one time and inhibit reproductive activity in either mares or stallions for one year would be very useful for performance horses. Also, there is a substantial need for a sterilant for controlling wild horses and burros that inhabit federal lands and parks. Ideally, an injectable agent that induced sterility in either females or males would help mitigate the burgeoning population of horses and burros that roam the west. Such a compound also could be used to chemically sterilize domesticated mares or stallions as an alternative to surgical castration. Goals 1. Develop procedures that will interfere with fertility in both mares and stallions for up to one year. 2. Develop procedures that will sterilize either mares or stallions to permanently interrupt reproductive processes. Reproductive Physiology Introduction Significant knowledge of normal reproductive physiology of the mare and stallion has been gained over the past 50 years. However, there is much more to learn and current research should build on past experiences as well as explore new diagnostic tests and procedures for enhancing reproductive efficiency. 20

23 Impact and Significance Reproductive physiology encompasses a wide variety of reproductive functions, including neuroanatomy and endocrinology, testicular and ovarian physiology, and physiology of the male and female reproductive tracts. Moreover, a better understanding of the factors responsible for controlling the transition from anestrus to the breeding season and back to anestrus are essential to optimizing reproductive performance in the horse. The ultimate goals of the male and female reproductive systems are to provide an opportunity for male and female gametes to combine their genetic material, provide a nurturing environment for that newly created individual during embryonic and fetal development, and provide immune protection and sustenance to the newborn after birth. Additional basic and applied research is needed for a better understanding of normal physiologic events which can be translated into enhanced success of equine breeding programs. The ultimate measure of reproductive success is the birth of live, healthy foals. Additional parameters that measure success may include ovulation rates, per cycle or per season pregnancy rates, embryo collection or transfer success rates, number of sperm per ejaculate, percentage of progressively motile sperm after cryopreservation, or other factors. Goals 1. Conduct basic research on physiologic mechanisms that control reproductive function in mares and stallions. 2. Develop new diagnostic tests to evaluate normal physiologic functions in mares or stallions. 3. Develop novel products, techniques, or protocols to enhance reproductive success in mares or stallions. Reproductive Pathology Introduction A wide variety of transient physiologic and permanent pathologic conditions adversely affect reproductive performance in mares and stallions. In non-pregnant mares, ovarian abnormalities such as failure of follicular development and failure of ovulation preclude mares from an opportunity to become pregnant. Persistent mating-induced endometritis and infectious endometritis create a uterine environment that is inhospitable to the developing embryo. Early embryonic death and bacterial placentitis are significant causes of pregnancy loss in mares. Testicular degeneration and epididymal dysfunction are two of the many issues that lead to reduced fertility of stallions. Impact and Significance Basic research is critical to understanding the pathophysiology of reproductive diseases in all domestic animals, including the horse. In many instances, the events that lead to a particular abnormality are not well understood. Advances in technology often lead to breakthrough scientific research that improves our knowledge of pathology or disease. 21

24 Applied or clinical research is needed to develop accurate and sensitive diagnostic tests for detection of the presence of a reproductive disease or abnormality. An accurate diagnosis is a key element in resolving reproductive abnormalities in the horse. Once an abnormality is detected, additional hypothesis-driven research is needed to develop and evaluate management or treatment strategies. Goals 1. Evaluate pathophysiologic mechanisms that lead to reproductive abnormalities in mares and stallions. 2. Develop new diagnostic tests or protocols to detect reproductive abnormalities in mares or stallions. 3. Develop novel therapeutic products, techniques or protocols to manage or treat reproductive abnormalities in mares or stallions. 22

25 Personnel Faculty Gerrit J. (Jerry) Bouma, PhD, Assistant Professor of Biomedical Sciences Dr. Gerrit (Jerry) Bouma is an Assistant Professor in the Department of Biomedical Sciences. He obtained his BSc degree in 1998 in Environmental Sciences (with an emphasis in environmental toxicology) from the Van Hall Institute in the Netherlands, his PhD degree in 2003 in Zoology (with an emphasis on Reproductive Biology) at the University of Idaho. From he was a post-doctoral fellow at the Jackson Laboratory (Bar Harbor, Maine), focusing his studies on the genetic control of mammalian fetal gonadal development. In 2006 he accepted a faculty position in the Department of Biomedical Sciences at Colorado State University. Dr. Bouma focuses his studies on (1) genetic control of mammalian fetal ovarian development, (2) the role of non-coding RNAs (mir- NAs) in regulating ovarian follicle development and growth, and (3) the function of pluripotency factors and non-coding RNAs in ovarian cancer and placental stem cells. In addition, he has active collaborations working on projects relating to the development of novel animal models to study the role of environmental contaminants in inducing testicular cancer. Recent data generated indicates the exciting possibility that a new form of cell communication exists within the mature ovarian follicle involving small cell-secreted vesicles containing mirnas and proteins. This study used the mare as an exceptional model as the mare s anatomy and physiology is such that it allows relatively easy access to ovarian tissue, enabling frequent sampling and monitoring of follicular growth and maturation, and also will enable us to study involvement of vesicles in cell communication and regulation of cell function in vivo. Data generated from these studies will have important application in reproductive physiology and provide unique opportunities to develop new diagnostic markers and tools for reproductive dysfunction and disease. Research Interests: Genetic and molecular control of fetal gonad development; Adult reproductive function 23

26 Jason E. Bruemmer, PhD, Professor of Animal Sciences Dr. Jason E. Bruemmer, Professor, received his BS degree in Animal Science and his MS degree in Physiology of Reproduction from Texas A&M University, and his PhD in Reproductive Physiology from New Mexico State University. Dr. Jason Bruemmer grew up in the horse industry in Texas and has continued to serve the industry as a scientist, researcher, lecturer and horseman. As a horseman, Dr. Bruemmer has been a cornerstone of the outreach program at CSU, working with breeders, owners, and clients to increase their knowledge of equine reproduction and management. Dr. Bruemmer s ability to interact and relate to horse owners and breeders has informed his research. He was instrumental in developing a protocol to harvest epididymal sperm from stallions who had died, allowing the owner to preserve the valuable genetics for future production. Stallion behavior, physiology, and management are major fields of interest to Dr. Bruemmer as he heads up the Stallion Services program at the ERL. His research in the cryopreservation of stallion semen and membrane integrity continues to work for improvement in stallion semen extenders. Stallion behavior is an often overlooked component of reproductive management and fertility assessment. Dr. Bruemmer s ongoing work in this field continues to have very productive results with stallions referred for breeding problems. A mentor to graduate students, veterinary students, and undergraduates, Dr. Bruemmer lectures on Equine Reproduction, Equine Behavior, Equine Management, Equine Industry, and the very popular Foaling module. As a researcher, Dr. Bruemmer has published over 40 scientific articles. In 2004, Dr. Bruemmer was a visiting scientist at the Vincent Center for Reproductive Biology at Massachusetts General Hospital at the Harvard School of Medicine. Recently, Dr. Bruemmer has been pursuing the answer to the age-old question: How does the mare know she s pregnant? Maternal recognition of pregnancy is well understood in most species...except the horse. The techniques in molecular biology acquired during his stay in Boston have allowed for some very interesting research and promising answers. As an avid horseman and polo player, Dr. Bruemmer still finds time to work with the CSU polo team and finds time for the occasional chukker himself. Research Interests: Cryopreservation of stallion semen; Sperm membrane integrity; Maternal recognition of pregnancy in the mare; Luteal tissue formation, function, and demise 24

27 Elaine M. Carnevale, DVM, PhD, Associate Professor of Biomedical Sciences Dr. Elaine Carnevale is an Associate Professor in the Department of Biomedical Sciences. She received her DVM in 1985 from Colorado State University. Dr. Carnevale continued at Colorado State University s Equine Reproduction Laboratory to receive her MS degree in equine reproduction, studying methods for the cooled shipment of embryos. For the next few years, Dr. Carnevale worked in the horse breeding industry in New Zealand, Ireland, and Australia. She returned to the United States to obtain her PhD degree from the University of Wisconsin under Dr. O.J. Ginther. Research for her PhD centered on reproductive function in the old mare. Upon graduating from the University of Wisconsin, Dr. Carnevale accepted a faculty position at Southern Illinois University in Carbondale, Illinois, where she taught undergraduate and graduate classes, conducted research in equine reproduction, and provided veterinary care for the University s animals. In 1997, Dr. Carnevale returned to private practice until accepting her current position at Colorado State University in Dr. Carnevale helps teach undergraduate, graduate, and veterinary students at the Equine Reproduction Laboratory. She also lectures and teaches laboratory sessions for short courses for veterinarians and horse owners. Dr. Carnevale heads the advanced assisted reproductive program at the Equine Reproduction Laboratory, using techniques, such as oocyte (egg) transfer and intracytoplasmic sperm injection (ICSI), to produce offspring from valuable horses that were previously thought to be infertile. Her team also receives ovaries from deceased mares in an effort to produce additional offspring. Research in Dr. Carnevale s laboratory focuses on reproductive aging in the mare and assisted reproduction. Studies have been conducted to examine age-associated changes in the follicles and oocytes of old mares, in an attempt to improve their reproductive performance. Dr. Carnevale s research in assisted reproductive techniques has resulted in many procedures, such as oocyte transfer, ICSI, and oocyte harvesting, that are currently being used in the clinical program. Developments continue in this area to aid in our understanding of equine reproduction and to assist in the production of offspring from valuable horses. Research Interests: Assisted reproductive techniques; Effects of maternal aging on oocyte quality and fertility Selected Honor: Pfizer Animal Health Award for Research Excellence,

28 Colin M. Clay, PhD, Head and Professor of Biomedical Sciences Dr. Colin M. Clay obtained his BS, MS, and PhD degrees at Colorado State University. Working with Drs. Edward Squires and B.W. Pickett at the Stallion Lab (known today as the ERL), Dr. Clay s graduate work provided the equine industry with the seminal studies regarding the effect of season and artificial photoperiod on the stallion. After post-doctoral training at Case Western Reserve University, Dr. Clay returned to a faculty position at CSU and now serves as a Professor and Head of the Department of Biomedical Sciences in the College of Veterinary Medicine and Biomedical Sciences. Dr. Clay teaches in a variety of undergraduate and graduate level physiology and biomedical science courses. He was a mentor and co-director for the Merck-Merial PVM summer research program from Dr. Clay s research program now focuses on the factors that control ovulation in the female. The region of the brain called the hypothalamus, which then signals to the anterior pituitary gland, is the primary source of endocrine hormones that will influence reproduction. The interaction of the hypothalamic peptide termed gonadotropin releasing hormone (GnRH) with specific receptors located on gonadotrope cells of the anterior pituitary gland represents a central point for regulation of reproductive function in all species. His laboratory uses a variety of mouse, sheep, and horse models to define the molecular mechanisms underlying transcriptional regulation of the GnRH receptor gene in an effort to understand the factors that mediate the surge in luteinizing hormone released from the gonadotrope prior to ovulation. Kisspeptin is another small peptide that may be the signal controlling seasonal reproduction in the horse as well as communicating signals for fetal-placental well-being. This work was initiated by Dr. Christianne Magee, who began as veterinary resident/ms student at the ERL, and after becoming board certified in Equine Reproduction by the American College of Theriogenologists also completed a PhD with Dr. Clay. Dr. Magee, in collaboration with other members of the ARBL and ERL, provided the first anatomical and biological evidence for a hypothalamic role for kisspeptin in the mare in a manuscript published in Endocrinology in Dr. Magee and her kisspeptin research were supported by the Preservation of Equine Genetics (PEG) Program at CSU, leading directly to numerous scientific presentations at the Society for Theriogenology, the Society for the Study of Reproduction, and the International Symposium for Equine Reproduction. Importantly, support from the PEG Program provided the preliminary data necessary to obtain a research grant in 2008 from the American Quarter Horse Foundation (AQHF); Dr. Magee was awarded the 2008 Grayson Jockey Club Foundation Storm Cat Award and the 2009 AQHF Young Investigator Research Award. Current work involves elucidating the role of kisspeptin in equine seasonal reproduction and pregnancy, and development of a radioimmunoassay for equine kisspeptin. The group is using non-terminal methods to study the effect of kisspeptin at the level of the hypothalamus in the horse and in the pregnant mare, as well as next-generation sequencing technologies to generate whole transcriptome libraries for equine reproductive tissues. Research Interests: GnRHR promoter regulation; Kisspeptin; Equine seasonal reproduction; Equine fetal-placental well-being; Next-generation sequencing 26

29 Ryan A. Ferris, DVM, Clinical Instructor Dr. Ferris graduated from veterinary school at Washington State University in He then completed an internship in equine surgery, medicine, and reproduction at the Equine Medical Center of Ocala in Dr. Ferris subsequently completed a 2-year Equine Theriogenology residency/master of Science program at Colorado State University through the Department of Clinical Sciences. Ryan became board certified by the American College of Theriogenologists in Dr. Ferris joined the CSU faculty as a Clinical Instructor in the Department of Clinical Sciences in He is primarily involved in the clinical broodmare, foaling, and embryo transfer programs at the Equine Reproduction Laboratory. Ryan also consults on reproduction cases and emergencies at the James L. Voss Veterinary Teaching Hospital. Research Interests: Bacterial and fungal endometritis; Post-mating induced endometritis; Embryo transfer James K. Graham, PhD, Professor of Biomedical Sciences Dr. James Graham is a Professor in the Department of Biomedical Sciences at Colorado State University. He received a BS at the University of Minnesota and PhD at Cornell University in reproductive physiology. Dr. Graham received his post-doctoral training at Michigan State University and Pennsylvania State University before moving to Colorado State University. He is involved in teaching courses at the undergraduate and graduate levels, as well as to veterinary students. His research concentrates on semen cryopreservation and analysis. Research Interests: Gamete physiology and cryobiology; Membrane biochemistry and function Selected Honor: Outstanding Mentor NIH Science Motivation Program,

30 Thomas R. (Tod) Hansen, PhD, Traubert Professor, Department of Biomedical Sciences and Director, Animal Reproduction and Biotechnology Laboratory Dr. Hansen received his BS in Animal Science from CSU in His MS (1984) and PhD (1986) in Reproductive Physiology were earned at Texas A&M University. Following postdoctoral training ( ) in molecular biology, he served as research Assistant Professor at the University of Missouri until 1990 when he joined the faculty as an Assistant Professor in Animal Science and Reproductive Biology at the University of Wyoming. He was promoted to Associate Professor in 1995 and to Professor in In 2005, he joined the faculty at CSU as the Mabel I. and Henry H. Traubert Professor in the Department of Biomedical Sciences and the Director of the ARBL. In 2006 he also started service as Director, Division of Physiology in the Department of Biomedical Sciences. Dr. Hansen contributes to teaching BMS 640 which is a graduate course focused on reproductive physiology and endocrinology. He trains undergraduate honors research, MS, DVM- MS, DVM-PhD, and postdoctoral students. He has authored or co-authored over 200 manuscripts and abstracts. He has published 2 patents and has 3 patent applications currently under review. Research Interests: Maternal uterine and ovarian responses to pregnancy; Mechanisms associated with implantation and placentation; maternal and fetal responses to infection with virus Selected Honors: Sorenson Outstanding Graduate Student Award, 1986 National Institute of Health FIRST Award, 1994 Mabel I. and Henry H. Traubert Professorship, 2005 Chair, USDA-AFRI Reproduction in Animals Grant Panel, 2007 Donald Henry Barron Lecturer,

31 Patrick M. McCue, DVM, PhD, Diplomate, American College of Theriogenologists, Professor of Clinical Sciences, Iron Rose Ranch Chair in Equine Reproduction Dr. McCue graduated from veterinary school at the University of California-Davis in He subsequently completed an internship in large animal medicine and surgery at the University of Pennsylvania and a residency in equine reproduction at the University of California-Davis. He received a PhD in comparative pathology from UC-Davis in Dr. McCue joined the faculty at Colorado State University as an Assistant Professor in 1994, was promoted to Associate Professor in 2000, and to full Professor in In 2010, Dr. McCue was named the Iron Rose Ranch Chair in Equine Reproduction. He is the author or co-author of over 200 publications, including journal articles, research abstracts, textbook chapters, and proceedings. In addition, he writes a monthly column called The Breeding Shed for the American Quarter Horse Journal. Dr. McCue was the recipient of the Carl J. Norden Distinguished Teacher Award and the AAEP Faculty Award for excellence in teaching while at CSU. He is actively involved in teaching veterinary students, providing continuing education for horse owners and veterinarians, and conducting research in equine reproduction. As Senior Veterinarian in Charge of ERL Clinics, Dr. McCue coordinates the Clinical Broodmare, Foaling, and Embryo Transfer Services. He also attends to equine reproduction cases at the James L. Voss Veterinary Teaching Hospital at Colorado State University. Research Interests: Reproductive pathology; Reproductive endocrinology; Hormone therapy; Embryo transfer Selected Honors: Diplomate, American College of Theriogenologists Norden Distinguished Teacher Award, 1996 Faculty Clinician Award, American Association of Equine Practitioners, 1996, 2003 Board of Directors, American Association of Equine Practitioners,

32 Terry M. Nett, PhD, Professor of Biomedical Sciences Dr. Nett earned a PhD in reproductive physiology from Washington State University in For his doctoral dissertation, he established assay systems to measure estrogens and progestins in mares and applied these assays to characterize secretion of these hormones during the estrous cycle, pregnancy, and the postpartum period. Dr. Nett served two years as a postdoctoral fellow in the Reproductive Endocrinology Program at Colorado State University beginning in 1972, then joined the faculty of the Animal Reproduction and Biotechnology Laboratory in 1974 where he continues his research today. Although not his primary focus, Dr. Nett has retained a strong interest in equine reproduction and has continued to perform research in horses throughout his career. He has published over 200 articles in refereed journals and contributed more than 30 chapters to text books. He served as Associate Dean for Research and Graduate Education in the College of Veterinary Medicine and Biomedical Sciences from Dr. Nett has taught various courses in reproductive physiology throughout his career and has trained 14 PhD students. His research interests include studying the mechanisms controlling seasonal reproduction, endocrine changes throughout the estrous cycle and pregnancy, and developing novel methods to measure reproductive hormones. He also is developing and evaluating methods for nonsurgical sterilization and/or contraception that may be applied to horses. Research Interests: Reproductive endocrinology of mares and stallions; Development of new techniques to assess/monitor changes in pregnancy status and/or the need for hormone therapy; Hormonal basis for testicular degeneration in stallions Selected Honors: Animal Physiology and Endocrinology Award, American Society of Animal Science, 1989 President, Society for the Study of Reproduction, Distinguished Alumnus Award, Washington State University, 1998 Distinguished Service Award, Society for the Study of Reproduction, 2001 Carl G. Hartman Award, Society for the Study of Reproduction, 2009 Oliver Pennock Distinguished Service Award, Colorado State University,

33 George E. Seidel, Jr., PhD, University Distinguished Professor of Biomedical Sciences George E. Seidel, Jr. was raised on a dairy farm in Pennsylvania. He received a BS degree from Penn State in dairy science in 1965, and MS (1968) and PhD (1970) degrees from Cornell. His Master s thesis concerned methodology of semen collection from bulls and biochemistry of semen. His PhD thesis concerned the endocrinology of superovulation of prepuberal calves and culture and transfer of the resulting embryos. He went on to Harvard Medical School to study electron microscopy. For the past 40 years, Dr. Seidel has been at the ARBL at Colorado State University. His current title is University Distinguished Professor. Initially, work in Colorado was primarily on reproductive physiology of bulls and stallions. In 1973, the Embryo Transfer Laboratory was established. Farmers brought their valuable donor cows to this laboratory for superovulation and recovery of embryos, which then were placed into the uteri of less valuable cows for gestation. Over the next decade, more than 6000 bovine embryos were transferred in this manner. Fees for these services funded the bulk of the teaching and research of the Laboratory. Techniques such as nonsurgical recovery and transfer of bovine and equine embryos and cryopreservation of embryos were developed and refined. The Laboratory became known for developing a simple, reliable procedure for bisecting embryos to produce identical twins in a variety of species, and for developing reliable methods for cryopreservation of embryos. Dr. Seidel also has helped to optimize many of these biotechnologies for equine reproduction. In 1978, Dr. Seidel spent a sabbatical leave at Yale and in 1986, a second sabbatical leave at the Whitehead Institute at MIT. Current research projects include in vitro oocyte maturation and fertilization, cryopreservation of oocytes and embryos, genetic control of early embryonic development, and sexing semen. The latter work resulted in founding the company, XY, Inc., which was owned in part by the Colorado State University Foundation, but now is an independent company in Texas. Research Interests: Reproductive physiology; Superovulation and embryo transfer; In vitro fertilization and culture of ova; Regulation of development of preimplantation embryos; Micromanipulation and cryopreservation of bovine and equine embryos; Sexed semen Selected Honors: Andrew G. Clark Award for Research and Creativity, Colorado State University, 1982 Alexander von Humboldt Award, 1983 Young Animal Scientist Award, Western Sect. American Soc. Animal Science, 1983 National Association of Animal Breeders Research Award, 1983 Upjohn Physiology Award, American Dairy Science Association, 1986 Governor s Award for Science and Technology, State of Colorado, 1986 Member, National Academy of Sciences, 1992 University Distinguished Professor, Colorado State University, 1993 Distinguished Service Award, International Embryo Transfer Society, 2001 Armsby Society Award, Pennsylvania State University, 2003 Pioneer Award, International Embryo Transfer Society, 2008 Researcher of the Year, Colorado State University Foundation, 2008 Animal Physiology and Endocrinology Award, American Society of Animal Science,

34 D.N. Rao Veeramachaneni, BVSc, MScVet, PhD, Professor of Biomedical Sciences D. N. Rao Veeramachaneni is a professor in the Department of Biomedical Sciences in the College of Veterinary Medicine and Biomedical Sciences at Colorado State University. He received his professional veterinary medical education from AP Agricultural University, India and PhD from University of Illinois, Urbana-Champaign. Research in Veeramachaneni s laboratory involves study of the impact of environmental pollutants on male reproduction using a variety of laboratory and wild animal models to address the question if deteriorating reproductive health in males is really idiopathic, as male infertility often is labeled, or a consequence of exposure to common chemical contaminants. He applies this knowledge in seminal evaluations of a variety of domesticated animals including stallions ( colostate.edu/bms/arbl/arbl_spermmorph.htm). His research has been featured on the cover pages of American Journal of Andrology and Toxicological Sciences. He has been invited to speak at several international forums and continuing education sessions at professional society meetings including International Congress on Animal Reproduction, Stockholm, Sweden; European Environmental Agency, Copenhagen, Denmark; Copenhagen Workshop on Environment, Reproductive Health and Fertility; VII Copenhagen Workshop on CIS and Testicular Cancer; European Union-Sponsored Credo Workshop on the Ecological Relevance of Chemically Induced Endocrine Disruption in Wildlife, Exeter, England; American Society of Andrology and Society of Toxicological Pathology. He has been serving on the editorial board of Reproduction (The Journal of the Society for Reproduction and Fertility, UK) since

35 Collaborating Scientists David J. Denniston, MS, PhD -- Associate Professor of Animal Sciences Research expertise: Equine reproductive physiology Christianne Magee, DVM, MS, PhD -- Postdoctoral Fellow Research expertise: Molecular endocrinology; Neuroendocrinology; Equine embryo transfer; Next-generation sequencing 33

36 Areas of Expertise of Core Personnel Gerrit Bouma, PhD Molecular and genetic control of fetal gonadal cell differentiation and adult reproductive function Jason Bruemmer, PhD Cryopreservation of stallion semen; Sperm membrane integrity; Maternal recognition of pregnancy in the mare; Luteal tissue formation, function, and demise Elaine Carnevale, DVM, PhD Assisted reproductive technologies; Follicular and oocyte interactions; Aging in the mare Colin Clay, PhD Molecular endocrinology Ryan Ferris, DVM, PhD Bacterial and fungal endometritis; Post-mating induced endometritis; Embryo transfer James Graham, PhD Semen freezing; Evaluation of semen Thomas Hansen, PhD Maternal recognition of pregnancy, including how the embryo signals to the mother; Maternal and fetal immune response to viral infection during pregnancy; Maternal adaptation to stress during pregnancy Pat McCue, DVM, PhD Equine embryo transfer; Hormone therapy; Ovarian pathology; Problem mares; Foaling and care of the newborn foal Terry Nett, PhD Evaluation of how changes in production of reproductive hormones impact the efficiency of reproduction in stallions and mares George Seidel, Jr., PhD Superovulation and embryo transfer; Cryopreservation of oocytes and embryos; In vitro oocyte maturation, fertilization, and culture of embryos; Micromanipulation, including splitting embryos and sperm injection; Sexing semen D.N. Rao Veeramachaneni, BVSc, MScVet, PhD Andrology; Reproductive toxicology; Impact of environmental pollutants on male reproduction 34

37 Staff Veterinarian Dr. Judy Merriott, DVM Dr. Merriott received her undergraduate degree from the University of Michigan. After working several years for a large corporation, she enrolled in veterinary school at Colorado State University and graduated in She was in private veterinary practice in Colorado for 8 years before joining the ERL team. Dr. Merriott is a staff veterinarian in the Assisted Reproductive Technology (ART) program where she manages the recipient and research mare herds, assists with collection of oocytes from donor mares, and provides instruction to veterinary students. 35

38 Research Associates JoAnne Stokes JoAnne received a Bachelor of Science Degree in Dairy Science from the Ohio State University. She worked for 14 years for private companies in Pennsylvania and Texas that provided services in bovine embryo transfer and in vitro fertilization. In 2003, JoAnne joined the ERL team to provide expertise in assisted reproductive technologies (ART). She works primarily with Dr. Elaine Carnevale and is responsible for care and nurturing of oocytes collected from valuable mares and performs intracytoplasmic sperm injection (ICSI) to fertilize the oocytes. JoAnne assists faculty and graduate students in a myriad of research projects and is involved in continuing education courses provided to horse owners and veterinarians at the Equine Reproduction Laboratory. Jillian Bishop Paula Moffett Jillian graduated with a Bachelor of Science degree from Kansas State University before completing a 6-month internship at Babcock Ranch in Texas. She then completed a one-year Equine Breeding Farm Internship at the Equine Reproduction Laboratory. Jillian now helps Drs. Pat McCue and Ryan Ferris in the Mare Program and coordinates mare and foal health care, and assists with breeding management and embryo transfer services. She also manages a clinical microbiology service and is actively involved in teaching CSU students, as well participating in the continuing education programs offered to horse owners and veterinarians at the Equine Reproduction Laboratory. Paula received a Bachelor s Degree in Agricultural Science and Natural Resources from Oklahoma State University. She joined the Equine Reproduction Laboratory in 2001 as a volunteer. She subsequently started a Master s program in Biomedical Sciences and eventually became a full-time Research Associate. Paula assists Dr. Jason Bruemmer in the Stallion Program and coordinates stallion care, semen collection and evaluation, and semen freezing services. She also participates in teaching undergraduate, graduate and PVM students as well as research projects involving stallions and/or semen. In addition, Paula is actively involved in the horse owner and veterinary continuing education courses provided through the Equine Reproduction Laboratory. 36

39 Current DVM Residents David B. Scofield, DVM, Equine Reproduction Resident Dr. Scofield grew up on the eastern shore of Maryland and in northern Delaware. He received a Bachelor of Science degree in biology at Wake Forest University in North Carolina in He subsequently enrolled in veterinary school at Oklahoma State University and received his DVM in Dr. Scofield completed an internship in equine medicine, surgery, and reproduction at Weatherford Equine Medical Center in Weatherford, Texas in He is currently in the first year of a 2-year Equine Reproduction residency at Colorado State University. He works primarily at the Equine Reproduction Laboratory and also provides equine reproduction services at the Veterinary Teaching Hospital. Research Interests: Pharmacokinetics of antimicrobial drugs; Embryo transfer; Hormone therapy Alicia R.G. Lindholm, DVM, Equine Reproduction Resident Dr. Lindholm was born and raised in Chelan, Washington, on the apple orchard her family owns and operates. She attended Washington State University and graduated with a Bachelor s degree in zoology in During her undergraduate studies she spent a semester studying abroad at James Cook University, Townsville, Queensland, Australia. Dr. Lindholm graduated from veterinary school at Washington State University in She completed an ambulatory internship at Rood and Riddle Equine Hospital in Lexington, Kentucky. In 2009, she began a 2-year combined Equine Theriogenology residency/master of Science degree at Colorado State University. A majority of her work is at the Equine Reproduction Laboratory working in the Clinical Broodmare, Foaling, and Embryo Transfer service. In addition, she consults on equine reproduction cases at the Veterinary Teaching Hospital. She will complete the program in June Research Interests: Ovarian cancer biomarkers; Uterine pathology; Embryo transfer; Hormone therapy 37

40 Graduate Students Ashley Cameron MS Student Alyssa Coville MS Student Joanna Hergenreder MS Student Elena Ruggeri PhD Student Juliano Silveira PhD Student ERL graduate students, left to right: Juliano Silveira, Elena Ruggeri, Alyssa Coville, Alicia Lindholm (DVM Resident), Joanna Hergenreder, David Scofield (DVM Resident), Ashley Cameron Postdoctoral student Dawn Sessions Interns Amy Anderson (left) Alexandria Hayes (right) 38

41 Synopsis of Program Overview The Equine Reproduction program at Colorado State University has a long history of developing reproductive techniques for the equine industry. Initially started in 1967 by Drs. B.W. Pickett and J.L. Voss, this program has evolved into a nationally and internationally renowned research enterprise. Over the past 40 years, techniques such as collection of semen and artificial insemination, recovery and transfer of equine embryos, shipping of cooled semen, shipping of cooled embryos, and freezing of oocytes, embryos, and semen have been developed and are now routinely used in the equine industry. More recent work includes assisted reproductive techniques, sexing of semen, intracytoplasmic sperm injection (ICSI), and oocyte transfer. Facilities have been built to accommodate the increased number of mares and stallions housed at the Laboratory. Typically several hundred mares are housed at this facility during the breeding season, as well as 25 stallions. The large research population of mares and stallions has provided a basis for teaching undergraduate, graduate, and veterinary students, as well as farm managers and veterinarians. The pharmaceutical industry has taken advantage of the fact that Colorado State University maintains a large number of horses, and numerous trials have been conducted in conjunction with the Equine Reproduction Laboratory, in which various drugs were evaluated to determine their effect on reproduction of mares and stallions. Transfer of technology from a research setting to the breeding farm always has been a high priority of our program. Since 1970, continuing education courses on equine reproductive management have been offered. Currently each year there are 7 to 10 continuing education programs available to the veterinarian and breeder concerning all aspects of reproduction, including reproductive management of mares and stallions, cooled semen, frozen semen, ultrasonography, and embryo transfer. The recent addition of laboratory space has allowed us to expand our continuing education programs as well as clinical services to the equine industry. This intimate contact with the horse industry has been extremely beneficial to our research program. Feedback from veterinarians and horsemen has helped to identify problems that are most pressing to the equine industry. 39

42 Facilities We now have a 4,000-square-foot office/laboratory complex that is used for teaching, research, and service, as well as the 22,000-square-foot Animal Reproduction and Biotechnology Laboratory, adjacent to the Equine Reproduction Laboratory, that houses 13 additional faculty members, graduate students and postdocs, and laboratory space that can be used for hormonal assay, cell culture, anatomical evaluation of sperm and facilities for in vitro embryo production. We have separate labs for embryo transfer, oocyte transfer, semen freezing, and sperm injection. Twenty stocks, divided among 5 locations, are available for mare examinations. A new building adjacent to the ERL contains a classroom, graduate student work stations, and a two-bedroom apartment for the interns. 40

43 Research Firsts The Equine Reproduction Laboratory is noted for having numerous firsts in the U.S., many of these are listed below. In addition, research in this laboratory has resulted in development of several drugs that now are used by veterinarians and breeders. These drugs include hormone preparations such as Regu-Mate, a hormone used for ovulation control, Ovuplant, and efsh used for stimulating follicular development. Other significant research findings include the mechanism of seasonal reproduction in both the mare and stallion. 1) Techniques for nonsurgical recovery and transfer of equine embryos (1980) 2) Freezing embryos (1985) The Equine Reproduction Laboratory was the first to produce live foals from frozen-thawed embryos in the U.S. 3) Identical twins (1985) Colorado State produced the first live foals from the bisection of a day-6 equine embryo. The resulting halves were then transferred into 2 separate mares who produced healthy male foals. 4) Shipping of cooled embryos (1988) This technique allows embryos to be collected on the farm and shipped to Colorado State for transfer into recipient mares. 5) Test tube horse (1996) Colorado State was the first to produce a foal by injecting a single sperm into an unfertilized egg. 6) Oocyte transfer (1999) This technique involves collection of an egg from a valuable donor mare and transferring that egg into a recipient, then breeding the recipient. 7) First foals from insemination of a mare with sexed semen (1999) In this technique, semen is sorted into X- and Y-chromosome-bearing spermatozoa. 8) Oocyte freezing (2000) Two mares became pregnant from transfer of frozen-thawed oocytes (eggs) and foaled in ) Gamete intrafallopian tube (GIFT) transfer (2000) This technique involves collecting an egg from a valuable donor mare and transferring that egg and sperm into a recipient. 10) Birth of first foal from ovaries taken from a dead mare (2002) 11) Injection of frozen/thawed sperm (2003) Commercial pregnancies were produced from injection of frozen/thawed sperm into oocytes from client mares. 12) Birth of a foal from use of frozen/thawed epididymal sperm (2005) 13) Successful freezing of in vitro-produced, 2- or 3-day-old embryos (2006) 41

44 Undergraduate Education The Equine Reproduction Laboratory offers a unique teaching and learning opportunity for undergraduate students because of the clinical reproduction service and the number of stallions and mares maintained at the facility. Students in the Equine Sciences, Animal Sciences and other undergraduate programs may take courses that have lecture or laboratory sessions at the ERL. In addition, ERL faculty members provide additional opportunities for undergraduate students in the form of independent study, honors research projects, work study, and other venues. ERL faculty coordinate or assist in teaching the following CSU undergraduate courses (annual enrollment for is shown in parentheses): Equine Behavior (49) Equine Management (42) Equine Nutrition (80) Equine Practicum (35) Equine Production and Industry (70) Equine Reproductive Management (90) Foaling Management (95) Introduction to Equine Sciences (140) In addition, Dr. Bruemmer works closely with the CSU Polo Team and Dr. Denniston not only coaches the CSU Horse Judging Team but also serves as the youth horse extension for the state of Colorado and, as such, coordinates the Colorado 4-H horse program. 42

45 PVM Teaching The Equine Reproduction Laboratory is the largest comprehensive teaching, research, and clinical service program in equine reproduction located at any College of Veterinary Medicine in the United States. The faculty and staff of the ERL are an integral part of the professional veterinary medicine (PVM) teaching program at Colorado State University. Faculty of the ERL teach the core course in Theriogenology (VM 744) to all 2nd-year PVM students and the equine reproduction component of the core Clinical Sciences III (VM 749) to the 3rd-year PVM students. In addition, elective courses in Foaling Management (ANEQ 445) and the reproduction section of Equine Medicine and Surgery I (VM 763) are taught by ERL faculty. Clinical education of PVM students also is facilitated by elective rotations at the Equine Reproduction Laboratory in the Junior and Senior Practicum courses. In these rotations, PVM students are provided the opportunity for hands-on clinical training concerning the mare, foal, and stallion. The large population of resident horses at the ERL provides additional opportunities for PVM students on the Equine Field Service, Independent Study electives, student research programs, and wet labs sponsored by clubs such as the Student Chapter of the American Association of Equine Practitioners and the Student Chapter of the Society for Theriogenology. Equine Theriogenology Residency Program The Department of Clinical Sciences at Colorado State University offers a 2-year clinical residency in Equine Theriogenology. The goals of the program are to provide the candidate with an opportunity to develop significant clinical expertise in all areas of equine reproduction and to prepare residents for a future career in academic or specialty private practice. Residents are enrolled in a non-thesis Master s (Plan B) degree program in the Department of Clinical Sciences. Residents are expected to be active participants in ongoing programs in clinical equine reproduction, teaching, continuing education, and research. The residency program is based at the Equine Reproduction Laboratory on the Foothills Campus of Colorado State University. In addition, residents provide Equine Theriogenology support to the Veterinary Teaching Hospital. Past DVM Interns and Residents Name Position Years Dr. Racquel Rodeheaver Intern 1997 Dr. Robert Foglia Resident Dr. Kory Niswender Resident Dr. Jason Hudson Resident Dr. Nichole Logan Resident Dr. Christianne Magee Resident Dr. Erica Gee Resident Dr. Catherine DeLuca Resident Dr. Ryan Ferris Resident Dr. Alica Lindholm Resident

46 Intern Program The ERL has an ongoing Equine Breeding Farm Internship program to provide advanced training in equine breeding farm procedures and management. Two interns, chosen through an application and interview process, are hosted each year. Successful candidates for an intern position have recently completed a Bachelor s degree in equine science, animal science, or a related discipline, and have a demonstrated desire to pursue a career in the equine breeding industry. ERL interns are provided a salary and housing at the Equine Reproduction Laboratory, along with a certificate upon successful completion of the one-year program and job placement assistance. Reproductive training of interns includes: Stallion Stallion handling Stallion management Semen collection Preparation of cooled-transported semen Semen evaluation Semen freezing principles Mare Hormone therapy Reproductive management of mares Artificial insemination Care of the pregnant mare and foaling Embryo transfer (assist) Care of the newborn foal Oocyte transfer (assist) Herd health management Job responsibilities include: Assist with the Clinical Equine Reproduction Program (horse handling, herd health management, record keeping, etc) Assist with general laboratory functions (including autoclaving, equipment cleaning, laundry, reception, etc) Participate in foal watch and foaling (provide back-up and assistance to students in the Foaling Management class) After-hours help with artificial insemination, embryo transfer, and horse arrivals/departures (rotate with or provide back-up to graduate students) Weekend on call (after-hours availability every other weekend for each intern) Night barn checks (i.e. monitor client-owned horses) Evening security check (i.e. gates shut and locked; facility doors locked) 44

47 Awards E.M. Carnevale Pfizer Animal Health Award for Research Excellence, 2008 D.J. Denniston Spurring Them to Excellence Teaching Award, Department of Animal Sciences, Colorado State University, 2009 P.M. McCue Iron Rose Ranch Chair in Equine Reproduction, 2007-present Nominated by MSNBC (NBC Nightly News) for 2010 Butterfly Award for Best Hometown Hero, 2010 G.E. Seidel, Jr. National Academy of Sciences, 1992-present University Distinguished Professor, Colorado State University, 1993-present Pioneer Award, International Embryo Transfer Society, 2008 Researcher of the Year Award, Colorado State University Research Foundation, 2008 Animal Physiology and Endocrinology Award, American Society of Animal Science, 2008 E.L. Squires Induction to University of Kentucky Equine Research Hall of Fame, 2007 Editorial Boards J.E. Bruemmer Journal of Animal Science, E.M. Carnevale Equine Veterinary Education, Assistant Editor, 2007-present C.M. Clay Biology of Reproduction, 2004-present P.M. McCue Journal of Equine Veterinary Science, 1998-present 45

48 G.E. Seidel, Jr. Reproduction in Domestic Animals, 2002-present Animal Reproduction, 2004-present Encyclopedia of Biotechnology in Agriculture and Food (Editorial Advisory Board), 2005-present Annual Review of Animal and Veterinary Sciences, 2011-present E.L. Squires Journal of Equine Veterinary Science (Editor), 2006-present Theriogenology, 1998-present D.N.R. Veeramachaneni Reproduction (SSF, UK), 2002-present Grant Proposal Review Boards J.E. Bruemmer Scientific Review Board, American Quarter Horse Association, 2008-present USDA, 2010 E.M. Carnevale CVMBS College Research Council, C.M. Clay NIH ICER Study Section, Ad Hoc member, 2008 J.K. Graham NSF, Ad Hoc Reviewer, 2008 G.E. Seidel, Jr. NIH Study Section on Cryobiology, 2008 NSERC (Canada), Ad Hoc Reviewer for Centers of Excellence Program, 2008 Dutch Government, Ad Hoc Reviewer for National Program,

49 Research Support The following individuals and companies have provided funds or other contributions toward ongoing research projects through the Equine Reproduction Laboratory. Mr. Jason Abraham American Quarter Horse Foundation Mr. Thomas H. Bailey Boisvert Farms, LLC Mrs. Barbara B. Chur Ms. Carri A. Copas Mr. Glenn Drake Dupree, Inc. Ms. Heather A. Hall Mr. Howard F. Henneman & Mrs. Barbara L. Henneman Hilton Inn Fort Collins Hole in the Wallet Farm Mr. Dave Huber & Mrs. Rhonda L. Huber Ms. Cecilia Hylton Ms. Dana Leigh Jones Mr. Harry Leatherwood Mr. Alexander Libfeld Mr. Christopher F. Long Marvine Ranch, LLC Mr. David E. McGlothlin Merial Mr. David G. Novat Partnar Animal Health Peterson & Smith Equine Hospital, LLC Eric S. Peterson, DVM & Kimberly M. Peterson, DVM Platinum Performance, Inc. Ms. Amanda B. Purdin Reproductive Management Rocky Mountain LAE Inc. Rodeo News Sandpoint Animal Medical Care Mr. Anthony F. Sears & Mrs. Merideth M. Sears Mrs. Carolyn D. Smith Mr. M. Wade Smith Mr. Peter D. Stent & Mrs. Nora Stent Mrs. Kathryn G. Stone & Mr. Fred W. Stone Sturgis Veterinary Hospital, P.C. The Walton Family Foundation, Inc. Ward & Ward Industrial Plaza Winegardner & Hammons, Inc. Mr. Jon Winkelried & Mrs. Abby Winkelried Major Donors Cecilia Hylton -- The Cecil & Irene Hylton Family Foundation Nora and Peter Stent Alice Walton -- Walton Family Foundation Jon and Abby Winkelried -- Marvine Ranch 47

50 Equine Scholarship Funding Through ARBL and ERL Abney Foundation Scholarship James N. Dupree Scholarship E.H. Honnen Scholarship Pattridge Family Scholarship Dr. Dean Pavillard Scholarship France Stone Scholarship Ray T. Walker Memorial Scholarship 48

51 Preservation of Equine Genetics (PEG) Program The Preservation of Equine Genetics (PEG) Program was developed approximately 10 years ago to allow us to focus our research on assisted reproductive technologies. Tremendous progress has been made toward the goals that were established for this program. Breeders have been very supportive in providing funds for development of reproductive techniques, such as semen freezing, oocyte and embryo freezing, superovulation, assessment of sperm damage, intracytoplasmic sperm injection, and oocyte collection and transfer. A very simplified technique has been developed for freezing equine embryos. Embryos are placed sequentially into three different solutions containing cryoprotectants, loaded into a 0.25-ml straw, and frozen quickly in liquid nitrogen. Progress also has been made in developing new methods for freezing semen and for assessing sperm damage after cooling and freezing. Furthermore, simplified procedures have been developed for insemination of mares with frozen semen, which makes the use of frozen semen much more practical. Through initial studies conducted in the PEG Program, we were able to solicit the help of a pharmaceutical company, Bioniche Animal Health, in developing a product to be used for induction of multiple ovulations in mares (efsh). This product has been demonstrated to safely stimulate multiple ovulations in cycling mares. Embryo recovery from superovulated mares averages approximately 2 embryos per flush, compared to 0.7 embryos per flush in untreated mares. The procedure for collection and transfer of oocytes now has become commonplace and is a major part of our commercial clinical program. Approximately 150 oocytes are collected and transferred each year from very valuable, subfertile mares. Techniques for assisting the subfertile stallion also have been developed, such as intracytoplasmic sperm injection and low-dose insemination. Recently, we were able to produce foals from very valuable mares in which oocytes were collected and then injected with sperm from an extremely valuable, subfertile stallion. This procedure is now being offered as a clinical service. Our plans are to continue to develop assisted reproductive techniques that can be used for valuable older mares and stallions. With the assistance of faculty in the Animal Reproduction and Biotechnology Laboratory, we plan to develop a strong program in molecular biology concerning maturation of equine oocytes, control of the corpus luteum and identification of embryonic factors responsible for recognition of pregnancy. The preliminary data obtained through the PEG Program can be used to acquire funding from federal agencies, such as the National Institutes of Health. We are fortunate to have a great group of faculty members, postdoctoral students, and graduate students to continue these very significant equine reproduction projects. Furthermore, we are extremely pleased to have good funding from equine breeders and the pharmaceutical industry to allow us to continue to conduct research. The significance of research within the PEG Program includes: 1) Preserve genetic material (eggs, embryos, sperm). 2) Improve fertility of frozen semen such that the genetic pool can be increased and the cost of breeding with frozen semen can be decreased. 3) Assess fertility of frozen semen prior to breeding using new laboratory techniques. 4) Understand the factors involved in follicular development. 49

52 5) Lower the cost of embryo transfer and improve pregnancy rates of mares by using superovulation. 6) Obtain pregnancies from older, valuable mares in which conventional embryo transfer has been unsuccessful. 7) Determine the causes of reduced fertility in mares. 8) Obtain pregnancies from stallions which are subfertile by using such techniques as intracytoplasmic sperm injection, in vitro fertilization, and oviductal insemination. 9) Allow sex selection of embryos. 10) Control sex of offspring by selecting X- or Y-chromosome-bearing sperm. 11) Improve the success of freezing day-7 or -8 embryos. The PEG Program is a joint effort involving faculty in the Departments of Biomedical Sciences, Animal Sciences, and Clinical Sciences. These faculty members include: Dr. Gerrit Bouma Department of Biomedical Sciences Dr. Jason Bruemmer Department of Animal Sciences Dr. Elaine Carnevale Department of Biomedical Sciences Dr. Colin Clay Department of Biomedical Sciences Dr. David Denniston Department of Animal Sciences Dr. Jim Graham Department of Biomedical Sciences Dr. Pat McCue Department of Clinical Sciences Dr. George Seidel Department of Biomedical Sciences Dr. Ed Squires Department of Biomedical Sciences Dr. Quinton Winger Department of Biomedical Sciences We feel that Colorado State University has the faculty, expertise and facilities to develop new and exciting techniques that will allow for propagation of superior equine genetics and preservation of equine genetics for the future. 50

53 Funded Research Projects Title Researchers Sponsor Dates Amount Cell-to-cell communication PI: G. Bouma PEG 7/09-6/10 32,080 within the equine follicle: Role Co-PI: E. Carnevale for microvesicle-mediated transfer of signaling molecules Gene expression in the equine Co-PI: J. Bruemmer, PEG 2/06-1/07 26,500 endometrium during maternal T.R. Hansen recognition of pregnancy Identification of maternal PI: J. Bruemmer PEG 7/07-6/08 31,618 recognition in the mare by endometrial gene expression Key to equine maternal PI: J. Bruemmer CRC 7/08-6/09 recognition: Apical versus basolateral secretion of endometrial oxytocin Equine in silico library database PI: J. Bruemmer PEG 7/09-6/10 50,000 development Co-PI: G. Bouma, C. Magee, C. Clay The role of prostaglandin PI: J. Bruemmer AQHF ,440 transporter in equine maternal recognition of pregnancy Development of assisted Co-PI: E. Carnevale, Hylton Fdn 7/05-6/08 250,000 reproductive techniques for E. Squires mares and stallions Effects of aging and efsh on PI: E. Carnevale PEG 5/06-4/07 45,000 initiation of oocyte maturation and oocyte developmental competence in the mare Transfer of in vitro produced PI: E. Carnevale P. Stent 6/06-8/07 embryos Comparison of laparoscopic PI: E. Carnevale CRC 7/06-6/07 19,956 techniques for oocyte transfer in the mare Differences in arrest and Co-PI: E. Carnevale, PEG 4/07-10/08 50,000 resumption of oocyte maturation E. Squires in young versus old mares 51

54 Title Researchers Sponsor Dates Amount Age-associated alterations in PI: E. Carnevale PEG 7/07-6/08 45,000 fertilization and blood flow Advanced age alters meiotic PI: E. Carnevale CRC 7/07-6/08 20,000 and fertilization competency of oocytes in mares Use of sexed semen to produce Co-PI: E. Carnevale, XY, Inc. 9/07-9/08 6,918 embryos in vitro E.L. Squires Effect of omega 3 fatty acid PI: E. Carnevale Platinum ,000 supplementation on systemic Performance product inflammation donation Improving fertility through PI: E. Carnevale Hylton Fdn ,000 assisted reproduction Role of microrna in equine PI: E. Carnevale PEG 2/09-1/10 oocyte maturation and competence Analysis of oviductal and uterine Co-PI: E. Carnevale, PEG 2/09-1/10 fluid collected in situ from mares P. McCue Pregnancy rates of equine PI: E. Carnevale PEG 2/09-1/10 embryos biopsied for genetic testing Direct molecular analysis of the Co-PI: E. Carnevale USDA ,370 equine cumulus-oocyte complex PI: P. Kayser using imaging mass spectrometry Vitrification of equine oocytes PI: E. Carnevale PEG 7/09-6/10 13,300 after in vivo or in vitro Co-PI: P. McCue, maturation G. Seidel Effect of aging and oocyte PI: E. Carnevale CRC 7/09-6/10 maturity on mrna content of maternal effect genes in the equine oocyte Extending equine fertility PI: E. Carnevale Hylton Fdn ,000 Evaluation of kisspeptin and its PI: C. Clay PEG 7/06-6/07 34,550 effect on the hypothalamicpituitary-gonadal axis in the mare Evaluation of kisspeptide in PI: C. Clay PEG 7/07-6/08 40,000 the mare Evaluation of Kisspeptin in PI: C. Clay AQHA 10/07-10/08 42,000 the mare 52

55 Title Researchers Sponsor Dates Amount Evaluation of Kisspeptin and PI: C. Clay AQHA 10/07-9/08 7,642 its role in the hypothalamicpituitary-gonadal axis Mapping the neuroendocrine PI: C. Clay PEG 7/09-6/10 30,000 control of reproductive function Co-PI: C. Magee, in the mare J. Bruemmer Determining the hierarchy of PI: C. Clay PEG 7/09-6/10 35,000 KiSS signaling in the mare Co-PI: C. Magee, J. Bruemmer Identifying stallion to stallion PI: D. Denniston PEG 7/09-6/10 8,450 differences that affect the ability Co-PI: J. Graham, of spermatozoa to survive J. Bruemmer, cryopreservation G. Seidel Development of a real time PCR PI: R. Ferris, Center for ,000 assay for the detection of P. McCue Companion pathogenic fungi in the uterus Animal Studies of the mare Development of a PCR assay PI: R. Ferris, Center for ,000 to predict genetic antimicrobial P. McCue Companion susceptibility in pathogenic fungi Animal Studies from the uterus of the mare Development of oral PI: J. Graham USDA- 9/04-9/09 199,000 contraceptives for the purpose APHIS of reducing reproduction in the wild pig Cryopreservation of sex-sorted PI: J. Graham XY, Inc. 9/05-8/07 41,826 stallion spermatozoa Improving capacitation of PI: J. Graham PEG 7/06-6/07 24,200 stallion sperm to enhance in vitro fertilization Improving stallion sperm PI: J. Graham PEG 7/07-6/08 21,500 capacitation to enhance in vitro fertilization II Development of new PI: J. Graham ARS 9/07-8/08 cryodiluents for stallion sperm Regulating gonadotropin PI: J. Graham CRC 7/08-6/09 releasing hormone in pigs GIFT Co-PI: J. Graham, PEG 7/08-6/09 30,000 E. Squires 53

56 Title Researchers Sponsor Dates Amount Capacitating stallion sperm PI: J. Graham PEG 7/09-6/10 28,002 to enhance in vitro fertilization Co-PI: E. Squires, G. Seidel Changing sperm membranes to PI: J. Graham CRC 7/09-6/10 enhance cryopreservation Gene expression in the equine PI: T. Hansen PEG 1/07-1/09 endometrium during maternal recognition Evaluation of kisspeptin in the PI: C. Magee Grayson equine -- Storm Cat Career Jockey Club Research Development Award Kisspeptin mediated regulation PI: C. Magee AQHA 10/09-9/10 of the equine hypothalamic pituitary gonadal axis Refining the techniques of PI: P. McCue PEG 7/06-6/07 50,600 using equine follicle-stimulating Co-I: C. Magee, hormone for superovulation E. Squires Characterization and prevention PI: P. McCue PEG 7/07-6/08 23,620 of luteal regression following Co-PI: E. Squires non-surgical transfer of equine embryos Efficacy of recombinant equine PI: P. McCue PEG 7/07-6/08 30,000 follicle stimulating hormone on Co-I: E. Squires superovulation in mares Evaluation of a Brix PI: P. McCue Schering ,400 Refractometer for determination Co-I: P. Dinsmore Plough of bovine colostrum quality Efficacy of medroxyprogesterone PI: P. McCue US Equest ,500 in suppression of estrus in mares Co-I: E. Gee Fdn Superovulation of mares with Co-PI: P. McCue, PEG 7/08-6/09 50,000 pfsh E. Squires MicroRNA profile of mares PI: P. McCue PEG 7/09-6/10 15,985 with granulosa cell tumors Pregnancy rates for embryos PI: G. Seidel PEG 3/09-12/09 18,221 biopsied for genetic testing Co-PI: P. McCue, E. Carnevale, G. Bouma 54

57 Title Researchers Sponsor Dates Amount Optimizing biopsy techniques PI: G. Seidel PEG 4/10-3/11 22,090 and comparison of two Co-PI: P. McCue, techniques for vitrifying E. Squires biopsied equine embryos Evaluation of procedures for PI: E. Squires Andreini 1/07-12/07 freezing day 7 and 8 equine embryos Production of in vitro equine PI: E. Squires XY, Inc. 7/07-12/07 6,918 embryos with sexed semen General studies on equine PI: E. Squires Private 1/07-12/07 55,331 reproduction donors Dietary supplementation of PI: E. Squires PEG 7/07-6/08 21,386 DHA and alpha-tocopherol and the effects on spermatozoal characteristics Fertility of mares inseminated PI: E. Squires PEG 7/09-6/10 30,870 with frozen/thawed semen centrifuged through a single layer of density gradient Effect of supplementary inositol PI: T. Suh PEG 7/06-6/07 20,603 1,4,5-triphosphate injection on Co-PI: G. Seidel fertilization and embryo development in equine ICSI 55

58 Scientific Publications 2007 Chapters in Books Bruemmer JE Sexual behavior of the breeding stallion. In: CICADE 2007 (Arno J, ed), San Jose, Costa Rica, p 10. Bruemmer JE Clinical case studies. In: CICADE 2007 (Arno J, ed), San Jose, Costa Rica, p 12. Bruemmer JE Success with cooled and frozen semen. In: CICADE 2007 (Arno J, ed), San Jose, Costa Rica, p 14. Bruemmer JE Anatomy and physiology of the stallion. In: CICADE 2007 (Arno J, ed), San Jose, Costa Rica, p 74. Bruemmer JE How to collect equine semen. In: CICADE 2007 (Arno J, ed), San Jose, Costa Rica, p 79. Carnevale EM Collection and transfer of oocytes in mares. In: Current Therapy in Equine Reproduction (Samper JC, Pycock JF, McKinnon AO, eds), Saunders Elsevier, pp Graham JK, Card C Preservation of genetics from dead or dying stallions. In: Current Therapy in Equine Reproduction (Samper JC, Pycock JF, McKinnon AO, eds), Saunders Elsevier, pp McCue PM Ovarian abnormalities. In: Current Therapy in Equine Reproduction (Samper JC, Pycock JF, McKinnon AO, eds), Saunders Elsevier, pp McCue PM Ovulation failure. In: Current Therapy in Equine Reproduction (Samper JC, Pycock JF, McKinnon AO, eds), Saunders Elsevier, pp McKinnon AO, Squires EL Embryo transfer and related technologies. In: Current Therapy in Equine Reproduction (Samper JC, Pycock JF, McKinnon AO, eds), Saunders Elsevier, pp Barfield JP, McCue PM, Seidel GE Jr, Squires EL The effect of dehydrating large equine embryos with galactose prior to cryopreservation. Proc 77th Int Symp Equine Embryo Transfer, pp Carnevale EM Cooling and cryopreservation of equine embryos. In: Equine Breeding Management and Artificial Insemination, 2nd Ed. (Samper J, ed), Saunders Elsevier, St. Louis, MO, pp

59 2010 Carnevale EM Horses: Commercial oocyte technologies. In: Encyclopedia of Biotechnology in Agriculture and Food (Heldman DR, Hoover DG, Wheeler MB, eds), CRC Press, Taylor and Francis Group, LLC, 1:1, pp Carnevale EM, Coutinho da Silva MA Assisted reproductive techniques. In: Equine Internal Medicine, 3rd Ed. (Reed SM, Bayly WM, Sellon DC, eds), W.B. Saunders, St. Louis, MO, pp Seidel GE Jr, Cullingford EL, Stokes JE, Carnevale EM, McCue PM Pregnancy rates following transfer of biopsied and/or vitrified equine embryos: Evaluation of two biopsy techniques. In: Proceedings of the International Symposium on Equine Reproduction, Anim Reprod Sci 121S:S297- S Bruemmer JE Freezing epididymal spermatozoa. In: Equine Reproduction (McKinnon AO, Squires EL, Vaala WE, Varner DD, eds), Blackwell Publishing, Ames, IA, pp Carnevale EM Mature oocyte collection. In: Equine Reproduction (McKinnon AO, Squires EL, Vaala WE, Varner DD, eds), Blackwell Publishing, Ames, IA, pp Carnevale EM Oocyte transfer. In: Equine Reproduction (McKinnon AO, Squires EL, Vaala WE, Varner DD, eds), Blackwell Publishing, Ames, IA, pp Gee EK, McCue PM Mastitis. In: Equine Reproduction, 2nd Ed. (McKinnon AO, Squires EL, Vaala WE, Varner DD, eds), Wiley-Blackwell, Ames, IA, pp McCue PM Inhibin immunotherapy. In: Equine Reproduction, 2nd Ed. (McKinnon AO, Squires EL, Vaala WE, Varner DD, eds), Wiley-Blackwell, Ames, IA, pp McCue PM Induction of ovulation. Chap. 43 in: Blackwell s Five-Minute Veterinary Consult Clinical Companion: Equine Theriogenology (Carleton C, ed), Wiley-Blackwell, Ames, IA. McCue PM Superovulation of the mare. Chap. 63 in: Blackwell s Five-Minute Veterinary Consult Clinical Companion: Equine Theriogenology (Carleton C, ed), Wiley-Blackwell, Ames, IA. McCue PM Disorders of ovulation. In: Equine Reproduction, 2nd Ed. (McKinnon AO, Squires EL, Vaala WE, Varner DD, eds), Wiley-Blackwell, Ames, IA, pp McCue PM, DeLuca CA, Wall JJ Cooled and transported embryo technology. In: Equine Reproduction, 2nd Ed. (McKinnon AO, Squires EL, Vaala WE, Varner DD, eds), Wiley-Blackwell, Ames, IA, pp McCue PM, Ferris RA The abnormal estrous cycle. In: Equine Reproduction, 2nd Ed. (McKinnon AO, Squires EL, Vaala WE, Varner DD, eds), Wiley-Blackwell, Ames, IA, pp McCue PM, McKinnon AO Ovarian abnormalities. In: Equine Reproduction, 2nd Ed. (McKinnon AO, Squires EL, Vaala WE, Varner DD, eds), Wiley-Blackwell, Ames, IA, pp McCue PM, McKinnon AO Pregnancy examination. In: Equine Reproduction, 2nd Ed. (McKinnon AO, Squires EL, Vaala WE, Varner DD, eds), Wiley-Blackwell, Ames, IA, pp

60 McCue PM, Scoggin CF, Lindholm ARG Estrus. In: Equine Reproduction, 2nd Ed. (McKinnon AO, Squires EL, Vaala WE, Varner DD, eds), Wiley-Blackwell, Ames, IA, pp McCue PM, Sitters S Lactation. In: Equine Reproduction, 2nd Ed. (McKinnon AO, Squires EL, Vaala WE, Varner DD, eds), Wiley-Blackwell, Ames, IA, pp McKinnon AO, McCue PM Induction of ovulation. In: Equine Reproduction, 2nd Ed. (Mc- Kinnon AO, Squires EL, Vaala WE, Varner DD, eds), Wiley-Blackwell, Ames, IA, pp McKinnon AO, McCue PM Uterine abnormalities. In: Equine Reproduction, 2nd Ed. (Mc- Kinnon AO, Squires EL, Vaala WE, Varner DD, eds), Wiley-Blackwell, Ames, IA, pp Olsen L, Carnevale EM Normal anatomy. In: Equine Reproduction (McKinnon AO, Squires EL, Vaala WE, Varner DD, eds), Blackwell Publishing, Ames, IA, pp Seidel GE Jr Future reproductive technology. In: Equine Reproduction, 2nd Ed. (McKinnon AO, Squires EL, Vaala WE, Varner DD, eds), Wiley-Blackwell, Ames, IA, pp Squires EL, McCue PM Superovulation. In: Equine Reproduction, 2nd Ed. (McKinnon AO, Squires EL, Vaala WE, Varner DD, eds), Wiley-Blackwell, Ames, IA, pp Refereed Journal Articles Lindbloom SL, Farmerie TA, Clay CM, Seidel GE Jr, Carnevale EM Potential involvement of EGF-like growth factors and phosphodiesterases in initiation of equine oocyte maturation. Anim Reprod Sci 103: Logan NL, McCue PM, Alonso MA, Squires EL Evaluation of three equine FSH superovulation protocols in mares. Anim Reprod Sci 102: McCue PM Evaluation of turbidimetric immunoassay for measurement of IgG concentration in foals. Am J Vet Res 68: McCue PM, Leblanc MM, Squires EL efsh in clinical equine practice. Theriogenology 68: McCue PM, Logan NL, Magee C Management of the transition period: Hormone therapy. Equine Vet Educ 19: McCue PM, Logan NL, Magee C Management of the transition period: Physiology and artificial photoperiod. Equine Vet Educ 19: McCue PM, Magee C, Gee EK Comparison of compounded deslorelin and hcg for induction of ovulation in mares. J Equine Vet Sci 27: Niswender GD, Davis TL, Griffeth RJ, Bogan RL, Monser K, Bott RC, Bruemmer JE, Nett TM Judge, jury and executioner: the auto-regulation of luteal function. Soc Reprod Fertil (Suppl) 64:

61 Purcell SH, Seidel GE Jr, McCue PM, Squires EL Aspiration of oocytes from transitional, cycling, and pregnant mares. Anim Reprod Sci 100: Seidel GE Jr Overview of sexing sperm. Theriogenology 68: Squires EL, McCue PM Superovulation in mares. Anim Reprod Sci 99:1-8. Vanderwall DK, Marquardt JL, Woods GL Use of a compounded long-acting progesterone formulation for equine pregnancy maintenance. J Equine Vet Sci 27: Carnevale EM Clinical considerations regarding assisted reproductive procedures in horses. J Equine Vet Sci 28: Carnevale EM The mare as a model for follicular maturation and reproductive aging in the woman. Theriogenology 69: Lawler JB, Hassel DM, Magnuson RJ, Hill AE, McCue PM, Traub-Dargatz JL Adsorptive effects of di-tri-octahedral smectite on Clostridium perfringens alpha, beta, and beta-2 exotoxins and equine colostral antibodies. Am J Vet Res 69: Lindbloom SM, Farmerie TA, Clay CM, Seidel GE Jr, Carnevale EM Potential involvement of EGF-like growth factors and phosphodiesterases in initiation of equine oocyte maturation. Anim Reprod Sci 103: Loomis PR, Graham JK Commercial semen freezing: Individual male variation in cryosurvival and response of stallion sperm to customized freezing protocols. Anim Reprod Sci 105: McCue PM The problem mare: management philosophy, diagnostic procedures, and therapeutic options. J Equine Vet Sci 28: McCue PM, Patten M, Denniston DJ, Bruemmer JE, Squires EL Strategies for using efsh for superovulating mares. J Equine Vet Sci 28: Mocé E, Graham JK In vitro evaluation of sperm quality. Anim Reprod Sci 105: Paccamonti D, Carnevale E Pregnancy loss during the embryonic period. Pferdeheilkunde 14: Squires EL Hormonal manipulation of the mare: A review. J Equine Vet Sci 28: Altermatt JL, Suh TK, Stokes JE, Carnevale EM Effects of age and equine follicle-stimulating hormone (efsh) on collection and viability of equine oocytes assessed by morphology and developmental competency after intracytoplasmic sperm injection (ICSI). Reprod Fertil Dev 21: Barfield JP, McCue PM, Squires EL, Seidel GE Jr Effect of dehydration prior to cryopreservation of large equine embryos. Cryobiology 59:

62 Bruemmer JE, Wilson CH, Coutinho da Silva M, Squires EL Effects of hyaluronan supplementation on cryopreserved equine spermatozoa hyaluronan and cryopreserved equine spermatozoa. J Equine Vet Sci 29: Campos-Chillon LF, Suh TK, Barcelo-Fimbres M, Seidel GE Jr, Carnevale EM Vitrification of early-stage bovine and equine embryos. Theriogenology 71: Cox TJ, Squires EL, Carnevale EM Effect of follicle size and follicle-stimulating hormone on ovulation induction and embryo recovery in the mare. J Equine Vet Sci 29: Gee EK, DeLuca C, Stylski JL, McCue PM Efficacy of medroxyprogesterone acetate in suppression of estrus in cycling mares. J Equine Vet Sci 29: Glazar AI, Mullen SF, Liu J, Benson JD, Critser JK, Squires EL, Graham JK Osmotic tolerance limits and membrane permeability characteristics of stallion spermatozoa treated with cholesterol. Cryobiology 59: Hackett ES, Bruemmer J, Hendrickson DA, McCue PM Buccal mucosal urethroplasty for treatment of recurrent hemospermia in a stallion. J Am Vet Med Assoc 235: Magee C, Foradori CD, Bruemmer JE, Arreguin-Arevalo JA, McCue PM, Handa RJ, Squires EL, Clay CM Biological and anatomical evidence for kisspeptin regulation of the hypothalamicpituitary-gonadal axis of estrous horse mares. Endocrinology 150: Barcelo-Fimbres ML, Campos-Chillon F, Seidel GE Jr In vitro fertilization using nonsexed and sexed bovine sperm: Sperm concentration, sorter pressure, and bull effects. Reprod Dom Anim PMID Bott RC, Ashley RL, Henkes LE, Antoniazzi AQ, Bruemmer JE, Niswender GD, Bazer FW, Spencer TE, Smirnova NP, Anthony RV, Hansen TR Uterine vein infusion of interferon tau (IFNT) extends luteal lifespan in ewes. Biol Reprod 82: Bott RC, Schwartzenberger SM, Bruemmer JE, Arreguin-Arevalo JA, Niswender GD Recombinant equine luteinizing hormone stimulates production of progesterone from murine Leydig, ovine small luteal, and equine granulosal cells. J Equine Vet Sci 31: Cullingford EL, Squires EL, McCue PM, Seidel GE Jr Attempts at superovulation of mares with porcine follicle stimulating hormone and recombinant equine follicle stimulating hormone. J Equine Vet Sci 30: Davis TL, Bott RC, Slough TL, Bruemmer JE, Niswender GD Progesterone inhibits oxytocin and prostaglandin F2α stimulated increases in intracellular calcium concentrations in small and large ovine luteal cells. Biol Reprod 82: Ferris RA, McCue PM The effects of dexamethasone and prednisolone on pituitary and ovarian function in the mare. Equine Vet J 42: Hackett ES, McCue PM Evaluation of a veterinary glucometer for use in horses. J Vet Int Med 24:

63 Mahmoud KG, Scholkamy TH, Ahmed YF, Seidel GE Jr, Nawito MF Effect of different combinations of cryoprotectants on in vitro maturation of immature buffalo (Bubalus bubalis) oocytes vitrified by straw and open-pulled straw methods. Reprod Dom Anim 45: Meyers-Brown GA, McCue PM, Niswender KD, Squires EL, DeLuca CA, Bidstrup LA, Colgin M, Famula TR, Roser JF Superovulation in mares using recombinant equine follicle stimulating hormone: Ovulation rates, embryo retrieval, and hormone profiles. J Equine Vet Sci 30: Paccamonti DL, Carnevale EM Early embryonic death in mares. Clin Theriogenol 2: Peel RK, Whittier JC, Enns RM, Grove AV, Seidel GE Jr Effect of sex versus twelve hour interval between two prostaglandin F2a injections administered with gonadotropin hormone releasing hormone and controlled internal drug release on pregnancy rates in beef cows. Prof Anim Sci. In Press. Slough TL, Rispoli LA, Carnevale EM, Niswender GD, Bruemmer JE Temporal gene expression in equine corpora lutea based on serial biopsies in vivo. J Anim Sci doi /jas Spizziri BE, Fox MH, Bruemmer JE, Squires EL, Graham JK Cholesterol-loaded cyclodextrins and fertility potential of stallion spermatozoa. Anim Reprod Sci 118: Tardif S, Brady HA, Breazeale KR, Bi M, Thompson LD, Bruemmer JE, Bailey LB, Hardy DM Zonadhesin D3 Polypeptides vary among species but are similar in Equus species capable of interbreeding. Biol Reprod 82: Abstracts Altermatt JL, Suh TK, Stokes JE, Squires EL, Seidel GE Jr, Carnevale EM Effects of age and FSH on collection of equine oocytes and developmental competency after intracytoplasmic sperm injection. Theriogenology 68: Bruemmer J, Lewis M, Prenni J, Squires E, Hansen TR Gene expression in the equine endometrium during maternal recognition of pregnancy. Biol Reprod (Special Issue):98 (abstr 98). Campos-Chillon LF, Barcelo-Fimbres M, Carnevale EM, Seidel GE Jr Use of bovine oocytes to evaluate in vitro fertilizing capacity of equine sperm. Theriogenology 68: Campos-Chillon LF, Clay CM, Altermatt JL, Bouma GJ, Carnevale EM Differences in resumption of oocyte maturation in young and old mares. Reprod Fertil Dev. Davis TL, Bott RC, Bruemmer JE, Niswender GD Progesterone alters membrane cholesterol content and oxytocin signaling in ovine small luteal cells. Biol Reprod (Special Issue):203 (abstr 497). French WB, LeClaire JD, Hurley K, Moffett PD, Bruemmer JE The effect of concentration on the motility of cooled stallion semen. Annual Meeting Equine Science Society. Luszcz NA, Bott RC, Squires EL, Graham JK, Bruemmer JE CD44 hyaluronon receptor in stallion spermatozoa. Annual Meeting Equine Science Society. 61

64 Magee C, Bruemmer JE, Foradori CD, McCue PM, Clay C Evaluation of kisspeptin in the hypothalamic-pituitary-gonadal axis of the mare. Theriogenology 68: Magee C, Foradori C, Bruemmer J, McCue P, Handa R, Clay C Distribution of kisspeptin and GnRH immunoreactive neuronal contacts in the preoptic area and hypothalamus of the mare. Biol Reprod (Special Issue):185. McCue PM, Gee EK, Magee C, Woods GL, Squires EL Hormones and other medical therapies in embryo transfer recipient mares. ET Proc, Soc for Theriogenology, pp Patten ML, Denniston DJ, Magee C, Bruemmer JE, McCue PM, Squires EL Superovulation in the mare: Evaluation of an efsh step down technique and ovulation inducing agents. Proc Ann Mtg Equine Science Society. Preis KA, Seidel GE Jr, Gardner DK Effect of in vitro maturation medium on subsequent quantity and quality of developing embryos. Proc Am Soc Reproduction Medicine. Scoggin CF, McCue PM How to assess and stabilize a mare suspected of periparturient hemorrhage in the field. Ann Conf Am Assoc Equine Pract. Squires EL, Patten ML, Zumbrunnen J, Bruemmer JE, Denniston DD, McCue PM Variation in progesterone in early gestation after nonsurgical embryo transfer. Proc 53rd Ann Conv Am Assoc Equine Pract. Vanderwall DK, Rasmussen DM, Woods GL Use of exogenous oxytocin to block luteolysis in mares: A plausible method of long-term suppression of estrus. Proc Ann Conv Am Assoc Equine Pract Altermatt JL, Suh TK, Stokes JE, Campos-Chillon LF, Carnevale EM Effect of mare age on oocyte morphology and developmental competence after ICSI. Reprod Fertil Dev 20:81. Barfield JP, McCue PM, Seidel GE Jr, Squires EL The effect of dehydrating large equine embryos with galactose prior to cryopreservation. Proc 7th Int Symp Equine Embryo Transfer, pp Barfield JP, McCue PM, Squires EL, Seidel GE Jr Cryopreservation of large equine embryos. Proc Soc Study Reprod, Biol Reprod 78:706. Bott R, Ashley R, Henkes L, Bruemmer J, Niswender G, Spencer T, Bazer F, Anthony R, Hansen T Uterine vein infusion of interferon-tau acts systemically to induce ISG15 in the ovine corpus luteum. 41st Soc Study Reprod, Annual Meeting and Abstracts, abstr #5636, p 221. Bruemmer JE, Hardy QP, Denniston DJ, Schumacher JK, Squires EL Effect of cooling prior to cryopreservation of stallion spermatozoa. Proc 5th Int Symp Stallion Reprod. Campos-Chillon LF, Clay CM, Altermatt JL, Bouma GJ, Carnevale EM Differences in resumption of oocyte maturation in young and old mares. Reprod Fertil Dev 20:81. Carnevale EM, Cox TJ How to do induction of ovulation. XIV Italian Assoc Equine Vet Practitioners, pp

65 DeLuca CA, McCue PM, Patten ML, Squires EL Comparison of three doses of refsh for superovulation of mares. Theriogenology 70: Gee EK, Bruemmer JE, Siciliano PD, McCue PM, Squires EL Effects of dietary vitamin E supplementation on spermatozoa quality in stallions with suboptimal post-thaw motility. Proc 5th Int Symp Stallion Reproduction, Anim Reprod Sci 107: Gee EK, McCue PM, DeLuca CA, Stylski JL Efficacy of medroxyprogesterone acetate in suppression of estrous behavior and follicular activity in cycling mares. Theriogenology 70:588. Gee EK, McCue PM, DeLcua CA, Stylski JL Efficacy of medroxyprogesterone acetate in suppression of estrous behavior and follicular activity in cycling mares. Proc Soc Study Reprod, Biol Reprod 78:706. Hollingshead S, Nett T, McCue P Development of an ELISA for analysis by a densimeter to quantify progesterone in mares. CVMBS Research Day Poster. Hudson MR, Seidel GE Jr, Squires EL, Spizziri BE, Walker DJ, Graham JK Capacitation of stallion spermatozoa evaluated by fertilization of bovine oocytes. Reprod Fertil Dev 20: Magee C, Arreguin-Arevalo A, Bruemmer J, Mrdutt M, Cantlon J, McCue P, Clay C Peripheral kisspeptide administration elicits an increase in serum FSH in the diestrous mare. Proc Soc Study Reprod, Biol Reprod 78:706. Magee C, Foradori CD, Bruemmer JE, Corning C, Farmerie TA, McCue PM, Handa RJ, Clay CM Evaluation of kisspeptin in the estrous mare. Proc First Ann Rocky Mountain Reprod Sci Symp. McCue PM Techniques to monitor foaling and care of the normal newborn foal. 18th Ann Conf Technicians, Colorado State University, Fort Collins, CO. McCue PM Clinical Cases in Equine Reproduction. 69th Ann Conf Veterinarians, Colorado State University, Fort Collins, CO. McCue PM Workup of the problem mare: Wet lab. Proc West Coast Equine Reprod Symp III. McCue PM The peripartum mare. Proc West Coast Equine Reprod Symp III. McCue PM The problem mare: Diagnosis and therapy. Proc West Coast Equine Reprod Symp III. McCue P, DeLuca C, Patten M, Squires EL Effect of a sham transcervical embryo transfer on plasma progesterone concentrations in recipient mares. Proc 7th Int Symp Equine Embryo Transfer. Spizziri B, Graham J, Hudson M In vitro capacitation of frozen/thawed stallion sperm. J Androl (March/April Suppl):38. Squires EL, Suh TK, Graham JK, Carnevale EM Use of sexed, refrozen spermatozoa for ICSI. Havemeyer Foundation 7th Int Symp Equine Embryo Transfer, p

66 2009 Antoniazzi AQ, Henkes LE, Bott RC, Ashley R, Bruemmer J, Niswender GD, Moss G, Alexander B, Oliveira JFC, Spencer ET, Bazer FW, Hansen TR Endocrine action of interferon-tau on the corpus luteum in sheep: Implication of antiluteolytic and luteotropic mechanisms. Proc Soc Study Reprod, Biol Reprod 81:573. Bott RC, Ashley RL, Henkes LE, Antoniazzi AQ, Bruemmer JE, Smirnova NP, Anthony RV, Niswender GD, Bazer FW, Spencer TE, Hansen TR Uterine vein infusion of interferon tau (IFNT) stimulates luteal gene expression, prevents anti-steroidogenic actions of prostaglandin F2alpha (PGF) and extends luteal lifespan in ewes. Biol Reprod 81(Suppl 1):577. Bott RC, Schwartzenberger SM, Bruemmer JE, Arreguin-Arevalo JA, Niswender GD Recombinant equine luteinizing hormone (relh) stimulates production of progesterone from murine leydig, equine granulosal and ovine small luteal cells. J Equine Vet Sci 29: Campos-Chillon LF, Bouma GJ, Seidel GE Jr, Clay CM, Carnevale EM Age effect on gene expression and mitochondrial DNA in the equine oocyte and follicle. Proc Soc Theriogenology 1:225. Carnevale EM, Graham JK, Suh TK, Stokes JE, Squires EL Foals produced after ICSI using frozen, sex-sorted, refrozen sperm. Proc Ann Mtg Int Embryo Transfer Soc, Reprod Fertil Dev 21:228. DeLuca CA, Gee EK, McCue PM How to remove large endometrial cysts with an improvised snare: a simple technique for practitioners. Proc Am Assoc Equine Practitioners 55: Ferris RA, McCue PM The effects of dexamethasone and prednisolone on pituitary and ovarian function in the mare. Proc Soc Theriogenology, Clin Theriogenol 1:225. Ferris RA, McCue PM How to use a quantitative turbidimetric immunoassay to determine immunoglobulin G concentrations in neonatal foals. Proc Am Assoc Equine Practitioners 55: Gee EK, Bruemmer JE, Siciliano PD, Morel PCH, Engle TE, Squires EL The influence of dietary vitamin E on serum vitamin E and fatty acids, and spermatozoa fatty acids in stallions. J Equine Vet Sci 29: Kaula N, Andrews A, Durso C, Dixon C, Graham JK Classification of hyperactivated spermatozoa using a robust minimum bounding square ratio algorithm. Proc 31st Ann Int Conf IEEE Engineering in Medicine and Biology Soc. Krull AL, Bouma GJ, Hansen TR, Bruemmer JE Transcriptional profile of day 18 pregnant and non-pregnant equine endometria: Insight into maternal recognition of pregnancy. J Equine Vet Sci 29: Lund G Comparison of timing of oocyte collection and methods of vitrification for ICSIproduced equine embryos. Proc Rocky Mountain Reprod Sci Symp, Colorado State Univ, Fort Collins, CO. McCue PM What s new in equine reproduction? 70th Ann Conf for Veterinarians, Colorado State Univ, Fort Collins, CO. McCue PM, DeLuca CA, Ferris RA, Wall JJ How to evaluate equine embryos. Proc Am Assoc Equine Pract 55:

67 Stokes JE, Squires EL, Suh TK, Altermatt JL, Carnevale EM Effect of developmental stage of ICSI-produced equine embryos on pregnancy rates. Proc Ann Mtg Int Embryo Transfer Soc, Reprod Fertil Dev 21:164. Verver JMP, McCue PM, Cullingford EL, Squires EL Comparison of porcine FSH and recombinant equine FSH for superovulation in mares. Proc British Equine Vet Assoc, p 165. Wall JJ, McCue PM, Brink ZA, Seidel GE Jr Pregnancy outcome following embryo transfer in the horse. Proc Rocky Mountain Reprod Sci Symp, Colorado State Univ, Fort Collins, CO. Yourey R Potential use of SDS-PAGE and silver staining to characterize composition of limited numbers of bovine and equine zona pellucidae. Proc Rocky Mountain Reprod Sci Symp, Colorado State Univ, Fort Collins, CO Bott RC, Ashley RL, Henkes LE, Antoniazzi AQ, Bruemmer JE, Niswender GD, Bazer FW, Spencer TE, Smirnova NP, Anthony RV, Hansen TR Uterine vein infusion of interferon tau (IFNT) extends luteal lifespan in ewes. Biol Reprod 82: Bruemmer JE, Bouma GJ, Hess A, Hansen TR, Squires EL Gene expression in the equine endometrium during maternal recognition of pregnancy. Anim Reprod Sci 121S: Burns PD, White NR, Cheatham RD, Romero R, Bruemmer JE, Engle TE Effect of fish meal supplementation on bovine plasma and luteal omega- 3 fatty acid content. Annual Meeting, Society for the Study of Reproduction. Burroughs C, Seidel G Jr Effect of addition of camp regulators to bovine in vitro maturation medium. Proc Ann Mtg Am Soc Anim Sci, J Anim Sci 88(E-Suppl):398, abstr T283. Cheatham RD, White NR, Burns PD, Chestnut ES, Hickman JS, Michishima JKK, Suh D, Bruemmer JE, Engle TE Effect of omega - 3 fatty acids on prostaglandin F2a induced mitogen-activated kinase signaling in bovine luteal cells in vitro. Annual Meeting, Society for the Study of Reproduction. Cleys ER, Bouma GJ, Bruemmer JE Effects of pregnancy status on organic anion transporters and prostaglandin receptors in the equine endometrium: Insights into maternal recognition of pregnancy in the mare. Annual Meeting, Society for the Study of Reproduction. Cullingford EL, Stokes JE, Bouma GJ, Seidel GE Jr Successful transfer of fresh and vitrified biopsied equine embryos. Reprod Fertil Dev 22: Davis TL, Bott RC, Slough TL, Bruemmer JE, Niswender GD Progesterone inhibits oxytocinand prostaglandin F2alpha-stimulated increases in intracellular calcium concentrations in small and large ovine luteal cells. Biol Reprod 82: Dixon C, Kaula N, Graham JK, Andrews A, Durso C Effect of Sildenafil, Vardenafil and Tadalafil on in-vitro sperm motility and hyperactivation in the stallion. Am Urology Assoc, San Francisco, CA. Ferris RA, Lindholm AR, McCue PM Relationship between donor mare age, semen type, and early embryonic development. Proc Soc Theriogenology, Clin Theriogenol 2:

68 Ferris RA, McCue PM Relationship between donor mare age, semen type, and early embryonic development. Clin Therio. Ferris RA, Veir JK, Lappin MR, McCue PM Development and clinical application of a broad range 16S quantitative PCR assay for the detection of bacteria in the uterus of the mare. Anim Reprod Sci 121S: Gee EK, Bruemmer JE, Siciliano PD, Morel PCH, Engle TE, McCue PM, Squires EL Effects on spermatozoa of dietary supplementation of vitamine E and omega-3 fatty acids in stallions with poor post-thaw motility. Anim Reprod Sci 121S: Giles R, Seidel G, McConnel C, McSweeney K A comparison of conception rates between new and re-used Eazi-Breed CIDRs. Proc Ann Mtg Am Soc Anim Sci, J Anim Sci 88(E-Suppl):677, abstr W286. Hackett ES, McGreevey N, Lunn DP, McCue PM Detection of bacteremia and host response in healthy neonatal foals. Proc Int Veterinary Emergency and Critical Care Symposium. Kaula N, Dixon C, Graham JK, Andrews A, Durso C A new method for computer aided in vitro identification and classification of hyperactivated spermatozoa. Am Urology Assoc, San Francisco, CA. Lindholm ARG, Bloemen EHG, Brooks RM, Ferris RA, McCue PM Comparison of deslorelin and buserelin in mares: LH response and induction of ovulation. Anim Reprod Sci 121S: Maclellan LJ, Stokes JE, Preis KA, McCue PM, Carnevale EM Vitrification, warming, ICSI and transfer of equine oocytes matured in vivo. Anim Reprod Sci 121S: Magee C, Bruemmer JE, Arreguin-Arevalo JA, Nett TM, Clay CM Effects of repeated injection of equine kisspeptide in diestrous and estrous mares. Anim Reprod Sci S121: McCue PM, Ferris RA, Lindholm AR, DeLuca CA Embryo recovery procedures and collection success: Results of 492 embryo-flush attempts. Proc Am Assoc Equine Pract 56: McCue PM, Wall JJ, Brink ZA, Seidel GE Jr Live foal rate: Effects of embryo transfer and donor age. Proc Ann Mtg Int Embryo Transfer Soc, Reprod Fertil Dev 22:247. Mocé E, Purdy PH, Graham JK Treating ram sperm with cholesterol-loaded cyclodextrins improves cryosurvival. Anim Reprod Sci 118: Mocé E, Blanch E, Tomás C, Graham JK Use of cholesterol in sperm cryopreservation: Present moment and perspectives to future. Reprod Dom Anim 45 (Suppl 2): Moraes EA, Graham JK, Torres CAA, Meyers M, Spizziri B Delivering cholesterol or cholestanol to bull sperm membranes improves cryosurvival. Anim Reprod Sci 118: Powers JG, Baker DL, Bruemmer JE, Ackerman MG, Nett TM Maternal passive transfer of GnRH antibodies does not change reproductive development in elk calves. Annual Meeting, Society for the Study of Reproduction. Purdy PH, Mocé E, Stobart R, Murdoch WJ, Moss GE, Larson B, Ramsey S, Graham JK, Blackburn HD The fertility of ram sperm held for 24 h at 5 C prior to cryopreservation. Anim Reprod Sci 118:

69 Rodrigues BL, Silveira JC, Bruemmer JE, Bouma GJ, Carnevale EM, Clay CM Caspase 3 expression in equine granulosa cells and oocytes in young and old mares. 43rd Annual Meeting, Society for the Study of Reproduction. Seidel G Current status of sexed semen technology. Proc Ann Mtg Am Soc Anim Sci, J Anim Sci 88(E-Suppl):783, abstr 887. Seidel GE Jr, Cullingford EL, Stokes JE, Carnevale EM, McCue PM Pregnancy rates following transfer of biopsied and/or vitrified equine embryos: Evaluation of two biopsy techniques. Anim Reprod Sci 121S: Spizziri BE, Fox MH, Bruemmer JE, Squires EL, Graham JK Cholesterol-loaded-cyclodextrins and fertility potential of stallion spermatozoa. Anim Reprod Sci 118: Spizziri BE, Kaula N, Squires EL, Graham JK In vitro capacitation of stallion spermatozoa. Anim Reprod Sci 121 (Suppl):S181-S183. White NR, Burns PD, Charumilinda J, Bryant AD, Prosser ZT, Bruemmer JE, Engle TE Effect of omega-3 fatty acids on prostaglandin F2a induced cyclooxygenase-2 (Cox-2) gene expression in bovine luteal cells in vitro. Annual Meeting, Society for the Study of Reproduction J.E. Bruemmer Invited Presentations Managing Stallions Conferencia International de Caballos de Deporte, Costa Rica Factors Affecting Fertility of Fresh, Cooled and Frozen Semen Conferencia International de Caballos de Deporte, Costa Rica E.M. Carnevale Clinical Use of Intracytoplasmic Sperm Injection in Horses 53rd Annual Convention, American Association of Equine Practitioners, Orlando, FL Emerging Embryo Technologies American Embryo Transfer Association and Society for Theriogenology Annual Conference, Monterey, CA Preservation of Equine Embryos American Embryo Transfer Association and Society for Theriogenology Annual Conference, Monterey, CA C.M. Clay Estradiol Regulation of the LH Surge Department of Biology, University of Northern Colorado, Greeley, CO 67

70 Estrogen Regulation of the GnRH Receptor and the LH Surge Department of Animal Science, University of Georgia, Athens, GA J.K. Graham Improving Gamete and Embryo Quality NIH Achieving High Throughput Repositories for Biomedical Germplasm Preservation Workshop, Bethesda, MD P.M. McCue Reproduction Problems in the Mare and Stallion Italian Equine Veterinary Association, Cremona, Italy Ovarian Abnormalities British Equine Veterinary Association, Edinburgh, Scotland Hormone Therapy British Equine Veterinary Association, Edinburgh, Scotland Freezing Equine Embryos Quarter Horse Congress, Columbus, OH Breeding Management Annual National Shire Horse Meeting, Ft. Lupton, CO Advanced Reproductive Procedures Annual National Shire Horse Meeting, Ft. Lupton, CO Ovarian Problems in the Mare Indiana Association of Equine Practitioners, Delphi, IN Uterine Prolems in the Mare Indiana Association of Equine Practitioners, Delphi, IN Hormone Therapy Indiana Association of Equine Practitioners, Delphi, IN Breeding Management Indiana Association of Equine Practitioners, Delphi, IN Advanced Reproductive Procedures Indiana Association of Equine Practitioners, Delphi, IN Clinical Cases in the Mare and Stallion Indiana Association of Equine Practitioners, Delphi, IN Foaling Management Countryside Large Animal Hospital, Greeley, CO Foaling Management Pioneer Equine Hospital, Oakdale, CA Equine Viral Arteritis Pioneer Equine Hospital, Oakdale, CA Prediction of Foaling, Foaling and Care of the Newborn Foal Owner Seminar, Glenwood Springs, CO Equine Viral Arteritis Annual Conference for Veterinarians, Colorado State University, Ft. Collins, CO G.E. Seidel, Jr. History of Procedures to Sex Sperm Select Sires Meeting, Loveland, CO Sexing Sperm Texas A&M University, College Station, TX Sexing Mammalian Sperm Cornell University, Ithaca, NY 68

71 Improving Gamete and Embryo Quality NIH Cryopreservation Workshop, Bethesda, MD How Identical Will Clones be with the Original? American Cutting Horse Association, Lexington, KY Sexing Mammalian Sperm XII Latin American Congress on Buiatrics, Acapulco, Mexico Sexing Sperm Annual Meeting of the American College of Theriogenologists, Monterey, CA Vitrification of Embryos American Embryo Transfer Association, Monterey, CA E.L. Squires Managing Stallions Conferencia International de Caballos de Deporte, Costa Rica Factors that Improve Embryo Transfer Rates Conferencia International de Caballos de Deporte, Costa Rica Vitrification, Superovulation and Endometritis Biotay Equine Practitioners Meeting, Argentina Breeding the Problem Mare Biotay Equine Practitioners Meeting, Argentina Advances in Equine Reproduction EquiScience Veterinary Spring Seminar, Bellingham, WA Advances in Equine Reproduction Equine Seminar, Emerald Downs, Auburn, WA Stallion Reproduction Florida Thoroughbred Association, Ocala, FL The Role of Omega-3 and -6 Fatty Acids in Regulation of Reproductive Function in Horses ADSA/ PSA/AMPA/ASAS Joint Annual Conference, San Antonio, TX Program Chair for Equine Embryo Transfer Symposium Society for Theriogenology Annual Meeting, Monterey, CA Assessing Embryo Quality Prior to Transfer Society for Theriogenology Annual Meeting, Monterey, CA Use of efsh for Broodmare Reproduction Bioniche Animal Health Annual Meeting, Athens, GA Anatomy and Physiology of the Mare Bioniche Animal Health Canada, Belleville, Ontario Use of efsh and Settle for Treatment of Mares Bioniche Animal Health Canada, Belleville, Ontario Status of the Use of efsh - Bioniche Research Meeting, Belleville, Ontario Use of Settle for Treatment of Endometritis Bioniche Research Meeting, Belleville, Ontario Stallion Management New England Regional Veterinary Conference, Portland, ME Hormonal Control of the Mare s Cycle New England Regional Veterinary Conference, Portland, ME Breeding the Problem Mare New England Regional Veterinary Conference, Portland, ME 69

72 Twinning and Early Embryonic Death New England Regional Veterinary Conference, Portland, ME Cooled and Frozen Semen New England Regional Veterinary Conference, Portland, ME New Reproductive Techniques New England Regional Veterinary Conference, Portland, ME Assisted Reproductive Techniques in the Horse Kentucky Veterinary Medical Association Annual Meeting/Annual Mid-America Veterinary Conference, Louisville, KY Recent Advances in Equine Reproduction Kentucky Veterinary Medical Association Annual Meeting/Annual Mid-America Veterinary Conference, Louisville, KY Hormonal and Non-Hormonal Management of the Broodmare Kentucky Veterinary Medical Association Annual Meeting/Annual Mid-America Veterinary Conference, Louisville, KY Improving a Stallion s Reproductive Performance Kentucky Veterinary Medical Association Annual Meeting/Annual Mid-America Veterinary Conference, Louisville, KY Advances in Equine Reproduction University of Kentucky, Lexington, KY Acceptance Speech Induction to Equine Research Hall of Fame, Gluck Center, University of Kentucky, Lexington, KY Diagnostic Techniques for Evaluating Stallion Sperm Select Breeders Service Annual Meeting, Sacramento, CA Invited Talk 5th International Conference on Equine Reproductive Medicine, Leipzig, Germany Invited Talk 4th Leipzig Expertworkshop on Equine Reproductive Medicine, Leipzig, Germany Variation in Progesterone in Early Gestation after Nonsurgical Embryo Transfer AAEP Scientific Program, AAEP Annual Convention, Orlando, FL 2008 J.E. Bruemmer Current Use of Cooled and Frozen Stallion Semen Countryside Veterinary Clinic, Greeley, CO Advances in Assisted Reproductive Strategies Kentucky International Equine Summit, Lexington, KY Vitrification of Equine Embryos Asociacion Mexicano Medicos Veterinarios Especialistas en Equinos, Puebla, Mexico (presented in Spanish) Freezing Epididymal Sperm Asociacion Mexicano Medicos Veterinarios Especialistas en Equinos, Puebla, Mexico (presented in Spanish) Advanced Methods in Semen Evaluation Asociacion Mexicano Medicos Veterinarios Especialistas en Equinos, Puebla, Mexico (presented in Spanish) Case Studies: Stallion Reproduction Bluegrass Reproduction Symposium, Lexington, KY Understanding Maternal Recognition Maxwell H. Gluck Equine Research Center, Lexington, KY 70

73 E.M. Carnevale Cooling, Shipping and Cryopreservation of Equine Embryos 54th Annual Convention of American Association of Equine Practitioners, Dry Lab, San Diego, CA Clinical Considerations Regarding Assisted Reproduction in Horses West Coast Equine Reproduction Symposium III, Buellton, CA Standard and Advanced Ultrasound Procedures for Monitoring the Mare s Reproductive Tract XIV Italian Association of Equine Veterinary Practitioners, Venice Effect of Aging on Reproductive Physiology of the Mare XIV Italian Association of Equine Veterinary Practitioners, Venice Embryo Collection and Effect on the Mare s Uterus XIV Italian Association of Equine Veterinary Practitioners, Venice Cooling and Cryopreservation of Equine Embryos XIV Italian Association of Equine Veterinary Practitioners, Venice Use of Assisted Reproductive Procedures for Subfertile Mares XIV Italian Association of Equine Veterinary Practitioners, Venice How to do Induction of Ovulation XIV Italian Association of Equine Veterinary Practitioners, Venice C.M. Clay New Insights into Gonadotropin Regulation 41st Annual Meeting of the Society for the Study of Reproduction, Kona, Hawaii Estrogen Regulation of the GnRH Receptor Gene: Elusive or Illusive? 27th Annual Symposium in Reproductive Science and Women s Health, University of Kentucky, Lexington, KY Estrogen Regulation of the GnRH Receptor Department of Biomedical Sciences, Cornell University, Ithaca, NY P.M. McCue Ovarian Problems in the Mare Proveto Equine Reproduction Seminar at Zwolle International, Utrecht, The Netherlands Clinical Cases of the Non-Pregnant Mare Proveto Equine Reproduction Seminar at Zwolle International, Utrecht, The Netherlands Evaluation and Examination of the Infertile Mare Proveto Equine Reproduction Seminar at Zwolle International, Utrecht, The Netherlands Evaluation and Examination of the Infertile Mare (wet lab) Proveto Equine Reproduction Seminar at Zwolle International, Utrecht, The Netherlands Work-up of the Problem Mare: Wet Lab West Coast Equine Reproduction Symposium III, Buellton, CA 71

74 Peripartum Problems in Mares West Coast Equine Reproduction Symposium III, Buellton, CA The Problem Mare: Management Philosophy, Diagnostic Procedures, and Therapeutic Options West Coast Equine Reproduction Symposium III, Buellton, CA Embryo Collection Techniques Annual Meeting, Florida Association of Equine Practitioners, Ocala, FL Equine Embryo Transfer (wet lab) Annual Meeting, Florida Association of Equine Practitioners, Ocala, FL Evaluation of Embryo Quality (dry lab) American Association of Equine Practitioners, San Diego, CA Ovarian Problems in the Mare Oregon Veterinary Conference, Corvallis, OR Uterine Problems in the Mare Oregon Veterinary Conference, Corvallis, OR Hormonal Therapy for Problem Broodmares Oregon Veterinary Conference, Corvallis, OR Endocrine Diagnostic Tests for Problem Broodmares Oregon Veterinary Conference, Corvallis, OR New Diagnostic Tests for the Periparturient Mare Oregon Veterinary Conference, Corvallis, OR Round-Table Discussion on Reproduction Intervet Schering-Plough Animal Health, Greenwood Village, CO Foaling and Care of the Newborn Foal Cottonwood Springs Ranch, Durango, CO G.E. Seidel, Jr. Sexing Sperm Annual Meeting, National Academy of Sciences, Washington, DC Cloning Horses AQHA Annual Meeting, San Francisco, CA Current Status of Sexing Sperm Italian Embryo Transfer Society Annual Meeting, Garda Lake, Italy Cloning Mammals Joy Goodwin Distinguished Lecture, Auburn University, Auburn, AL Sexing Mammalian Sperm Auburn University, Auburn, AL Sexing Mammalian Sperm Kinki University, Wakayama, Japan Cloning Horses AQHA Annual Meeting, San Francisco, CA Cloning Mammals and Related Technology University of Illinois, Champaign, IL The National Academy of Sciences: How it functions; What it does Cell and Molecular Biology Program, Colorado State University, Fort Collins, CO Sexing Sperm Biomedical Student Association, Colorado State University, Fort Collins, CO 72

75 2009 J.E. Bruemmer Foaling Seminar Ted Robinson Ranch, Oak View, CA Basic Molecular Biology: Terminology and Techniques American Quarter Horse Association Grant Review Board, Amarillo, TX Current Understanding of Maternal Recognition in the Mare Maxwell Gluck Equine Research Center, University of Kentucky, Lexington, KY Reproductive Techniques in Horse University of Wyoming E.M. Carnevale Effect of Maternal Age on Assisted Reproductive Technologies in the Mare CVMBS Research Day, Colorado State University, Fort Collins, CO Foals Produced after ICSI Using Frozen, Sex-Sorted, Refrozen Sperm International Embryo Transfer Society Annual Meeting, San Diego, CA C.M. Clay Novel Actions of Steroid Hormones in the Hypothalamic-Pituitary-Ovarian Axis Annual Meeting of the Endocrine Society, Washington, DC Estrogen Regulation of the LH Surge Department of Veterinary Science, Maxwell Gluck Equine Research Center, University of Kentucky, Lexington, KY P.M. McCue How to Perform a Flush for Embryo Transfer 15th Societá Italiana Veterinari per Equini Congress, Bologna, Italy Managing Hydrops of the Fetal Membrane 15th Societá Italiana Veterinari per Equini Congress, Bologna, Italy Advanced Topics in Hormone Therapy 15th Societá Italiana Veterinari per Equini Congress, Bologna, Italy The Periparturient Mare and Newborn Foal: Review of Diagnostic Tests and Management Practices 15th Societá Italiana Veterinari per Equini Congress, Bologna, Italy Hormone Therapy: Basic and Advanced Universidad Nacional Autonoma de Mexico, Mexico City The Periparturient Mare: Problems and Management Universidad Nacional Autonoma de Mexico, Mexico City The Problem Mare: Diagnosis and Management Universidad Nacional Autonoma de Mexico, Mexico City 73

76 Saving Foals: The Veterinarian s Role in Education of Horse Owners and Foaling Attendants Universidad Nacional Autonoma de Mexico, Mexico City Ovarian Pathologies Modul Gynaecology, Biotechnology, Andrology, Vetmeduni Vienna, Austria Hormone Therapy in the Mare Modul Gynaecology, Biotechnology, Andrology, Vetmeduni Vienna, Austria Manipulation of Ovulation in Estrous Mares Modul Gynaecology, Biotechnology, Andrology, Vetmeduni Vienna, Austria Breeding the Difficult Mare Pilchuck Veterinary Hospital, Snohomish, WA The Problem Mare: Horse Owner s Guide to a Successful Season Scottsdale Arabian Horse Show, Scottsdale, AZ Foaling and Care of the Newborn Foal South Valley Large Animal Clinic, South Jordan, UT Saving Foals: A Horse Owner s Guide to a Successful Season Veterinary Teaching Hospital, CSU, Fort Collins, CO Problem Mares How to Optimize Pregnancy Rates VTH Practitioners Breakfast Meeting, Veterinary Teaching Hospital, CSU, Fort Collins, CO Breeding the Problem Mare Rocky Mountain Horse Expo, Denver, CO Foaling and Care of the Newborn Foal Littleton Large Animal Clinic, Parker, CO G.E. Seidel, Jr. Sexing Sperm Biomedical Student Association, Colorado State University, Fort Collins, CO Sexing Sperm Producer Meeting, National Center for Physiology and Animal Breeding Research, Queretaro, Mexico The Future of Research in Reproductive Physiology Producer Meeting, National Center for Physiology and Animal Breeding Research, Queretaro, Mexico Challenges, Successes, and Recommendations for Operating an Established Rancher Coalition Conference on Collaborative Conservation, Fort Collins, CO Conservation Easements Conference on Collaborative Conservation, Fort Collins, CO 2010 J.E. Bruemmer Gene Expression in the Equine Endometrium During Maternal Recognition of Pregnancy Tenth International Symposium on Equine Reproduction, Lexington, KY Postsecondary Equine Education International Equine Summit, Lexington KY 74

77 E.M. Carnevale Effect of Equine Oocyte Donor Age on Success of Oocyte Transfer and Intracytoplasmic Sperm Injection Tenth International Symposium on Equine Reproduction, Lexington, KY R.A. Ferris How to Use a Quantitative Turbidimetric Immunoassay Assay to Determine IgG Concentrations in Neonatal Foals 55th Annual Convention, American Association of Equine Practitioners Quantitation of Foal IgG 72nd CSU Annual Veterinary Conference, Fort Collins, CO Detection, Treatment, and Prevention of Endometritis in the Horse ARBL Seminar Series, Colorado State University, Fort Collins, CO P.M. McCue Hormone Therapy (Moderator) Tenth International Symposium on Equine Reproduction, Lexington, KY Embryo Transfer (Table Topics Moderator) American Association of Equine Practitioners, Baltimore, MD Embryo Recovery Procedures and Collection Success: Results of 492 Embryo Flush Attempts American Association of Equine Practitioners, Baltimore, MD Relationship Between Donor Mare Age, Semen Type, and Early Embryonic Development Society for Theriogenology, Seattle, WA New Techniques in Breeding Soundness Evaluation of the Mare The Ohio State University Equine Reproduction Conference, Columbus, OH Management of the Problem Mare: Behavioral Issues The Ohio State University Equine Reproduction Conference, Columbus, OH Management of Embryo Donors and Embryo Recovery The Ohio State University Equine Reproduction Conference, Columbus, OH Critical Care of the Newborn Foal CSU Collegiate Horsemen s Association, Fort Collins, CO Saving Foals Horses Forever, Longmont, CO G.E. Seidel, Jr. Whole Genome Selection in Cattle Using SNPs Satellite Symposium, International Embryo Transfer Society, Cordoba, Argentina Sexing Mammalian Sperm Technical University of Valencia, Valencia, Spain 75

78 Short Courses CSU Equine Reproductive Management and Artificial Insemination, Jan 12-15, 2007 Techniques for Handling and Utilizing Cooled Semen, Jan 16, 2007 Techniques for Handling and Utilizing Frozen Semen, Jan 17, 2007 Basic Equine Reproductive Ultrasonography, Oct 1, 2007 Equine Embryo Transfer, Oct 2-4, 2007 Annual Reproduction Symposium, Oct 5-6, 2007 Equine Reproductive Management and Artificial Insemination, Nov 1-4, 2007 Techniques for Handling and Utilizing Cooled Semen, Nov 5, 2007 Techniques for Handling and Utilizing Frozen Semen, Nov 6, 2007 Equine Reproductive Management and Artificial Insemination, Jan 10-13, 2008 Techniques for Handling and Utilizing Cooled Semen, Jan 14, 2008 Techniques for Handling and Utilizing Frozen Semen, Jan 15, 2008 Diagnostic Imaging in Equine Reproduction, Sept 25, 2008 Annual Reproduction Symposium Mare and Stallion Reproductive Behavior, Sept 26-27, 2008 Equine Embryo Transfer, Sept 28-30, 2008 Equine Reproductive Management and Artificial Insemination, Oct 30-Nov 2, 2008 Techniques for Handling and Utilizing Transported Cooled Equine Spermatozoa, Nov 3, 2008 Techniques for Handling and Utilizing Transported Frozen Equine Spermatozoa, Nov 4, 2008 Equine Reproductive Management and Artificial Insemination, Jan 8-11, 2009 Techniques for Handling and Utilizing Transported Cooled Equine Spermatozoa, Jan 12, 2009 Techniques for Handling and Utilizing Transported Frozen Equine Spermatozoa, Jan 13, 2009 Foaling and Care of the Newborn Foal, Jan 16-17, 2009 Equine Oocyte Collection and Handling, Sept 26, 2009 Equine Embryo Transfer, Sept 27, 2009 Equine Reproductive Management and Artificial Insemination, Nov 5-8, 2009 Techniques for Handling and Utilizing Transported Cooled Equine Spermatozoa, Nov 9, 2009 Techniques for Handling and Utilizing Frozen Equine Spermatozoa, Nov 10, 2009 Techniques for Handling and Utilizing Transported Frozen Equine Spermatozoa, Nov 13,

79 Equine Reproductive Management and Artificial Insemination, Jan 16-19, 2010 Techniques for Handling and Utilizing Transported Cooled Equine Spermatozoa, Jan 20, 2010 Techniques for Handling and Utilizing Frozen Equine Spermatozoa, Jan 21, 2010 Foaling and Care of the Newborn Foal, Jan 30-31, 2010 Equine Embryo Transfer, Sept 24-25, 2010 The Problem Mare, Sept 26-27, 2010 Reproductive Management and Artificial Insemination, Nov 4-7, 2010 Techniques for Handling and Utilizing Cooled Semen, Nov 8, 2010 Techniques for Handling and Utilizing Frozen Semen, Nov 9, 2010 Foaling and Care of the Newborn Foal, Jan 9-10, 2011 Reproductive Management and Artificial Insemination, Jan 12-15, 2011 Techniques for Handling and Utilizing Cooled Semen, Jan 16, 2011 Techniques for Handling and Utilizing Frozen Semen, Jan 17,

80 Visiting Scientists 2007 Maria Augusta Alonso Brazil Catalina Castaneira Veterinarian and PhD candidate, Argentina Ants Kavak Veterinarian, Estonian University of Life Sciences Dr. Marcello Veterinarian, Brazil Phillip Mendoza 4th-year medical student, University of Colorado Health Sciences Center Roberto Sanches Veterinarian, Germany Sophie Sitters Utrecht, The Netherlands Jennifer Winkelried New Jersey 2008 Kelly Campbell Australia Thais Marques Granato Brazil Annett Heise Oonderstepoort, South Africa Jimena Mugica Mexico Kathleen Mullen Cornell University, Ithaca, New York Renata Cristina Uliani Brazil Jorris Verver Utrecht, The Netherlands Leila Zekkour Mexico 2009 Ester Bloemen Wageningen, The Netherlands Sergio Luiz da Silveira Camargo, Jr. Veterinarian, Brazil Elizabeth Johnson Cornell University, Ithaca, NY Lisa Maclellan Australia Daniele Mendes Brazil 78

81 2010 Fernanda Jordao Affonso Brazil Ina Carrick Australia Hermione Gabriella Finch Scotland Sara Fleck Liverpool, England Guadalupe Adelia Garcia Paredes Mexico Thais Marques Granato Brazil Elise Kelly SUNY, Oneonta, NY Lisa Maclellan Australia Jessica Rivera Costa Rica Venancio Orozco Rogers Mexico Pamela Peters Vienna, Austria Brennan Pitard Arkansas Julhiano Rossini Veterinarian, Brazil Elena Ruggeri Italy Ruud Vullers Wageningen, The Netherlands 79

82 Past Graduate Students / Placement Name Degree Year Current Position James Cranwell MS 1970 Unknown Arthur Roberts MS 1970 Unknown E. Wayne Anderson MS 1971 Retired, Fort Collins, CO Whit Byers MS 1971 Director, Select Breeders Services, Aubrey, TX Merlin Gebauer PhD 1973 David G. Back MS 1974 Farm & Breeding Manager, Kristull Ranch, Bowling, KY Leslie D. Burwash MS 1974 Manager of Horse Program, Animal Industry Foundation, Alberta Agriculture, Food and Rural Development, Airdrie, Alberta, Canada Diane S. Demick MS 1975 Unknown Carl Rugg MS 1976 Unknown Jerry Billquist MS 1977 Veterinarian, Boerne, TX Ron Wallace MS 1977 Consulting Service, Lexington, KY Jana Hoyer MS 1978 Veterinarian, Crawford Animal Hospital, Greenfield, WI Larry Johnson PhD 1978 Professor, Dept of Veterinary Anatomy and Public Health, Texas A&M University, College Station, TX Win Stevens MS 1978 Veterinarian, Millbrook, NY Donald L. Thompson, Jr. PhD 1978 Professor, Dept of Animal Science, Louisiana State University, Baton Rouge, LA Dave McGlothlin MS 1980 Horse Division Manager, Harris Farm, Coalinga, CA Gail Robb-Todter MS 1980 Owner, Integrative Health Services, Charlottesville, VA Dennis Eikenberry MS 1981 Owner, Sportack Equestrian Products, Park City, UT Pete Heesemann MS 1981 Stock Broker, Goshen, KY Kathy Imel MS 1981 Public Safety Communications Consultant and Artist, Westminster, CO Paul R. Loomis MS 1982 Owner, Select Breeders Services Inc., Colora, MD Dave Monroe MS 1982 Breeding Farm Manager Matt Iuliano MS 1983 Vice President for Registration Services, The Jockey Club, Lexington, KY James Maher MS 1983 Sculptor, Belle Fourche, SD Jim Neil MS 1983 Breeding Farm Manager, Kerrville, TX Colin Clay MS 1984 Professor and Head, Dept. of Biomedical Sciences/ ARBL, Colorado State University, Fort Collins, CO Jeffrey D. Cochran MS 1984 Ultrasound Technician, Bear Lake Memorial Hospital, Montpelier, ID Terry T. Olar MS 1984 Director, Center for Reproductive Studies, Christ Hospital, Cincinnati, OH 80

83 Name Degree Year Current Position Neville Slade MS 1984 Instructor and Dept. Chair, Agriculture and Natural Resources Dept., Victor Valley College, Apple Valley, CA Martin J. Cristanelli MS 1985 Veterinarian, Pilot Point, TX Pam (Hart) Silvia MS 1985 Student Affairs Officer, University of Kentucky, Versailles, KY Marcelo Villahoz MS 1985 Vice President of Global Business Development, BioShield Technologies, Inc., Norcross, GA Karla Clark MS 1986 Veterinarian Gail Colbern MS 1986 Director of Biopharmacology, CellGenesys, Inc., South San Francisco, CA Jim Kubiak MS 1987 Owner, The Breeder s Assistant Equine Reproduction Schools & Services, Ramona, CA Robert Morris MS 1987 Breeding Farm Manager Greg Wiepz MS 1987 Associate Scientist, Dept. of Biomolecular Chemistry, University of Wisconsin, Madison, WI Karen Baucus MS 1988 Breeding Manager, Amarillo, TX Edward Blach MS 1988 Owner, Dr. Ed, Inc., Business Development Consulting, Monument, CO Elaine Carnevale MS 1988 Associate Professor, Dept. of Biomedical Sciences/ ARBL, Colorado State University, Fort Collins, CO Colin Clay PhD 1988 Professor and Head, Dept. of Biomedical Sciences/ ARBL, Colorado State University, Fort Collins, CO Joan (Naden) Clay MS 1988 Dressage Trainer, Stone Gate Ranch, Fort Collins, CO David Frantz MS 1988 Swine AI Technician, OK Craig Shoemaker MS 1988 Equine Professional Services, IDEXX Pharmaceuticals, Inc., Nampa, ID Kent Barnes MS 1989 Stallion Manager, Shadwell Farms, Lexington, KY Jack Clayton MS 1989 X-Ray Technician, Ventura, CA Becky Terrell MS 1989 Instructor, Equine Science Program, North Central Texas College, Gainesville, TX Ben Blue MS 1990 Veterinarian, Wellington, FL Leslie Harrison MS 1990 Veterinarian, H&H Performance Horses, Colorado Springs, CO Melinda (Hermenet) MS 1990 Veterinary Associate, Santa Anita Park, Arcadia, CA Blue Lynn Ionata-Skutches MS 1990 Chemistry Teacher, Sonora High School, Sonora, CA Paulette (Long) Hope MS 1990 Research Associate, Ethicon Endo-Surgery, San Francisco, CA Hallie Sue Rowley MS 1990 Owner, Efficient Arena, Inc., Canby, OR Tina Anderson MS 1991 Equine Specialist, Purina Mills, Fort Worth, TX Hilda Baisel MS 1991 Veterinarian, Oakdale, CA Nick Carney MS 1991 Veterinarian, Dixon, CA 81

84 Name Degree Year Current Position Vito Del Vento MS 1991 Equine Veterinarian, Apex Equine, Paeonian Springs, VA Kevin Dippert MS 1991 Owner, Equine Reproduction Concepts, Amissville, VA JeanPatrick R. Kayser MS 1991 Associate Professor, Agribusiness and Agriscience, Middle Tennessee State Univ., Murfreesboro, TN Meg (Farlin) Anema MS 1992 Equine Studies Program Director, Arizona Agribusiness & Equine Center High Schools, Phoenix, AZ Anne Francl-Mask MS 1992 Breeding Farm Manager, Kapoa, HI Diane Moran MS 1992 Breeding Manager, ND Valentine (Shabpareh) Williams MS 1992 Equine Manager, Fort Dodge Animal Health, Overland Park, KS Stacy Truelson MS 1992 Unknown Sylvia Bedford-Guaus MS 1993 Assistant Professor of Theriogenology, Dept. of Clinical Sciences, Cornell University, Ithaca, NY John Knowles MS 1993 Veterinarian, Pioneer Equine Hospital, Oakdale, CA Jennifer Phillips MS 1993 Unknown Amy Heitland MS 1994 Pharmaceutical Sales Representative, Wyeth, Colorado Springs, CO Liz Mumford MS 1994 Veterinarian, Global Influenza Programme, World Health Organization, Geneva, Switzerland Kristine Wilhelm MS 1994 Owner, Edgewood Meadow Farm, Fleetwood, PA Sandy Badzinski MS 1995 Client Services Director, Crawford Animal Hospital, Milwaukee, WI Kevin Dippert PhD 1995 Owner, Equine Reproduction Concepts, Amissville, VA Joel Brubaker MS 1996 Veterinarian, Falcon Veterinary Services, Peyton, CO Nancy Cook MS 1996 Veterinarian & Owner, Advanced Equine Reproduction, Steamboat Springs, CO Susan (Leipold) Johnson MS 1996 Veterinarian, Buck Hill Veterinary Service, Spencer, MA Bradley Ray MS 1996 Owner/Manager, Premier Breeding Services, LLC, Kiowa, CO Tom Sissener MS 1996 Veterinarian, CARE Centre Animal Hospital, Calgary, Alberta, Canada Steve Van Wagner MS 1996 Veterinarian, New Waverly Veterinary Clinic, New Waverly, TX David Denniston MS 1997 Associate Professor, Dept. of Animal Sciences, Colorado State University, Fort Collins, CO Kitren Nickerson MS 1997 Veterinarian, Mesa Veterinary Hospital, Golden, CO Chrissy Young MS 1997 Research Technician, Memphis, TN SaTrice Coleman MS 1998 Unknown Sarah Keith MS 1998 Unknown Patricia Salazar MS 1998 Veterinarian 82

85 Name Degree Year Current Position Rebecca Schmid MS 1998 Veterinarian, Delta, Colorado Bryan Buchanan MS 1999 Veterinarian, Weatherford, TX Lilian Franz MS 1999 Product Development Specialist, Fort Dodge Animal Health, Princeton, NJ Anne Hurtt MS 1999 IVF Technician, Weatherford, TX Ricardo Ramirez MS 1999 Veterinarian Ryan Coy MS 1999 Veterinarian, Royal Vista Southwest, Purcell, OK Erin (Crockett) Bishop MS 2000 Breeding Technician, Select Breeders Southwest, Inc., Aubrey, TX Allison Lindsey MS 2000 Instructor, Equine Studies, Laramie County Community College, Cheyenne, WY Nicole Oberstein MS 2000 Breeding Farm Manager, Select Breeders, Europe, Murg, Germany Mary O Donovan MS 2000 Veterinarian, Weathervane Farm, Middlebury, VT Thomas J. Scott MS 2000 Supervising Public Health Veterinarian, USDA and Owner Tom Scott Veterinary Services, Equine Ambulatory Practice, Nampa, ID Valerie (Farquhar) Linse MS 2001 Veterinarian, Hagyard-Davidson-McGee, Lexington, KY Bob Foglia MS 2001 Veterinarian, Royal Vista Southwest, Purcell, OK Emily Kirk MS 2001 Unknown Charles Scoggin MS 2001 Resident Veterinarian, Claiborne Farm, Lexington, KY Marco Antonio Coutinho da Silva MS 2002 Assistant Professor of Theriogenology, Dept. of Veterinary Clinical Sciences, The Ohio State University, Columbus, OH Lilian Franz PhD 2002 Product Development Specialist, Fort Dodge Animal Health, Princeton, NJ Brielle (Hecht) Weldon MS 2002 Veterinarian, Indian Tree Animal Hospital, Arvada, CO Heather Reger MS 2002 Veterinarian, Midway, UT Tawna Backman MS 2003 Veterinarian, Equine Medical Services, Columbia, MO Luke Bass MS 2003 Veterinarian, Pioneer Equine Hospital, Oakdale, CA Wendy Eldridge- MS 2003 Owner, MW Equine LLC, Pamplin, VA Panuska Brett Glazar MS 2003 MEA Laboratory Manager, VitroLife, Inc., Englewood, CO Karly Macon MS 2003 Veterinarian, Cranbrook Veterinary Hospital, Cranbrook, British Columbia, Canada Paula Moffett MS 2003 Research Associate, Equine Reproduction Laboratory, Colorado State University, Fort Collins, CO Amanda (Moore) Glazar MS 2003 Director, Adult Learning & Outcomes, Global Education Group, Littleton, CO 83

86 Name Degree Year Current Position Kimberly Preis MS 2003 Senior Embryologist, Colorado Center for Reproductive Medicine, Lone Tree, CO Kira Zerkel MS 2003 Veterinarian, Anchorage, AK Sara Lindbloom MS 2004 Research Technician, Biotech Company, Kansas Kory Niswender MS 2004 Veterinarian, ESMS on the Brazos, Weatherford, TX Chris Wilson MS 2004 Embryologist, HRC Fertility, California Amanda Beisner MS 2005 Barn Manager, Montana Marco Antonio Coutinho da Silva PhD 2005 Assistant Professor of Theriogenology, Dept. of Veterinary Clinical Sciences, The Ohio State University, Columbus, OH Quinn Hardy MS 2005 Broker, Billings, MT Jay Hudson MS 2005 Veterinarian, Lazy H Large Animal Services, Gillette, WY Amanda (Moore) Glazar PhD 2005 Director, Adult Learning & Outcomes, Global Education Group, Littleton, CO Teresa Slough PhD 2005 Assistant Professor, Animal Sciences and Industry, Kansas State University, Manhattan, KS Susan (Welch) Mc- Cutcheon MS 2005 Pharmaceutical Sales, Precision Pharmacy, Weatherford, TX Nichole Logan MS 2006 Veterinarian, Dakota Hills Veterinary Clinic, Rapid City, SD Beth Spizziri MS 2006 Teaching Instructor, Dept. Biology, Delta College, Saginaw, MI Timothy Cox MS 2007 DVM candidate, Colorado State University, Fort Collins, CO Christianne Magee MS 2007 Postdoctoral Fellow, ARBL/Biomedical Sciences, Colorado State University, Fort Collins, CO Melinda Meyers MS 2007 Melissa Hudson MS 2008 DVM candidate, Colorado State University, Fort Collins, CO Joy Altermatt MS 2009 Veterinarian and Clinician, Equine Reproduction Innovations, Inc., Wellington, CO Rebecca Bott PhD 2009 Assistant Professor, Equine Management & Reproduction, South Dakota State Univ., Brookings, SD Fernando Campos- Chillon PhD 2009 Veterinarian and Clinical Manager, Equine Reproduction Innovations, Inc., Wellington, CO Catherine DeLuca MS 2009 Veterinarian, Buckeystown Veterinary Hospital, Buckeystown, MD Gretchen Lund MS 2009 DVM candidate, Colorado State University, Fort Collins, CO Rebecca Yourey MS 2009 DVM candidate, Colorado State University, Fort Collins, CO Brad Daigneault MS 2010 PhD candidate, Dept of Animal Sciences, University of Illinois at Urbana-Champaign, Urbana, IL 84

87 Name Degree Year Current Position Bethany Frank-Guest MS 2010 Veterinarian, Baronne Veterinary Clinic, Sunset, LA Ellane Cleys MS 2010 PhD student, ARBL/Biomedical Sciences, Colorado State University, Fort Collins, CO Ryan Brooks MS 2010 Facilities and Breeding Manager, Middleburg Agricultural Research and Extension Center, Virginia Tech, Blacksburg, VA Bernardo Rodrigues MS 2010 Veterinarian, Shogun and Aldedarã Equine Embryo Transfer Farms, Goiás, Brazil Erika Cullingford MS 2010 Junior Embryologist, ART Institute of Washington, Washington, DC Ryan Ferris MS 2010 Clinical Instructor, Dept of Clinical Sciences, Colorado State University, Fort Collins, CO Beth (Spizziri) Roznowski PhD 2010 Teaching Instructor, Dept. Biology, Delta College, Saginaw, MI 85

88 Animal Reproduction and Biotechnology Laboratory COLLEGE OF VETERINARY MEDICINE AND BIOMEDICAL SCIENCES Sex, Science, and Survival in Academe A History of the Animal Reproduction and Biotechnology Laboratory at Colorado State University B.W. Pickett and Colleagues July 2011 Coming Soon The Animal Reproduction and Biotechnology Laboratory at Colorado State University has been around, in one form or another, since the early 1940s. In that time, ARBL has made many groundbreaking discoveries and seen many momentous events. After years of combing the archives, we re proud to offer Sex, Science, and Survival in Academe, the complete ARBL history book. The book was written by longtime CSU professor, researcher, and administrator, B.W. Pickett, along with several colleagues to document the history of an important phase of animal reproductive research, teaching, outreach, philosophy, and service; and the benefit of those activities to the livestock industries and public that we serve. Please call (970) or visit edu/bms/arbl for information on ordering your copy of this book. 86