Clinical Records NONINVASIVE METHOD OF SEMEN COLLECTION FOR SUCCESSFUL ARTIFICIAL INSEMINATION IN A CASE OF RETROGRADE EJACULATION

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1 FERTILITY AND STERILITY Copyright The American Fertility Society Vol. 36, No.2, August 1981 Printed in U.s A. NONINVASIVE METHOD OF SEMEN COLLECTION FOR SUCCESSFUL ARTIFICIAL INSEMINATION IN A CASE OF RETROGRADE EJACULATION MAHENDRAN MAHADEVAN, B.V.Sc. JOHN F. LEETON, M.B., B.S., F.R.C.O.G.* ALAN O. TROUNSON, B.Sc., M.Sc., PH.D. Department of Obstetrics and Gynaecology, Monash Medical School, Queen Victoria Medical Centre, Melbourne, Victoria 3000, Australia In retrograde ejaculation, semen flows into the bladder because of failure of the bladder neck to close. This defect may be caused by trauma, inflammation, senescence, damage to the sympathetic nerves of the bladder, or malfunction ofthe posterior urethra. The incidence of male infertility due to retrograde ejaculation is reported to be increasing.1 The pathophysiology of retrograde ejaculation and successful therapeutic methods have been reported by Barwin et al./ Fischer and Coats,2 Hotchkiss et al.,a and Glezerman et al. 4 Success of pharmacologic treatment reportedly is low.1, 5, 6 Antegrade semen collection procedures involving ejaculation with a full bladder 6 or defecation following ejaculation have been suggested for treatment of patients with retrograde ejaculation. 7 Although these methods of treatment can be useful in some individuals, they are not reliable and cannot be used in all cases. For patients with severe damage to the bladder neck, ejaculation with a full bladder is contraindicated. However, many pregnancies have been achieved by retrieval of spermatozoa from the bladder after masturbation or intercourse. Two methods currently are used to recover spermatozoa from th bladder: (1) In the invasive method, the urine is replaced with a few milliliters of an isotonic buffer solution by catheterization before masturbation. The patient is again catheterized to recover the sperm suspension, and his wife is inseminated with the sperm obtained. a (2) In the noninvasive method, the urine is neutralized by Received December 29, 1980; revised and accepted February 20, *Reprint requests: Dr. John F. Leeton, Department of Obstetrics and Gynaecology, Monash Medical School, Queen Victoria Medical Centre, Melbourne, Victoria 3000, Australia. 243 the oral ingestion of alkalinizing agents (NaHCOa) because the normally acidic urine is considered spermicidal. The urine voided after masturbation is centrifuged and the wife is inseminated with the sperm recovered.2 The invasive method of Hotchkiss et ala is not acceptable to most patients because of the psychologic stress and the pain involved in the catheterization procedure, which may also result in severe urethral infection.2, 4 The noninvasive method of Fischer and Coats2 has been reported to be unreliable because of the variable quality of semen retrieved. Repeated inseminations for many cycles are required to achieve a pregnancy.1, 5, 6 It was suggested that the reduction of motility of the sperm obtained by noninvasive methods was due to their exposure to the excessively high osmolarity of urine. 6 We report the development of a noninvasive method for adjusting the osmolarity of the urine present in the bladder at the time of ejaculation, so that high-quality sperm may be retrieved repeatedly. Clinical Records METHODS AND RESULTS A 29-year-old man and his 26-year-old wife were first evaluated in 1978 because of primary infertility of 2 years' duration. Results of routine infertility investigations of the wife were normal, but the husband had never experienced antegrade ejaculation. He had a complicated history of pediatric bladder surgery and had suffered recurrent bouts of urinary infection in adult life. Panendoscopy and retrograde pyelography revealed a large subtrigonal bladder diverticulum with an obstructed atrophic pyelonephrotic left kidney. A

2 244 COMMUNICATIONS-IN-BRIEF August 1981 TABLE 1. Development of the Semen Collection Method for a Patient with Retrograde Ejaculation Osmolarity of urine Date (1980) Comments on semen collection Motility Sperm concentration Before ejaculation After ejaculation March May 27 Trial semen collection by method of Fischer and Coats 2 Patient 500 ml of water 3 hr before semen collection June 3 As above but patient drank 8 water only 1 hr before semen collection; urine osmolarity checked before semen collection June Repeated as above 145 Osmolarity checked after min June 17 July 17 August 14 asee Figure 2. Change in osmolarity determined after patient drank 300 ml of water" First cycle of insemination First collection 312 Next collection 80 After 30 min 280 Second cycle of insemination First collection 285 Next collection 207 mosmoleslkg water % millionslml left nephro-ureterectomy excision of the subtrigonal bladder diverticulum and reconstruction of the bladder neck were carried out in 1978 that resulted in the production of sterile urine, but the problem of retrograde ejaculation remained. Because of the complicated nature ofthese congenital and acquired urinary abnormalities and the fact that surgery had been carried out on two occasions, another operation specifically aimed at correcting the patient's ejaculation problem was considered unjustified, especially as narrowing of the internal meatus could complicate his otherwise now normally functioning urinary tract. It was therefore decided to attempt sperm recovery from the bladder with the noninvasive method so that the risk of additional urinary infections was avoided. Development of the Semen Collection Method Semen was recovered from the patient's bladder according to a modified method of Fischer and Coats. 2 The volume, ph, and osmolarity of the urine specimens were recorded. The percentage motility and the sperm concentration of the semen sample were determined according to the methods described by Eliasson. 7 Adjustment of Urine Osmolarity. As shown in Table 1 and Figure 1, the quality of the semen recovered was poor when the osmolarity of the urine deviated from the normal osmolarity ofhuman semen. The osmolarity of normal human seminal plasma has been reported to be 366 ± 16 mosmoles/kg.8 This finding confirms the suggestion by Crich and Jequier 6 that the quality of the semen recovered from the bladder depends upon the osmolarity of the urine. The procedure for neutralization of urine resulted in ph in the range of 6.7 to 7.4. It was considered that human spermatozoa could survive well in this ph range. The osmolarity of the urine voided just before ejaculation was also measured in order to predict the osmolarity of the urine in the bladder at the OSMOLARITY (m. OSMOl.ES/Kg of WATER) FIG. 1. The effect of urine osmolarity on the percentage motility of human spermatozoa.

3 Vol. 36, No.2 COMMUNICATIONS-IN-BRIEF !oJ :i 800 o tio 700 Ii '" ::: INGESTlml OF WATER I 000 ml) TIME IN HOURS :: I 60 s )0 FIG. 2. The effect of drinking 300 ml of water on the osmolarity and volume of urine voided every 30 minutes. time of ejaculation. It was found that, 3 hours after the ingestion of 500 ml of water, the osmolarity was still high (Table 1, May 27), and after 1 hour it was too low (Table 1, June 6 and June ). To find the rate of change in osmolarity of the urine, the patient was asked to void 30 minutes after first voidance (Table 1, June ). The rate of change in the patient's urine osmolarity was 340 mosmoleslhour, but it was noted that this value could vary depending on the water balance of the body. A semen sample with acceptable motility (40%) was recovered when the osmolarity was 316 mosmoleslkg. On the basis of these findings and the basic physiology of water regulation by the kidney and the bladder, we hypothesized that after the ingestion of water the osmolarity of urine in the bladder would initially decrease, owing to increased water excretion (dilution phase), and then increase because of the concentrating effect of the bladder (concentration phase). To verify this hypothesis, a volunteer agreed to void every 30 minutes following the ingestion of 300 ml of water. The osmolarity and the volume ofthe voided urine were recorded and are shown in Figure 2. The decrease in the osmolarity of the urine with a corresponding increase in the volume of the urine voided confirmed our hypothesis. The rate of change in osmolarity during the arbitrary dilution phase (1st hour after ingestion of water) was very high (approximately 540 mosmoles/kg). However, the rate of change during the concentration phase (next hour) was very similar to the value found for the patient (about 340 mosmolesl kg). Usually about 15 to 30 minutes are required for ejaculation to occur either by masturbation or intercourse. It was considered that semen was best collected during the concentration phase and when the osmolarity of the voided urine was approximately 200 to 300 mosmoles/kg. The patient 20 was advised not to delay ejaculation any longer than necessary. Semen Collection and Insemination. Using the following semen collection and insemination protocol, two inseminations were attempted during the wife's first ovulatory cycle. The day of expected ovulation was determined from previous temperature charts and the average length of the menstrual cycle. From about 5 days before the expected day of ovulation, the rate of excretion of urinary luteinizing hormone (LH) was determined each morning. The time of ovulation was predicted from the start of the LH increase and the LH peak as described by Trounson et al. 9 The husband was asked to abstain from ejaculation for at least 3 days before the expected day of ovulation to increase the sperm concentration in his semen, to take alkalinizing tablets (4 gm of NaHC0 3 ), four times daily, and to avoid ingesting toxic drugs and alcohol. On the day of the LH increase (Fig. 3), the husband was asked to void and then to drink about 500 ml of water, 1 hour before ejaculation. The husband again voided completely just before the semen collection to determine whether the urine osmolarity was in the range of 200 to 300 mosmoleslkg. When the osmolarity was low, it was checked again after 15 to 20 minutes; if it was high, the patient was asked to drink about 200 ml of water and the osmolarity was rechecked. When the osmolarity was within the recommended range, the husband was asked to ejacu- - ::0 "',0.\l!{! 30 :r:! 20 '-l :/. - -:: z - :r: :.:...J AIH DAY OF CYCLE FIG. 3. Timing of ovulation using urinary LH excretion rates. AIH, artificial insemination with the husband's semen.

4 246 COMMUNICATIONS-IN-BRIEF August 1981 late (by intercourse in a room. near the laboratory) without unnecessary delay. Urine was voided completely immediately after ejaculation (within minutes). The volume of urine voided was in the range of 15 to 50 ml. The urine was immediately dispensed into -ml sterile plastic centrifuge tubes and centrifuged at 500 x g for 5 minutes. The osmolarity of the supernatant urine was checked and the remainder was discarded. The sperm sediment was resuspended in 0.25 to 0.5 ml of Ham's FlO medium (Flow Laboratories, Rockville, Md.) containing human serum albumin, 4 mg/ml (Sigma Chemical Co., St. Louis, Mo.). The sperm suspension from all the tubes were pooled, mixed gently, and inseminated immediately after removing an aliquot for semen analysis. Approximately 0.3 ml of the sperm suspension was placed intrauterine and the balance was placed intracervical and pericervical. The same procedures were repeated the next day (Fig. 3). For the first cycle of insemination it was found that the quality of semen recovered was not good, although acceptable (Table 1). This may have been due to the high osmolarity of the urine after ejaculation (560- mosmoles/kg). We speculated that, because of a negative water balance in the patient, his bladder had concentrated the urine at a faster rate (about 520 mosmoles/kg) than expected (340 mosmoleslkg). For the next insemination cycle the husband was asked to drink as much fluid (water, soft drinks, tea or coffee) as possible every 3 hours from 5 days before the expected day of ovulation. Semen of good quality was recovered on both days when insemination was attempted during the second cycle (Table 1), and a pregnancy resulted. The pregnancy has progressed satisfactorily to date. DISCUSSION It is recommended that for patients showing a very fast rate of change in osmolarity, as was found for our patient during the first insemination cycle, the osmolarity of the urine just before ejaculation should be in the lower range of the recommended values (about 200 mosmoleslkg). Alternatively, just before semen collection it may be worthwhile for the husband to drink an additional 200 ml of water to lower the rate of increase in osmolarity. It is possible that the incidence of retrograde ejaculation is more prevalent than has been reported. Retrograde ejaculation is usually suspected if no semen is ejaculated (aspermia). However, azoospermia and hypospermia have been reported to occur when part of the ejaculate is retrograde and part antegrade. When the semen arrives at the prostatic portion of the urethra, the bladder neck is closed by contraction of the internal urethral sphincter under sympathetic control. Closure of the bladder neck prevents the semen from flowing back into the bladder.l, 4,11 If the internal urethral sphincter is slow to contract or constricts only partially upon the arrival of the semen a portion of the ejaculate will pass into the bladder, resulting in hypospermia. Moreover, it is well known that the first portion of an ejaculate usually has a higher sperm concentration. Therefore, if a greater portion of the initial fraction of the ejaculate passes into the bladder, a reduction in sperm concentration may result. Barwin et au and Kapetanakis et al. 5 have reported pregnancies following insemination of semen recovered from the bladder and cryopreserved. Cryopreservation of the recovered semen was said to be necessary because recovery of goodquality semen at the time of ovulation was unpredictable. As reported here, a method is now available for the consistent recovery of semen of acceptable quality. Cryopreservation is not essential and is probably contraindicated, since the lower-quality semen usually recovered from a patient with retrograde ejaculation will be further lowered by freezing and thawing. Acknowledgments. We wish to thank Miss Barbara Beech (insemination), Miss Mandy Besanko (radioimmunoassay), Mr. Warren Shepherd (medical illustration), and Miss Mandy McLaughlan (typing). REFERENCES 1. Barwin BN, McKay D, Jolly EE, Dempsey A: Retrograde ejaculation. In Homologous Artificial Insemination (AIH), Edited by JC Emperaire, A Audebert, ESE Hafez. London, Martinus Nijhoff, 1980, p Fischer IC, Coats EG: Sterility due to retrograde ejaculation of semen. Obstet Gynecol 4:352, Hotchkiss RS, Pinto AB, Kleegman S: Artificial insemination with semen recovered from the bladder. Fertil Steril 6:37, Glezerman M, Lunenfeld B, Potashnik G, Oelsner G, Beer R: Retrograde ejaculation: pathophysiologic aspects and report of two successfully treated cases. Fertil Steril 27:796, Kapetanakis E, Rao R, Dmowski WP, Scommegna A: Conception following insemination with a freeze-preserved retrograde ejaculate. Fertil Steril 29:360, 1978

5 Vol. 36, No.2 COMMUNICATIONS-IN-BRIEF Crich JP, Jequier AM: Infertility in men with retrograde ejaculation: the action of urine on sperm motility, and a simple method for achieving antegrade ejaculation. Fertil Steril 30:572, Eliasson R: Standards in evaluation of human semen. Andrologie 3:49, Valazquez A, Pedron N, Delgado NM, Rosado A: Osmolality and conductance of normal and abnormal human seminal plasma. Int J Fertil 22:92, Trounson A, Herreros M, Burger H, Clarke I: The precise detection of ovulation using a rapid radio-immunoassay of urinary LH. Proc Endocr Soc Aust 23:75, Keiserman WM, Dubin L, Amelar RD: A new type of retrograde ejaculation: report of three cases. Fertil Steril 25:71, Potts IF: The mechanism of ejaculation. Med J Aust 1:495,1957