BDWIH PERRY SARTII1. for t!» degree MASTER OP SCIEHCE. Department of Zoology KAHSAS STATE COLLEGE. A» Tim, Cornell College, t.?

Transcription

1 )P SEX HOHHOPISS OH.TIBER ASD IJBMOOLOBIH COHCElJTRATIOff OP WHIT. IOBH CIHCKESS by BDWIH PERRY SARTII A» Tm, Cornell College, t.?e»non, Iowa, 347 A THESIS sub:;tted In partal fulfllment of tba roqurexaents for t!» degree MASTER OP SCIEHCE Department of Zoology KAHSAS STATE COLLEGE OP AGRICULTURE AND APPLIED SCIE3CB 948

2 LC ~J OP COOT" W# c, 3^ DOGTXffl... -» ««PROCEDTJBB* *? 5 HES;JLTS 6 COSCLUSIOlS. - M DISCU35I0H. -.. H SUHKMr/.... M ACKKOmjSDOMSKT» S3 LITERATURE CITED»«. H

3 XKTRODOCTIQR For aany years bologsts have known that male nwnreels and brd* had a consstently hgher erythrocyte count and a greater haaoglobn concentraton In the blood than fenale audaals of the axae speces* A thorough search of the avalable lterature faled to dsclose the frst report of ths dfference by an expermental bologst, however, an etologcal relatonshp between ths dfference n the blood pcture and the sex glands was reported by several workers. In 928 Bluoel, a orraan zoologst, found that the erythrocyte count of xaale rata dropped sharply aftor castraton* lurtltor evdence t)at the sex glands exorcsed an nfluence on the blood pcture was reported by Juhn and Doom (930), w:o found that the red blood cell counts n sexually Esoature chckens dd not show a sex based dfference, but that a harp dvergence between the sexes occurred at puberty when the male brds counts rose very rapdly. Adult nale chckens had an average count about 30 per cent hgher than the fenale average, whle castrated nale brds approached the fenale lovol. more defnte proof was offered by an Italan, JBaraanuclc tll Padoa (03), who reported that the roaoval of a hen's ovary, wth a resultng prolferaton of testcular tssue and elaboraton of testcular horoono, caused a slarp rse n the erythrocyte nutaber of the brd, r.inlar results were obtaned n nvestgatons wth pgeons (Rddle and fsraucher, 934), ( ;ddle and Gauthen, 933), rats (stenglass et al., 94), and golden hamsters (rten

4 and Jacob r, en, 94)* The purpose of ths nvestgaton was not only to chock and clarfy tlls relatonshp between the sex hormones and the blood pcture, but also to secure nformaton concernng the met od of acton of estrogenc hormone on the erythrocyte numbor and the hemoglobn concentraton* Past workers have generally agreed that androgenc hormone ncreased the erytrocyto and the hemoglobn levels, although the acton s not thoroughly understood* Before the era of synthetc testosterone, workers such as Truffl (927) reported that testcular tssue, grafted nto female anmals, produced a sudden rse n erythrocyte numbers* More recently, ncreased erythrocyte counts and hemoglobn concentratons caused by synthetc male sex hormone have been reported by Tabor et al. (94, 943) n chckens, Dona et al* (943) In chckens, Vollmer and Gordon (94), /ollmer et al* (942) n rats, lnklosten et al # (944) n rats, and McCullagh and Jones (942) n eunuchod male human bengs* Concernng te effect of estrogenc hormone, however, no such wdespread agreement exsts. Dave and Boynton (94) found that stlbostrol, a synthetc estrogenc compound, caused an Increase n t-e number of red blood cells* They attrbuted ths ncrease to a drect stmulaton of the hematopoetc system by the ostrogen* On the other hand, Tyslowlts and Bartman (94) observed a decreased orytrooyte count n doge of both sexes after the admnstraton of estrogen. They concluded, as dd Castrodale et al# (94), that the estrogen n some way nhbted the hematopoetc system* Only a alght anema wae reported by

5 3 Feuchnger (940) after treatment of female rata wth ostrogc and the anmals soon returned to ther normal level although the Injectons were contnued. le also noted a smlar drop n crytlrocyte number durn; each per'od of estrus n rats. Tabor ot al. (94* 943) and Crafts (94), workng wth aalo chckens and rats, reported consstent decreases n both erytlrocyte counts and hemoglobn concentratons after njectons of estrogenc hormone# Tljer evdence Indcated a possble antagonsm between male and female hormone nsofar as ther relaton to the blood pcture was concerned. However, usng the two hormones together on capons, Doara et al. (945) reported a resultng Increase n erythrocyte number, whle Tyslowta and Dlngemanse (94) observed a decrease n erythrocyte number resultng from the seme combnaton n castrated dogs. In lght of such dscrepances, addtonal research seened ndcated. The wrter hoped that ths experment would ad n clarfyng both the effect of estrogen on the blood pcture and ts method of acton thereon. pr m Ths experment was begun In October, 947, and contnued r*ough June, 943* All of te work was done n the laboratory of the Department of Zoolo tc^r of Kansas tate Colloge n Manhattan, Kansas. A total of 42 */hto Leghorn chckens was used as expermental anmals. They were selected from two separate shpcwnts, snce the experment was dvded nto two seres; and each brd marked by attaclng a numbered metal clp to ts left wng*

6 All brds wore weghed to the nearest gran before the experment began end agan at the concluson of the work* The frst seres conssted of 6 brds, eght capons and eght normal males. All of these brds wore receved on Hay 3, 947, and were about fve months old when the experment began* The capons were castrated by Dr* IS* U«nerrok of the Department of Zoology on July 5, 947. These 6 brds were furter subdvded nto four groups. Group conssted of four normal males and served as a control group. These brds lved wth the others, havng an Identcal envronment n every way, but they receved no hormones. Group 2 conssted of four capons whch receved njectons of male sex hormone* Pour more capons made up Group 3, and ths group receved njectons of both male and female hormones* The fourth group contaned four normal males and was treated wth estrogenc hormone* W the gocok! seres, a total of 26 brds was used* Tlrteex of tcn were capons, 0 were normal males, and three were normal females* They were receved on January 8, 943, and were about three months old when they were placed on experment* These capons were castrated on February 7, 948* The brds n ths seres were dvded nto sx groups* The control group (Group 5) ths tme contaned fve normal males* Group 6 contaned fve capons whch were treated wth male sex hormone* Group 7 also contaned fve capons, but ths group was treated wth a combnaton of male and female sex hormones. c do loraone was admnstered to Group 3, whch contaned fve normal males* Groups 9 and 0, whch also receved female sex hormone, conssted

7 of three capons and thro normal females respectvely. The expermental brds were njected wth t'o ormone naterlal three tmes a week for a perod of sx weeks* "Orotom," propared by t: rng Corporaton, was used as the male sex hormone. It conssted of testosterone proponate n a sesame ol soluton, and contaned 25 mllgrams of actvo "onaone materel per cubc ocntmoter of soluton* Each tr«weekly dose was twotenths of a cubc centmeter of Cretan or fve rdlllgrams of testosterone proponate* Each brd thus receved a total of 90 mllgress of actve oraonc. Durng the frst seres* *Theeln, n a Parke-Davs product* was used as the female sex hormone. It was used n an aqueous suspenson* wth et or one or two nlllgrama of actve hormone per cubc centmeter of water. trl-weokly dose was one mllgram of actve lormone materal and each brd rooelved a total of 3 mllgrams durng the expermental perod* or the groups recevng both hormonos tbw above doscrbed dosages were combned and gven on the tame tr-wwekly schedule. njectons were ntramusoula:. Pror to the frst njecton n each seres two erythrocyte counts and two Iemoglobln determnatons were taken on ea. Thus an ndvdual norm was establshed for each, tarng the course of the njectons, weekly erythrocyte counts and hemoglobn determnatons were made, and after the concluson of the njectons, the weekly determnatons were contnued untl the brds approxmated ther pre-njocton level* Blood for the tests was drawn from a ven on the Inner surface of a wng* For erythrocyte

8 6 counts* a one to two hundred dluton wt": ayom'o soluton was snde* and the red blood sella were counted on a Spencear Brght-lne Hemacytometer* A one to two hundred and ffty dluton wth onetenth normal hydrochlorc acd was used for the hemoglobn actor. -lnatons, and the her.; In concentraton In grams per one hundred cubc centmeters was read fron the scale of a Fsher ITLectro-hemoraater* LTS Tl»o recorded results of t.dm Invest gat Ion conssted of week* ly erythrocyte counts* weekly determnatons of hemoglobn concentraton, and the weght ncrease of each brd durlr. 3-wook expermental perods :,o attempt was made to treat those results statstcally snce the number of ndvdual brds nvolved was consdered too small for such treatment* Interpretaton of the results therefore requred consderaton of the physologcal dfferences between n als of the same speces as well as the dfferences generally found between any gven ndvdual end the so-called norm of the speces concerned* In calculatng mean data for each group of brds no allowance was nado for ndvdual devaton from the mean* but all observed results were Include. "oan weekly data for each {^roup were prepared n both graphc and tabular form and ncluded n ths report. Fgs. -0* The naxlraum varaton n or;-throcyto count* hemoglobn concentraton, and body weght of each ndvdual brd was also tabulated and added to ths report. Table,

9 . - n \ - ',! - - j r 3.5 RBO ^ ^^~ _^^ g 3.0 ft H H e o OC 2.5 J Hb. >" - a -.. m.-r*" -- «- >.- --» o 3 8 u P. w 6 J , weeks 8 Fg.. Group ; fve normal male controls. 3.5 m C o 3.0 o 2.5 3d J_ njecton perod [ ' weeks 9 8 Fg, 2. Group 2; four capons wth male hormone.

10 mllons mllons RBC ".. * -! ' ( sj\ ', -. ' I,.. ' ' :. j : ' H RBC %»* ' ]A ^ Pg. ] m - - Jy A.>--- '*> r. ~ Inject}Lon ! &»rt edj \ V 4 3 2, ^ws? * 0 4-j. -_ ! ' weeks 3. Group 3; four capons wth male and female h Drmo ass. o 8 H U to t R5. ; f 3.5 x ston perod 3.0 '- -- Bb T 2.5 b\,. \ \~ >*. ' Ny' - L_ -- y^ 4.» -»"' *»«.- -* 2 ^.-- 0 o 8 H U e P. to P ff ! Or oup 4; r ( J r 3 wee ks 2 four ma les w th female hormon 9. J 8 3

11 .... I. j I. l '...\. l j t I ' ' ',3 --> +- - ^^^ ' -h ; 4 o o : L! IV - _»«.....«... ***«.Bat * -.,.-' « T bo weeks Fg. 5. Group 5; fve normal male controls _> rl H #'» W J / L> o >* s j / PS B.O ^^, ',.-» r +"j ^ J 8CWr "Ol o * a --. V >- -! M s 8 H U 2 I m *» >J "- «0 -«.. '"\ weeks Pg. 6. Group 6; fve capons wth male hormone. 9 s

12 - -.»njeoto I :. -! r. I. J I 0! I-T- It 3.5 m > \ so g a s S-O «-> w RBO H X r - -. ^l : > X ^r»-«'»«-'^ <- X.--' «4 >«h *r 4 > k «** ^. l " *< [V 3 2 o o o U o «> 0 I j. S > t 2 e n 8 s 0 ] 2 > 3 ] weeks Fg. 7. Group 7; fve capons wth male and female hormones. " n g RBO E o 2.0 I weeks 9 8 Fg. 8. Group 8; fve males wth female hormone.

13 -. ', ' _ C 3.0! o H, -»Pfl BU 8CD 2.5 * r.lec,on perod. «'*,-«**«. V»-«*»»*... -^ o o 8 H 2 «0 to weeks 8 Fg. 9. Group 9; three capons wth female hormone ; HBO F H njecton e perod J j- p, C^- " 0 23 T weeks o 2 H Pg. 0. Group 0; three females wth female hormone. 9 8 to g

14 ,: 8 Tabc *.on- weekly data for four normal attc. /M.&.^rs : :;;.: t RBC , , u Table 2. H»tta weelly data for four capons treated wth androgenc horaone. -. t : BL , IS _ l

15 four 3 Table 3. Mean weekly data for capons treated wth androgenc and estrogenc hormones*.- : * RBC : s» G , G , * , , , G Table 4. Mean weekly data for four normal males treated wth o stroble honaono. oc.:: : : s RBC : : : m # ,0 G , , IS

16 4 Cl-.. data for fve nornal male controls* 3 RBC t lb » « * C * C, C C. Table G. Moan weekly data for ffcl capons treated wth androgenc homono. cc;: : RBC lb * * G

17 - : 5 Table 7, Hean weekly date for fve capons treated wth snc and estrogenc honaonea. t RBC nb o, * a G Table 3, Mean weekly data for fve norml aalea treated wth estrogenc* hornone. :::-; :» a I HBC : 3 : r , , IS H

18 6 Table Mean weoltly data for treated wth estrogenc a,.. ; : SBC : O00 0, Table 0* aan weekly data for tlree feroalcs treated wth estrogenc.o".: : BBC t m G :

19 : ES - 7 Table. Data for ndvdual brds on changes In RBC* Sb* M weght. SI : s t : SeT 5 Weknht Per cont 296 aalo none ded 297 male none + 2, male MM male f capon xlrn^atla capon androgenc capon androgenc capon androgenc capon androgenc and estrogenc capon androgenc and cstro^enlc capon androgenc and estrogenc +27. C capon androgenc ad estrogenc US estrogenc IMS sale estrogenc male estrogenc alt estrogenc ,4 347 aalo none male MM aalo BMM aalo none male none capon androgenc > capon androgenc capon , capon androgenc ded 342 capon dc ca;>o: androgenc na estrogenc capon androgenc aod estrogenc +32.C capon androgenc an estrogenc + 32.' capon androgenc ttd estrogenc + 32.C capon androgenc t c: Erogenc +26.; aalo estrogenc ded 355 raalo estrogenc aalo trogenc male trogenc , aalo C , mpm estrogenc + 6,7 2 capon estrogenc cupo... estrogenc feraale Erogenc +4, feoale estrogenc +42, fomale estrogenc +42,56

20 0 Group, consstng of four normal male brds, 296, 297, 300, and 327, was used as tle control group for the frst seres of experments. The mean ntal erythrocyte count for tlls group was 3,380,000 and the aean Intal hemoglobn concentraton was.3* Ths group receved no >nos and no bland njecton* were used. The lowest mean erythrocyte count recorded for ths group durng the nvestgaton was 3,30,000 whle the hghest was 3,420,000 observed on the thrteenth and fnal count. Durng the 3 weeks, the curve vared slghtly from week to week but In no case wore the varatons sgnfcant, Plgt. The slght ncrease In erythrocyte count ovor tle entre perod was probably due to the maturng and growth of the brds,.loan hemoglobn concentraton readed ts hgh pont of 2.7 on tle fnal determnaton and ths curve showed a rather steady (gradual ncrease from week to week. As wth the erythrocyte count, the net Increase n hemoglobn concentraton was attrbuted to growth and maturty* One Indvdual of ths group, brd 296, ded from njures sustaned w>lle fghtng durng the sxth week of the experment. A radcally dfferent curve was produced by tle four members of Group 2, brds 236, 290, 23, and 293, Pg. 2. These brds were all capons and receved njectons of testosterone proponate* Fre-njecton counts shewed a mean erythrocyte count of 2,440,000 for ths group. Immedately after the frst njecton the mean count began to rse rapdly. In two weeks t reached 3,060,000 and after droppng to 3,040,000 for one week, clmbed less steeply to reach 3,330,000 after fve weeks of Injectons, For one week after the last njecton t remaned at approxmately ths level

21 9 and then began a rapd decrease. At the end of the thrteenth week* t had dropped to 2,430,(300 almost dentcal to the prenjecton level. Tlss average ncrease In erythrocyte count for the group was per cent and three of the ndvdual brds wore wthn three per cent of ths avorage. The fourth brd, 293, showed a per cent Increase, but snce ths ndvdual began the experment wth an oxtrentely low erythrocyte count It was assumed that the relatvely large ncrease was due to a temporary anema at the begnnng of the experment. Ths assucton was further supported by the fact tlat the erythrocyte count of brd 293 lad decreased only to 2,300,000 at the concluson of the experment. Mean weakly hemoglobn concentratons for ths group roee steadly from an ntal average of 9»2 to a peak of 4*0. The frst two weeks of njectons caused a steep Increase from 9.4 to 2,0 and the subsequent ncrease was more gradual. The peak of 4.0 was readed one week after the last njecton and two weeks later tlmn the erythrocyte peak. The hemoglobn curve form* ed a plateau between 3.5 and 4.0 begnnng durng the ffth week of njectons and extendng for fve weeks, or four past the fnal Injecton. Durng the ffth post-njocton the hemoglobn concentraton dropped sharply from 3.7 to 0.7 and dd not drop any further durng the expermental perod. The four members of ths group vared wdely In ther percentages of ncrease In hemoglobn concentraton, averagng as a group a 52.7 per cont ncrease, but as ndvduals, rangng from a per cent ncrease to a per cont ncrease. There wae not a

22 consstent correlaton between eryt t count and hemoglobn centraton, snce brd 293 bad the greatest ncrease n erythrocyte number and the smallest ncrease n hemoglobn concentraton. Brds 294, I and 235, all capons, made up Group 3, whch receved njectons of both :aale and fonale hormone, An average eryt'rocyte count of 2,470,000 was estal J. for *s croup pror to the frst njecton. Durng the frst four weeks of njecton, the mean count roso to 3,020,000 and added a very small ncrease the followng week to rer.ch a peak of 3,060,000 after fve weeks of njectons*?or tlls group the curve rose less steeply and to a lower peak than the curve for the precedng group, Fg* A plateau between 3,020,000 and 3,060,000 extended over a perod of three week3, t.'se last three durng whch the hormones were admnstered* Immedately after cessaton of tl* njectons, the mean erythrocyte count began a gradual decrease whch was accelerated durng tl& last three weeks of the expermental perod* At the end of the 3-week perod, the mean erythrocyte count for the group was 2,420,000, slghtly lower than the pre-njecton level* As a group these four brds ncreased ther erythrocyte count 28*4 per cent but there was a wde devaton n percentage of ncrease among tlje four ndvduals* The wean hemoglobn concentraton for ths group rose stoadly from 9*6 to 3*3 durng the frst fve weeks of njectons. For three weeks, the last two of the njecton perod and the frst after the njectons had ceased, t remaned between 3*0 and

23 4*0, reachng tlls peak one week after the end of the Injecton schedule* It then fell rater sharply and steadly untl, at the end of the 3-woek perod, t was 9*9, a slghtly hgher level than that observed n pre-njecton determnatons* The mean Increase n hemoglobn concentraton for ths _p was 47*32 per cent, but once agan vdua consderably from the oml.. The four brds n Group 4, 209, 290, 329. and 328, were normal males and receved njectons of ostrogeu*.or tls group the curve was very nearly an nverted replca o o for Groups 2 and 3, Fg* 4* These brds allowed a marked decrease n both erythrocyte count and hemoglobn concentraton under t>*b nfluence of estrogen* A pro-njecton mean erythrocyte count of 0, JO was establshed for tha group and the mean weekly counts decreased steadly after the njectons began. low mean count of 2,750,000 was reached after fve weeks of njectons and durng the sxth and last week of njectons an nsgnfcant ncrease of 20,000 was recorded* Immedately after the njectons ceased the mean erythrocyte count began a steady rse and at the end of 2 weeks stood at 3,o;>0,000* At the end of the 0-week expermental perod the mean count had slumped to e» *. a level apprecably lower than the pre-njecton level* As a group these brds showed an average decrease of 20*28 per cent* Te mean hemoglobn concentrat r these brds was not nfluenced by the estrogen to the same extent as the oryt srocyte count* After four weeks of njectons t had dropped only onetenth of a gram from the pre-njecton level of 2.0*./urng the

24 JJC last two weeks of the njecton perc, sever, the mean bn concentraton droppod to a mnmum of UU4. Izanedately after tbo njectons ceased t began a steady rse and reached 2*9 at the end of the twelfth week* At the end of the rteenth creek, the mean concentraton was 2.4, whch waa fourtenths of a gram hgher than the pre-njeeton lovel* The slg decrease durng the thrteenth week accompaned the prevously descrbed slump for the mea erythrocyte count The decrease n ::»an hemoglobn concentraton for ths group* jo\xp was 6.66 por cent but once a^a era were wde dfferences n the decreases observed n the ndvdual brds* Groups 5-0 were used dur o second seres of experlslants* Group 5 ncluded brds 347, 35, 372, 34, and 357, all nonaal males, and was used as the control group for ths aeres* V7eekly nean erythrocyte counts for ths group ranged from a low of 3,G0,000 to a hgh of 3,250,000. Ths range waa lower than that observed n the other control group because these brds wore two months younger than the brds In Group. sudden or sgnfcant changes occurred n the curve, smlar results were obtaned from weekly hemoglobn determnatons. In ths ease* the mean concentraton vared between 0*8 and. and no sudden or sgnfcant changes were observed,.. Fve capons, 333, 340, 343, 336, and 342, wore ncluded n Group 6 and receved Injectons of testosterone propone Prom a pre-lnjocton lovel of 2,440,000 the mean erythrocyte count rose to 3,450,000 at the end of the sx week njecton perod* The curve of ths rse waa farly regular, although slghtly

25 zz ctooper dor. r»st four weeks of njectons,. a soon as the njectons were stopped the mean eryt:.rocyte count bc-an a sharp decrease, fallng to 2,000,000 at the and of the twelfth week and decroas 00 at the end of tle 3-week experaental pc: he ncreaso for the mean erytrocyte count of the group was per cent but Indvdual ncreasea ranged from 24*3 p< (t to por cent. A pre-njocton level of 9,3 was establsled for the neon hemoglobn concentraton of ths group. Ths rose rapdly durng the frst four weeks of njectons* reac ng 2.3 at the end of fourth week. Durng the ffth week of njectons t rose only one-tenth of a gran to reach ts peek of 2,4 and durng the sxth and last wee: of Injectons returned to 2.3. Tms a plateau between 0.3 and 35.4, coverng the last t'ree weeks of the njector. od, appeared n the curve. After tle last njecton the mean eoneentraton od steadly and at the end of Mm 3-week perod stood at 9.4, only one- of a gran her than the pre-injocton level. As a group, these brda jwed an ncrease n ther xaean hemoglobn concentraton of per cent and the Indvdual ncreasea ranged fro 3.00 per oent to 4.36 per cent. Group 7 ncluded brds 339, 33G, 345, 335, and 344, and aa n Group 3, these brds were all capons and were treated wth both male and feme! ^ones. In ths group the mean erythvoeyta count rose rapdly from a pro-njecton level of 2,560,000 to 3,30,000 t the end of the thrd week of njectons. Durng last three weeka of njectons tl*e mean count contnued to ncrease.

26 M but less rapdly, Pg, 7, t reached a peak of 3,250,000. at the end of the sx week Injecton perod* Irraodately after the njectons ceased, t:«.nean count began a steady decreaae and rcrched 2,450,000 at the end of the 3-week Injecton perod* Ths was defntely lower than the pro-injecton level* The ncrease n the nean erythrocyte count for tls group amounted to 5 per cent* In ths croup, sooe decree of consstency was observed for the only tr» dur; nvest' I as three brds, 338, 345, and 335, showed dentcal ncreases of 32*65 pe b, whle brd 339 ncreased 34*69 per cent* The fft amber of t jup, brd 344, was the only one varyng aarkedly wth an ncrease of only 26*52 per cent* A raarked ncrease n heno^lobln concentraton was also observed n morabors of the group. Pros a pro-njecton level of 9*8 the aean hemoglobn concentraton rose to a peak of 2*5 after four weeks of njectons. It then docroased two-tenths of a gjjses durng the last two weeks. of the njecton perod and greatly accelerated Its decrease after the njoctlons had ceased* At MM end of the 3-week expernrental perod t. had fallen to 9*7, one-tenth of a gram lower than the pro-injecton level* The.^ease In asan hemoglobn concentraton for» was per cent rle ndvdual brds vared from 6*36 per cent to per cent n ther ndvdual ncreases* The curve f >up was smlar to that for ^-roup 4, Ths group, consstng of fve noraal male brds, 356, 355, 352, 353, and 373, resolved Injectons of estrogen* The erythrocyte count for tms group dropped steadly from a

27 jyte 86 njecton levol of 3,000,000 to 2, 70 at ad of the sx week course of In Joc ts poot*njectl^ ;e was At almost e sarso rate as Its procodln/* decrease, and at ~.nd of ths 3-week expermental perod the mean evrthaseyta t vas 3,00,000, only 20,0?0 less than tlc px>e-ajccton lovol. o ^crease In : count ap was. pc I and the Indvdual decreases were all n ths coj^paratvely narrow rongo of 8.75 per co:t to.t» o ond of tlta - jectlons, the mean h^^lobn conccntr- was 3 of a cram lower than the pro-njecton lovel,,e next threw wsslrs, thw decrease was more rapd and at the end of ths sx week njecton perod the aean concentraton was dot (. n cessaton of tho njectons t row* steadly to reach.9 at the end of the 3-week expermental part. As a group, these brds showed a decrease n mean het aton of. 9 por eent, but ndvdual decreases ranged from G.3G per cent to D.3 per cont. One brd, 3C, a durng the eghth week of the Investgaton, apparently from a vtamn defcc jsted a oo ssms, 2,, 239, c, oevod njectons of female hormone. a ason eryt ovocyte eount for ths group dropped from a pro-njecton lovel of 2,650,000 to 2,400,000 after three weeks of njc, h** roturned to,500 before the end of the sx week njecton perod, l*an hemoglobn concentraton vared between 0.4 and Ifl.,g the nvestgaton,.. -ar results were observed for Croup 0, nclude*

28 Pr e results obtaned Igatlons r.jg. I '.. three norraal fertales, 25S, 259, a receved Injectons of estroge raean erytlrobe count dropped to 2,350,000 from ft pr of 2,550,000 durng the course of Injectons, but had returned to 2,560,000 at the end of the ss woe jd* The booh hemoglobn concentraton rr between 0# and B*8.s grotm. t was consdered that these changes were Iagnfleant. Fg. o«ii of the brds were weghed b ;xperasnt started and agan at Its eonolus *u The pore aln of oach brd was recorded and Included In t ".per ' evdence was obtaned whlo I a correlaton between the each brd receved and Its rate of weght Increase* Group 2 m concluson that testosterone proponate sed an Increase n tle erythrocyte nurabor and In the heooglobln concentraton was an Inevtable dervatve * lor frst njectons these brd od froa the oont roups only n lackng tec;. jaaantal factors were Identcal. Yet In every case, the eryt rocyto count and tl* heoo- >bn concentraton of the capons was defntely lowc* tlvm la the raale brds. r pon the admnstrat orone proponate to these capons ther raaleness was physologcally restored *od number and hetaoslobln concentraton roso to

29 27 at least ap-roxnate, and n many cases, to oxceod that of the ale brds* Whan the hormone was wthhold, the erythrocyte number and the hemoglobn concentraton of the capons returned to pre-njecton levels n a few weeks* These changes occurred n every ndvdual belongng to the groups concerned* To summarse, removal of the testes wth the resultng decrease n testosterone level, resulted n decreased erythrocyte numbers and hemoglobn concentratons; roplacevert of the testosterone by njecton resulted n the restoraton of the erytlrocyte numbers and the hemo» globn concentratons to the male level, and wthdrawal of the replacement horaone resulted n a return of the erythrocyte count and the hemoglobn concentraton to the capon le.el, Thcroforo, o testosterone was responsble for the observed ncreases n erythrocyte numbers and hemoglobn concentraton* Group 4 and Croup 8 provded the data from whch was derved the concluson that estrogen caused a decrease n erythrocyte number and hemoglobn concentraton of males* The brds n these groups were all normal males and lved under dentcal condton* wth the control groups pror to the course of njectons* Durng the perod of njecton they dffered from the controls only n rocevng estrogen* Ther pre-njooton erythrocyte and hemoglobn levels were very smlar to the control brds, but after recevng the estrogen njectons tusy deoreased untl they approached the oapon level* After the njectons ceased the erythrocyte numbers and Jemoglobn concentratons returned n a few weeks to the pro*njecton level. All members of tlk>ae two groups responded n the same fashon, rnco stoppng the adans-

30 tratlon of estrogen resulted n a return to te nalo level. It was concluded t at the estrogen was responsble for the observed decrease. The concluson tlat estrogen repressed the erytlropootc acton of testosterone proponate on capons was supported by the data secured from Group 3 and Group 7* These two groups, lke Group 2 and Group 6, conssted of capons that receved testostorone proponate njectons* owever, these groups dffered from Group 2 and Group 6 n that they also receved estrogen njectons* Whle the eryt rocyte number and hemoglobn concentraton of these groups also ncreased, ther mean ncrease was far loss than that of Group 2 and Group 6. Snce the only varyng factor was the estrogen, t was concluded that t was responsble for the losser ncrease* These same groups also supported the concluson that the estrogen took effect zaore slowly than the testosterone proponate* vvhen tle curve of Group 2 was compared to tlat of Group 3 the two were found to be very smlar for the frst tlree weeks of the njecton perod* After that pont, however, the curve of Group 2 contnued to rse at about the same rate wllle the curve of Group 3 flattened suddenly* A comparson of the curves of Group 6 and Group 7 revealed the same stuaton* Tme, t was reasonable to hold the flattenng to be due to the estrogen becomng effectve at that pont* The next concluson, that s that estrogen dd not act upon the blood pcture through a drect effect upon the blood elements or blood formng elements were obvous results of the data obtaned from Group 9 and Group 0. Group 9 ncluded three capons

31 HI and Group 0 three fonaloo end ell sx of tlseso brds receved Injectons of ostr. Although the erythrocyte count and the hemoglobn concentraton decreased slghtly durng the course of njectons, they recovered tlelr pre-lnjocton level before the end of tl-m njecton perod. Thus t-:c o.ccrcaoo was only temporary and was so snal as to bo cozsdered nsgnfcant* It was as* suned that ths temporary depresson was probably due to a general endocrne mbalance whch was soon corrected by t?je brd. If the estrogen acted drectly on the blood the erytjrocyto count and the honoglobln concentraton would have reacted as t ey dd n Group 4 and Group 3. Ince no smlar decreases were observed n these groups, te concluson that ths was not the method by whch estrogen acted was a Justfable one. An examnaton of all the data secured see^ed to ndcate that the ostrogen exerted ts nfluence on the blood pcture through ts represson of the anteror ptutary,.e result of estrogen's effect on the anteror ptutary Is two-fold. It reduces the amount of gonadotropc horraone produced, ndrectly reducng testosterone secreton n a male, and t reduces the erythropoetc secreton of the ptutary. These phenomona occur only when an excess of sex hormone s present. In the case of Group 4 and Group 3, the ostrogen reduced the androgen secreton and repressed the erythropoetc secreton, thus actng to reduce the erythrocyte count and the hemoglobn concentraton n two ways. In Group 3 and Group 7, both made up of capons, the only acton was the reducton of erythropoetc socreton. A cooparson of t-ve curves of Groups 3 and 7 wth the curves of Groups 4

32 30 and 8 showed that the estrogen had a much creator offect on the blood pcture of the males o t nhbted both erythropoetc and* ndrectly, androgenc secreton than t dd upon the blood pcture of the capons. Therefore, tlls hypothess could satsfactorly explan all cases observed* wmnm Almost wthout excepton, the results of endocrnologcal research n the past have agreed that testosterone cauooe an ncrease n erythrocyte number and hemoglobn concentraton n Mwwln and brds* These results :aave been obtaned wth a varety of expermental methods as well as wth many dfferent expermental anmals* -nklostoln et al* (944) examned the regeneraton of erythrocytes and hemoglobn after a heorrhogonduced anema In rats and reported that testosterone accelerated o erythrocyte regeneraton n normal males and females, hyporsoctoased males and females, and castrate mlos. About the regeneraton of hemoglobn, lowevor, ther results were not consstent, snce n many eases the testosterone njectons caused no acceleraton of the regeneraton process* Examnaton of the blood pcture of eunuchod men by UeCullagh and Jones (942) revealed that durng testosterone therapy both ther erythrocyte count and ther hemoglobn concentraton ncreased; and that upon cessaton of the therapy both returned to ther pre-treatment level. lar results wore obtaned wth rats by Crafts (94G), Vollmor and Cordon (94), Vollmer et al. (948), and Stenglass

33 s 3 et al* (04), tlth chckens by Tabor ot al. (04, 043), Jura* and Doom (030), and Doom et al* (043), and wth golden hamstors by Sten and Jacobsen (044) The results of the wrter* nvestgaton were n eomplete agreement wth such conclusons* The physology of ths phenooonon, however, was not defntely explaned by anyone* Each worker advanced one or more possble explanatons but none of them were able to secure expermental proof for ther hypotheses. lccullagh and Jones (042) suggested that the explanaton mght le eltljor In the effect of androgens on the metabolc rate or n the effect of androgens on the blood formng elements. They supported the lattor vew, although tley advanced an equally strong case for each explanaton. Support* ng evdence for the drect acton theory was found by Vollmor and Gordon (04) and Vollmer et al* (04S) workng wth rats* From ther examnaton of the red bone marrow they dscovered that testosterone caused a hyperplasa of the erythroblastc elements. They also found, as dd Pnkesten et al. (044), that tostooterone tended to prevent te hypoplasa of orytlro*. bastc tssue that ordnarly resulted froa Irjpopljysectomy* All of those workers suggested that to prolferaton of the red marrow mght be only a specal manfestaton of I oneral stmulaton of somatc growth by testosterone as descrbed by Goldzlehor (94) and Rubensten and Solomon (04). Other workers have been nclned to thnk trjt the affect of testosterone on the blood pcture s a result on an ncreased basal metabolc rate. Alt lough Beyer and Danow (942) reported that no change n the basal metabolc rate of rats was caused

34 ether by castraton or testosterone njecton*, most nvestgators have reported opposte results* A defnte ncrease n basal metabolc rate, caused by testosterone, was reported by Jones ot al. (94), Kenyon et al* (940). McCullagh and Hossmlller (94), and Sandlford et al* (94)* On the bass of such reports as t ese, Crafts (946) and Stenglass et al. (94) concluded t mt testosterone rased t*\«basal metabolc rate, oroby creatng an Increased oxygen demand} and that the response of the erythropoetc systea to ths demand resulted n tle ncreased oryt rocyte number and hemoglobn concentraton so generally observed after testosterone admnstratons* Ths theory also covered t:e known prolferaton of the erythroblastc elenents, snce t assumed that the prolferaton was n response to the ncreased oxygen demand* A few workers, such as Adams and Shevket (929) and l.ioycr et al. (940) suggested that any nfluence upon the basal metabolc rate was exerted tlrough the thyrod or ptutary, glands whch have a known and drect effect upon basal metabolc rate. 80 data were secured from ths nvestgaton whch permtted the addton of anythng to the dscusson concernng the method by whch testosterone exerted ts nfluence on the blood pcture. Past workers dd not agree completely as to the effect of estrogen on the blood pcture* Davs and Doynton (94) reported that stllbestrol, a synthetc estrogenc compound, had a benefcal effect on the hematopoetc system of human bengs of both sexes, but ther results were not supported by any other workers. Only a very slght decrease n erythrocyte number and

35 89 hemoglobn concentraton of male and female does after estrogen admnstraton was reported by Tyslotrta and Hartaan (94) but the saos workers later (Tyslowta and Dngemense, 94) agreed wth Castrodale et al» (94) that estrogen caused such a severe aneraa n dogs of both sexes that t proved fatal* Vollmer and Gordon (94) and Grafts (94) reported moderate, but defnte, decreases n erythrocyte and emoglobn levols caused by estrogen n doga and monkeys of both sexes, as well as castrate anmals} and Fnklesten et al* (944) found that estrogen nhbted tre regeneraton of erythrocytes n anenc rats, male, female, and castrate* A quanttatve dfference n the reacton of male, female, and castrate chekens to estrogen was reported by Tabor ot al* (943) who found that the erythrocyte number of male brds dropped much farther than dd the erytrocyte number of female brds or capons recevng the same amount of estrogen* Results of ths nvestgaton partly agreed wth ths concluson, as estrogen had a greater effect on male brds (Groups 4 and 8) than t dd on female brds (Group 0) or on capons (Groups 3, 7, and 9). A German worker, Feuchnger (940), found that estrogen caused only a temporary decrease n erytlrocyte number when gven to female rats, rabbts, and human bengs} and le further reported that a smlar transtory anon&a appeared durng eaeh perod of e3trus n rats. th ths last result, the prevously descrbed experence wth Group 0 was n oomplote agreement* Conclusons drawn from tlxs nvestgaton were that estrogen depressed the erythrocyte number and the hemoglobn concentraton n males and repressed the response to testosterone when both

36 M hormones were gven to capons. Ho such decrease was observed n the case or females and capons wthout androgen stmulaton, but only a small and temporary decrease* Thus, the wrter* s conclusons agreed wth some reached by past workers and dsagreed slarply wth others* The probable method of acton of estrogen on the blood pcture, as suggested by past nvestgatons, has boon a very confused matter* Perhaps a majorty of the workers assumed that estrogen actod drectly on the blood pcture* Crafts (94), Tyslowts and llertman (94), Castrodale ot al» (94), and Taber et al» (94) all advanced ths assumpton* Ther conclusons were generally based upon two premses* frst, that the estrogen caused a decrease n erythrocyte number of both sexes; and second, that a hypoplasa of the red bone marrow resulted from estrogen njectons* One of these workers (Tabcr et al«94) also thought that an antagonsm to testosterone was n part rosponsblo for tle effect of estrogen on tho blood pcture* The fact that the decrease of erythrocyte number due to estrogen was so much greater n males than In females or capons was the bass for ths oory* stll a thrd suggeston was that estrogen reduced the basal metabolc rate and thus the demand by the body for erythrocytes. Stonglass et al. (94) supported ths theory, although the effect of estrogen on the basal metabolc rate Is tself a moot pont. Sherwood and Bowers (956) found that ovaran hormones defntely depressed the metabolc rate of dogs whle Danforth ot al. (937) concluded that t had no effect on the metabolc rate of rats*

37 89» of the above descrbed theores satsfactorly explaned the results obtaned n ths nvestgaton, v sble excepton of the antagonsm theory. Snco t.ere s no expermental evdence of a chenlcal conflct between estrogen and testosterone, I wrter doubted that such an antagonsm exsted* Prom the data obtaned n ths Investgaton* t was concluded that the estrogen probably acted on the blood pcture through ts effect on the anteror ptutary. The fact that the ptutary was related to the blood pcture was deaonstrated by Crafts (94, 946), Bayer ot al«(940), and Vollmer et al. (939), They all agreed that hypophysectomy caused a decreased eryt to number and a degeneraton of the erythroblastc tssues, :erts and Meyer (937) and Belly and Solomon (940) demonstrated that the sex hormones depressed the actvty of the anteror ptutary and Rubensten and Kurland (940), Rubensten an omen (94, 94) and Edelsberg and Ornston (940) proved that only an excess of sex hormone caused ths depresson* Reasonng from the data secured from ths nvestgaton, n the ] o above lsted facts, t was concluded that a workable lypotheas could be constructed t would explan the acton of estrogen on the blood pcture n terms of ts In the caso acton on the ptutary* of Group 4 and Group 3 n ths experment, whch ncluded normal males recevng estrogen, the hormone, throu^ ts acton on the ptutary, exerted a dual nfluence on the blood pcture. By reducng the secroton of Gonadotropns t Indrectly reduced the producton of androgen by the testes and so decreased the stmulaton of the hematopoetc system* By reducng

38 86 the secreton or the eryt ropoletc fracton of the ptutary* t decreased the hematopoetc stmulaton from that source* The lack of snllar results n the female brds and n fcfca capons of Group 9 ndcated that the dosage used dd not consttute an excess of hormone for those partcular brds* In tlyo caso of the capons n Group 3 and Group 7, n eh e androgen nduced ncrease n erythrocyte count repressed by estrogen* the effect ess entrely due to the reducton of the erythorpoletc secreton from the ptutary, naturally* tlje effect was not as marlsed n those brds as t was n the normal males* snes a sngle acton was not as effectve as a two-fold acton* The decreases n erythrocyte number observed n female and castrate anmals after estrogen admnstraton by prevously mentoned workers ra^ very well have been due to the fact t hat tle dosage they used consttuted a very defnte excess and exerted a strong repressve Influence on the ptutary* SUMMAKY -am the procedure and results descrbed n the precedng pages, the followng conclusons were derved* * Testosterone proponate caused an Increase n the erythrocyte number and the hemoglobn concentratons of oaponaod Whts Leghorn olckens* S«-*ogen caused a decrease n the erythrocyte number and the hemoglobn concentraton of nale whte Leghorn chcken;. 3 # In capons whch receved both horaonos, estrogen

39 89 the erythropoetc acton of testosterone proponate on caponsod Whte Leghorn chlcccn:. 4* ICstrogen took effect acre slowly than testosterone proponate when both horraooe* were gven to caponaed :/hlto Leghorn ckens* trogen caused s(. angos In erythrocyte count and hemoglobn concentraton only n sales and n capons who also receved testosterone proponsto* C # ^strogen does not act drectly upon the blood elements or upon the blood forrang eleoe.ta of Whte Leghorn chcuens. 7..?ogeu nay act upon the eryt rocyto nusaber and upon the hemoglobn concentraton tl-rough ts effect on tle anteror lobe of the ptutary gland of Whte Leghorn chclre

40 II ACKHOKLEDGMEHT The wrter wshes to express Ms grattude to Dr*. Herrlck, Professor of Zoology* Kansas State College of Agrculture and Appled Scence, for?s supervson, advce, and assstance, both n the Investgaton and n the preparaton of the thess. The wrter also wshes to thank Oro* Los Montagus for her knd techncal assstance.

41 » LITbKATUHE CITED aabeth and Faele Shevket, o noraal pctu*e c.3 n the pcture followng thyrod feedng, Phyaol, Zool, 2: Aprl, 02: Bolly, Jacob S. anc. La.uel Solanen* The nhbton of lactaton post-partum wth testosterone proponate* Endocrnol.. Bluael, Paul* Castraton and erythrocyte count, (n German), Lfcd. Cln, 24: C; Caetrodale, Dante,.ja Bcrbaum, bson, 2*lg# end Cyr~~. HaeBryde, Comparatve studes or of ostradol and stlbestrol upon the blood, lvor, and bone marrow* Endocrnology. j 3C3-37. eptoaber, 94, Craft: The effect of endoorlnes on the forced olorents Bndoc.. : , October, of the blood, Effects of hypophysectoray, castraton, and testosterone pvayl oaate an bnapalwaa *.. t a adult nala n ( >- crnology, 39 40*43. Decenbor, 948# Danforth, E. ::,, R. R, Greene and A, C, Ivy, J3 effect of female sex hormone* on the oxygen conaaapton rate of normal rata, and upon the tolerance" to desscated jrod, bndocrnology, 2: 86~867«bay, 937, ' Davs, bdward II, and "elbouroe W, Boy». Indcatons, cllnleal use, and toxcty of 4-4» dlydroxydethyl stlben. aw, lft, ndoornc :, , Aprl, 94. L, v., ;slo Taber, and Davd E. Davs, parson of e lumbers n normal and loraooe treated brown leghorn fowl. Soc. bxpt. Bol, and Had, Proc. 62: 49*50. ft olaberg, Josop, and bllot /., Ornate L. Observatons on the contnued use of male sex hormone over a long perod of tne, Endocrnol.26*. January,

42 0 tngor, Oswald* Follcle hormone and the blood pcture* (n Oer.an). Arch* s* Sxpt. '.. u* Pharmakol. 96! G44-G70* 0< uen, "race, Albert on and "or: harrper* The effect or sex hormones on the anema nduced by hemorrhage n the rat- xlocr: 35: 2G7-277,, 944. dseher* Sax A* Growth and sex hormones. Jour* Cln. Endocrnology* $ * November, 94* -ta, Roy, and Roland K* Meyer* Tha effect of testosterone, testosterone proponate, and dhydroe berone on tle secreton of the gone complex as evdenced n parabotc rats, jcrnolo. 8t 73G-744. Hovonher, 9. Jones* Held, s* Perry KcCullc..,oy EScCullagh and 3. W* Buckaloo. Observaton on the hyper-metabolam nduced by metlyl testosterone* Jour* 6.docrnology* Is August, 94* Juhn, Hary and L. V* Doom* The relaton of gonadal condton to ^ocvte number n fowl* Amer. Jour. Physol. 94s G0-G2. September, 930* Kenyon, Allan T*, totlryn Knowlton, Irone Sandford* ft G. and Gertrude Lotwn* A comparatve study of the metabolc effoots n normal men and women and n eunuchodsm, ulocrnology. 2G-45. January* 940* MaCullagh. K. Perry and II* R. Rossmller. I testostorono - oalorgenc r.ctlvty* Jour* Cl» Sadocrlnology* It June, 94* HoCullagh, lb Perry and Rad Jones* Effect of androgens on tle blood count of Endocrnology* 2t Aprl, 94:,.on. Jour. Cln. Meyer* Arthur K. and alan l*now* Influence of raethyltestosterone on metabolsm of normal* castrate, and thyrodectomsed rats. r >oc. xpt* Bol, and Bed* Proc Aprl, 94.' Heyer, Ovd.. ol W, Thewlls and Harold P* Ruse. torn broep nu end Hfglfj Mteovlmfeenrt. :z NMM Padoa, Emmanuelo. ["ho change n erythrocyte number wth sexual nverson of the fowl, (n Italan)* Bu. Italan Soo* Expt. 3ol. G:

43 . Rddle, Oscar and Pela Fay Breachcr. Hemoglobn and erythrocyte dfferences accordng to sex and season In doves and pgeons* Amer. Jour* Phy ?!arch, 934* Idle, Oscar and George E* Caul ->yte number n young pgeons and Its relaton to 3x>redty, growth, and mc. Amor. Jour. Physol. 22: * May, Rubensten, B# s* and A. orland* :3e of the tests to snal doses of t03to3torc proponate* Endocrnology Joembor, 940* Kubensten, and M. olomon* 3 grow' rosel t of large doses of testosterone proponate n the castrate albno rat* uadocrnolo<;y January, Tle growth stmulatng effect of small doses of testosterone proponate n the castrate albno rat r/»nolo ~r. 'J 3: February, 34* Sandford, Irene, athryn Knowlton and Allan T» Kenyo^. Basal loat producton n hypogoadsn n men and ts ncrease by protracted treatment ulth testosterone proponate* Jour* Cln. Endocrnology* It * Deoeaber, 94* Sherwood,? C* and Lnwood M«Bowers* ct of ovaran hormone on the basal metabolsm of expermental hyperthyrold rats* Amor. Jour* Physol* 5: May, 93. ftfe latlryn and Barbara Jaeobsca. o effect of castraton and cryptorchdzatlon on the blood pcture of tl» golden laamotor. Anatomcal Kooord * (abstract). sngles*, Paul, 'Albert s* aordon and Harry A. Charrloer. ~*ect of castraton and sex hormones on tle blood" ncture of rat3* >c«:-:xpt* Bol* and Bled* Proc* October, 944* Teber, Mlsle, Dave ntl... jobb* The affect of sex hormones on the erythrocyte number n the blood of the fowl. Anatosdeal tecord* Bt 39* 94. (abstract). Effect of sex hormone* on the erytlrooyte number n the blood the domestc fowl. Amer. Jour* Physol * February, 943*

44 49?ru Influence of castraton and the transplantaton of gonads on the erythrocyte nusbor and the heno^lobn quant E - (n Italan). Bu, Italan *oo. :xpt. Bol. 2: 036-r>>39. s 7. 3 Tyslowtz, R, and Carol ::. P nt«auj J?lue;co of large doses of cstrogoa on the blood pcture or osus monkeys. Endocrnology, 29: ,. September, 94, Tyslowts, dm and I* Jngexaanee, large doses of estrogen on the blood pcture of dogs* Endocrnology, 89: * JVttotMT, 94 Vollraer, ; >rarn F 9 $ Albert. ordon, Irvng Lovensten and larry A*Charrper» Effects of hypophysectorjy on the blood pcture of the rat, endocrnology, 25: , Deeenber, 939, Vollaer, Erwn P, and Albert S«0o:*do:. Effect of sex and gonadotropc omones on the blood pcture of the rat, "ndocrl: olo t» 20* , member, 94, lnor t "Orwn P Albert r>, Gordon and larry A, Charrper,..ts of horaones on erythropoesle n aypopr rats, ISndocrlnolocy, 3: , December, 94.

45