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o mcfiuoouov mo vogquN mofioxo umoE mo omcmm: .o.w :mozH whoa onoxzo w.N m.om w upozom Numa Vim wmuom Nmmmm w hmzmz Huma m.omm.v H.ohm.m m.HHm.Hm n.0nm.mm camcouumz onmfi Hownofiz o.~no.wm w ocho>m¥ onma schemafiaa Hauumm H.mnu.ea m mm.wnum amcocaz moma w.wm :Houmxum mm wig w.» all: ma Isl; «vogue: Hommzm amasoflfiflom Hecaummso 30am Hapmcoq oHoxu waHm fi.e.u:ouv H mgm . m momcoz . z :oHumcHmeo onoomOHmmmH . m 2 z z z N H mN 0N mH 0H m N H P— l P PP PF 4 H a 14 u H m m m mewv Nm u oHoxu HmEHozv Hm onEom z z z z z m N H mN ON mH 0H m N H La; ».beb. H; . 4 aweawaam a m m memw wN u oHoxu HmEHozv we onEom z 2 z 2 2 m N H mN ON mH mH NH 0H m m w bh. EIP __ p _ . m miw . . memw wN u oHuxo Hmspozv me onEmm z 2 2 2 z m N H mN ON mH NH 0H m N H HFFIT H u L Lytpu [Hi Ji uLfL w m.w m.mem m 1m. Hmzmw om u mHoxo HmEHozv ow onEom quoHHoxu Heapumcoz co xmoomOHmmmq HmHHom mo muoowmm m mmzuHm 3O wchmoz . 2 :oHuHHSHHmm u a Hmoomoammmq : m momcoz u z couuafis>o - >0 «N ON mm NH w w 2 >0 a m. m w meoozv :OHumuomq manuocsmmco> wee mHmogumoc< moH see me New ........... made mm ................ T - LI 1 a db a w w we: >0 :oHumumou Nm onEom mo :oHumuoma wee coHumpmoo v mmDon 31 close association with the follicle. Ovarian enlargement and a darkening of the reddish color were other indications of follicular growth. These characteristics were usually observed 24 to 48 hours prior to ovulation. Eight to twelve hours before ovulation, the vascular network was observed to deve10p to such a degree as to circumscribe a portion of the follicle from which the stigma will develop. The fully developed stigma appeared as a smooth, translucent, hemispherical structure. The moment of follicular rupture (ovulation) was observed and photographed in three animals (Figures 5 and 6). These are believed to be the only photographs of observed ovulation in nonhuman primates, although others have observed rupture during laparotomy. Immediately prior to extrusion of the cumulus mass, the clear follicular fluid was observed protruding from the site of rupture (Figdre 5). Figure 5. Immediately prior to follicular rupture, follicular fluid (between arrows) can be seen protruding from the stigma. 32 Following expulsion of the cumulus mass the stigmal membrane collapsed and considerable bleeding from the follicle was observed. The entire cumulus mass could then be seen adhering to the ovarian surface (Figure 6). Although follicular rupture and release of the ovum was rapid, it was not the explosive phenomenon observed in the rat by Blandau, and it required approximately 30 seconds for completion. After laparotomy to recover the ovum, the viscid properties and adhesiveness of the cumulus mass allowed it to be stretched 1.5 cm before it could be freed from the ovarian surface. Under microscopic examination the ovum was recovered from the cumulus mass. Figure 6. Immediately following expulsion of the follicular contents, the hemorrhagic cumulus mass (arrow) can be seen adhering to the ovarian surface. During the course of these studies, two general types of pre- ovulatory follicular development were observed. In order to classify differences between individual follicles, they were labeled as type 1 33 or type 2 depending on their morphological appearance. Follicles were classified as type 1 if the stigma was large, clear, and protuberant (Figure 7). There were no vascular elements associated with the transluscent stigma. The follicular vessels are usually seen circumferential to the stigma conforming to the ovarian surface. Following ovulation the corpus luteum forms an extensive mass that does not greatly protrude from the ovary. Follicular boundaries are sometimes difficult to detect in a type 1 follicle. The type 2 follicle generally encompasses a greater portion of the ovary than the type 1, and has a stigma that is smaller and less distinct than that of the type 1 follicle (Figure 8). The entire follicle is raised from the ovarian surface making the follicular boundaries quite distinct. Following ovulation, the corpus luteum develops into a highly protuberant yellow mass. Table 4 illustrates ovulatory morphology of individual animals. Cases where classification was questionable have been omitted. From these 46 follicles 30 (65.2%) exhibited the pre-ovulatory morphological appearance consistent with a type 1 follicle, while 16 (34.8%) resembled the type 2 configuration. It must be emphasized that these are generalized descriptions useful in standardizing classification. However, variations are observed in normal animals from time to time. One of the more interesting variations seen is illustrated by Figure 9. This poly- stigmal appearance has been observed to date in one normally cycling female. Five of seven follicles of this animal were of this type. In one additional, noncycling animal induced to ovulate with clomiphene citrate two of three follicles had this appearance. 34 Figure 7. Type one follicle showing large, clear stigma (a) circumscribed by follicular vessels (b) (Photo — top view; sketch - lateral view). Figure 8. 35 Type two follicle showing elevation of the follicular vessels (a) and the small clear stigma (b). The distinct fellicular borders are also observed (c). 36 TABLE 4 Follicular Morphology Female Type 1 Type 2 11 * * * 12 * 14 * * 11: 17 * 18 *(p) *(2) 40 * 'k * *- ‘k 41 *(L) *(R) * 'k 42 * 'k 43 *(L) *(R) * 44 *(p) *(p) * *(p) *(p) * *(p) 37 TABLE 4 (cont'd.) Female Type 1 Type 2 51 'k 53 'k * 54 * * p - polystigmal follicle 38 Figure 9. Polystigmal follicle observed in one normally cycling animal. 39 DrugEffects on Ovulation Clomiphene citrate Administration of clomiphene citrate to induce ovulation and menstruation in amenorrheic and in normally cycling animals was found to be quite successful. Figure 10 illustrates the time relationships between drug administration, ovulation and menstruation. Of the eight animals used in this study, six (75%) ovulated and menstruated within three treatment cycles. No clomiphene was administered during the rebound cycle. In those animals that were previously amenorrheic, three of four responded to the drug treatment by ovulating during the first rebound cycle. The fourth animal required further clomiphene administration but did ovulate during the second rebound cycle. Laparoscopic examinations and uniformity of menstrual periods following ovulation give evidence that ovulation was induced at the normal time during the cycle. Of the regularly cycling animals, only one ovulated during the first treatment cycle. Two of the remaining animals ovulated suc- cessfully during the rebound cycle while the fourth animals's cyclicity was interrupted by the clomiphene administration. These aniamls, like those in the earlier group, show uniform menses following the drug-induced ovulation. The possibility exists that the clomiphene was acting differently in these two groups of animals. Laparoscopic examinations at intervals optimum for recording the drug effects on follicular morphology revealed that many small follicles were seen which did not ovulate and later seemed to disappear. Of those animals that did ovulate fOIIOWing clomiphene treatment, the follicles were invariably larger than normal, and in 40 z oo oo h b bli- 14. 1 >o oz 1 one»: 1 mm mafia—om so no no go so _ p h H H _ LL oz 1 oz 1 oz oz 1 oz 1 mm ofimsoz z z oo go i p _ FL 1 >o 1 >o mom»: 4 oz 1 oz oHaEma z oo 4o 1 1 >o moazz 1 a>o ill o onEom Huguenozo m mzuzo a oHozo m ofiozo N ouuzo H oHuzo mHoEHem oHoznhocoE< eoHposoopmom co muoommm oumanu ozozmHEoHu mo Humeesm oH mmDuHm ki 41 :oHuahSmHofiaoa oeozoflsoHo - so momcoz I z :oHuoHoeHumuomx: - awe»: coupaao>o - >o 2 Ho Ho _. FL 1 a >o 1 mom»: a mom»: Hm onEom 2 z 2 mo .1 _ l 1 1 1 >o 1 me oHeEom 2 Ho 40 r F? I 1 >o 1 oz 1 oz ow oHNEom AU AU 1 u» i 41 oz . oz 1 02 NH oneom zaaaosazo m oHoxu v oHozo m oHoxu N mHoxu H oHoxu mHmchm hmstom H.o1u:ouo oH mzooHa 42 several cases had abnormal appearances (Figures 9 and 11). Another frequent consequence of this treatment was ovarian hypertrOphy in the absence of ovulation. Figure 11. Abnormal ovulation following clomiphene citrate administration to an anovulatory macaque. Megestrol acetate The effects of megestrol acetate (MA) on follicular development and ovulation is another example of the type of study facilitated by the technique of laparoscopy. Of the three animals receiving MA for eight consecutive days two ovulated between days 14 and 18 of the 'treatment cycle (Figure 12) and again successfully in the subsequent, untreated cycle. The third animal did not ovulate until late in the cycle following MA administration. Of those animals receiving MA on four consecutive days, two ovulated during the treatment cycle, while the other two did not ovulate until the cycle following MA adminis- tration. Withdrawal bleeding occurred in all cases to verify the 43 om om om m1---m1 1221. 1 >o 1 22 >QTT 1 m onEom <2 om om mm m1 ........ o F _ v r .11! 2222 >o 1 222 2o 1 NV meEQm <2 om om om m1 ........ m H r T 1 22 >o 1 221 12o 1 av mfimfiom <2 m1 om om mN-1~ m1 ........ o 21 >2 1 222221 1 1 1m o1aaoa :oHuoswonmoz so mpoommm oumuoo< Hoaumomoz mo AHmEESm NH mmDuHm 44 :011a1oso - >o momma: .. z <2 om m1---N1 w >6 “ 22:2 1 _ M m 0 H me m <2 om m1---m1 P r l 1 a 222 >ol 1 A1 o1asoa <2 om 1 m1---m1 1T >m 1 2221 1 1 <1 o1asoa H.w1u:oov NH m4m<9 45 progestational action of the compound in the animal and normal cyclicity returned during the subsequent cycle. The effects of MA on follicular development were minor, and included a possible decrease of follicular vascularity and delayed follicular rupture. All of those animals ovulating during the treatment did so between days 14 and 18. DISCUSSION Reproductive Cyclicity_ The similarities among Macaca mulatta, Macaca fascicularis and the human female in regard to menstrual cycle parameters represents a major reason for the increasing usage of nonhuman primates in research. The 30.8 r 1.0 day mean cycle length of the macaques from the present study agrees closely with the averages previously determined (Jewett and Dukelow, 1972; Nawar and Hafez, 1972), as does the 30 day median and mode. Calculations made from the data presented in Table 1 reveal that the average menstrual cycle length of the rhesus macaque (M: mulatta) is 27.9 days while the average of the crab-eating macaque (M: fascicularis) is 30.4 days. These calculations were made by omitting studies of each species in which the mean cycle data was inordinately high (Corner, 1932; Kerber and Reese, 1969; Kaverne and Michael, 1970). All of the studies deleted had mode cycle lengths that corresponded with the group modes (35.0 for M: :3 from Corner, 1932; 28 for M: m: and 26 to 28 for M: E: from Kerber and Reese, 1969; and 28 for M: m: from Kaverne and Michael, 1970), and it was considered that a small number of amenorrheic or abnormal animals had skewed the average lengths. This agreement confirms the normality of the animals used while the data from Table 2 suggest that 46 47 ovarian alteration is not essential for normal cyclicity since a high percentage of the ovulations occurred on the same ovary during consecutive cycles. The fact that there is a similarity among the reproductive cyclicity of these three primate species suggests many questions concerning the feasibility of using the nonhuman primate as a model for the study of human reproduction. Differences between species do exist, such as the summer amenorrhea ("summer sterility") exhibited by Macaca mulatta (Riesen et al., 1971). Recently Macdonald (1971) concluded his study comparing three macaque species by stating that the M: fascicularis seemed to be an excellent model for study of the control of ovulation, implantation, pregnancy, parturition and the teratogenic effects of foreign agents because its reproductive function proved to be continuous and infrequently hampered by amenorrhea. Ovulation Timing_ Knowledge of the time of ovulation is very important when studying the reproductive physiology of an animal, and is necessary where accurate conception times or fertilizable gametes are sought. Investigations designed to detect the time of ovulation have shown that direct visualization of the ovaries offers the most accurate method of studying the temporal relationship between the follicular and luteal phases of the menstrual cycle. Presently there are only two methods of observing ovaries in vivo, laparotomy and laparoscopy. Studies which were designed to reveal the normal follicular structure at laparotomy (Johansson et a1., 1967; Betteridge et al., 1970) were handicapped since only a few observations could be made during a cycle. 48 These workers did not discuss relationships between the follicular and luteal phase lengths. Laparosc0py has provided a means of observing follicular deve10pment and predicting ovulation times without the trauma concomitant with abdominal surgery. In this study, ovulation timing was possible without altering the lengths of the menstrual cycles. The 14.0 i 1.1 day follicular phase length observed confirms an earlier observation of a 14.1 i 3.1 day follicular phase in the Macaca fascicularis revealed by rectal palpation (Mahoney, 1970). The luteal phase length in this study was 15.6 i 1.8 days and did not differ statistically (p < .10) from the follicular phase length. From the data, it appears that ovulation in this animal occurs slightly before mid—cycle with a relatively stable follicular phase and a more variable luteal phase. Similar reports by Hartman (1932), who attributed almost all variation in cycle length of the M: mulatta to the post-ovulatory interval, and Mahoney (1970) who found a greater standard deviation in the luteal phase length than the follicular phase lengths of Macaca fascicularis, suggest that the growth and rupture of the follicle may require a shorter, more exact interval than does the formation, function, and lysis of the corpus luteum. Since reproductive functioning can be interrupted by stress, as is often the case when performing laparotomies (Betteridge, 1972), it was necessary to evaluate the effects of the laparoscopic technique on cyclicity. The data in Table 5 compare the cycle parameters found during four separate investigations of Macaca fascicularis, using the first three as control studies for the present work. In none of the three control studies were the animals exposed to surgical procedures. The fact that 210 laparosc0pic examinations during the TABLE 5 49 Cycle Lengths and Ovulation Times of Cynomolgus Monkeys Source Mean Cycle Lengths Ovulation Time Technique1 Fujiwara et al. 28.513.3 (S.D.) 12-17 Laparotomy 1969 (N=272) (N=4) Mahoney 31.1:8.9 (S.D.) l4.li3.l Palpation 1970 (N=25) (N=18) Macdonald 31.3:l.5 (S.E.) 1971 Present Studies 30.8:1.0 (S.E.) 14.0:1.1 Laparosc0py (N=115) (N=19) 1Technique of Ovulation Timing course of this study did not alter menstrual cyclicity suggests that the anesthesia and laparosc0pic procedure are not a sufficiently stressful stimulus to interfere with normal reproductive functioning. In several of the animals where serial laparoscopic examinations were made during the same cycle, menstrual periods were normal. One animal that was anesthetized daily for a month and laparoscoped twice, ovulated, conceived, and delivered a normal infant after normal gestation. Further laparoscopy was not found to affect lactation. Nprmal Ovulatory Morphology_ Direct observation of the ovarian changes is the best method 0f determining whether or not ovulation has occurred. Hartman's studies using rectal palpation (1932, 1933, 1939) showed amazing accuracy when predicting the time of ovulation but were of little 50 value to the study of the physical nature of follicular development. Most previous attempts to observe the ovarian surface before and after ovulation, have been done at laparotomy, due to a lack of suitable technological means. One of these studies by Johansson et al. (1968) concluded that the appearance of pre-ovulatory follicles varies considerably but that the stigma could be found within two days of ovulation. Another study (Betteridge et al., 1970) took issue with this statement and, based on inability to observe stigmal morphology, suggested that if such changes occur in the rhesus macaque they must be abrupt. Jewett and Dukelow (1971, 1973) using the same colony as the present study, adapted laparosc0py to the nonhuman primate and described the progressive development of the follicle before and after ovulation. These workers did not attempt to characterize the morphological structures observed during consecutive cycles in the same animals, and follicular rupture was not observed. The present study confirms the descriptions of follicular enlargement prior to ovulation and the relative time sequence of events. The normal pre-ovulatory follicles can be observed in various stages of deve10pment for intervals of two to four days, and the specific arrangement of follicular vessels can often be observed for intervals of 15 to 24 hours. The observation and photographic recording reported herein of ovulation in the nonhuman primate is the first report of its kind, although cinematography of follicular rupture has been recorded for both the rat and rabbit (Blandau, 1955). Photographs of ovulation and subsequent recovery of the ovum have confirmed the relationships 51 observed previously during this study and have shown that the point of follicular rupture is located on the stigma. The fact that the cumulus mass is very adhesive and remains in close proximity to the ovarian surface immediately after ovulation suggests that the transfer of the ovum to the oviduct may occur as a result of the sweeping motion of the fimbria over the ovarian surface as observed in the rabbit. Another observation supporting this hypoth- esis is the close association of the fimbria to the pre-ovulatory follicles. The predominance of the type 1 follicle during normal ovulatory cycles suggests that its characteristics may be those associated with normal ovulation. The other major type of pre-ovulatory follicular development occurs with sufficient frequency in normal cycles to suggest that its appearance is not abnormal. One can speculate that since these two types have been seen in the same animal, and even on the same ovary, the difference may be a function of the physical nature of the ovarian covering (the thickness or tenacity of the tunica albuginea), and the location of the devel- oping follicle in the ovary (depth in the interstitium, or proximity to the suspensory ligaments). Polystigmal follicles were observed in one female during 70% of her ovulations. Such abnormal morphology may reflect a variation in endocrine regulation. Further evidence that hormonal regulation may be the cause for this type of morphology comes from observations in an animal induced to ovulate with clomiphene citrate who responded with two polystigmal follicles out of three ovulations. 52 Drug Effects on Ovulation Clomiphene citrate The ability to induce normal ovulation in an abnormally cycling animal would be a valuable addition to the area of reproductive physiology. The induction of ovulation by clomiphene citrate was examined laparosc0pically during this study. The results show the general applicability of laparoscopy to the screening of compounds of this kind. Ovulation was successfully induced in at least onei cycle in 88% of the animals studied. All of the amenorrheic animals that were treated with clomiphene ovulated and, in all cases, the clomiphene induced single ovulations, which were charac- terized by large, in some cases abnormal appearing, follicles. The ovulations usually occurred mid-way in the third (rebound) cycle and were invariably followed by menstruation within 10 to 20 days. These findings agree with those observed by Wan and Balin (1970) who found an ovulation rate of 94% in amenorrheic Magaga_ mulatta. Their hypothesis that the mechanism of action of clomiphene in these animals includes an anti-estrogenic inhibition of the hypothalamic negative feedback loop causing subsequent release of gonadotropins is supported by the present study. Administration of clomiphene to regularly cycling animals resulted in a cessation of ovarian activity in 75% of the cases studied, a result inconsistent with the proposed mechanism of action. Wan and Balin attributed their failure to detect ovarian hyper- stimulation to a possible species difference, or an effect of varying endogenous gonadotropin levels, a variation often seen in human patients (Kistner, 1965). 53 In view of the fact that ovarian hyperstimulation was observed during this study, and that cessation of ovulatory activity followed- clomiphene administration in the previously normal animals, it is conceivable that their latter suggestion may explain the differences in response. If the anti-estrogenic effect of clomiphene in animals with a normal endogenous estrogen level is to inhibit the stimulation of gonadotropins at the pituitary level, such a cessation of activity might be expected. In any case, further study of the mechanism of action of this potentially valuable clinical aid are needed. Megestrol acetate Megestrol acetate (MA), an effective progestational agent, when administered at 500 ug daily on days 8 to 16 or 12 to 16 of the menstrual cycle was not effective in blocking ovulation in Macaca fascicularis. At this dosage, MA has been shown to be completely successful in blocking ovulation in the squirrel monkey (Saimiri sciureus) (Harrison and Dukelow, 1971). Daily administration of this level in women was shown to be an effective contraceptive program without blocking ovulation (van Leusden, 1969). The dif- ference in response patterns among these species may reflect either the variation in animal size, since the squirrel monkey is approxi- mately 10% as large as a mature macaque, or possibly a variation of endocronological regulation. A recent study by Spies and Niswender (1972) indicated that when 500 pg of progesterone was injected into rhesus monkeys on days 2 to 10, 8 to 16 or 8 to 22, ovulation was inhibited in all cases. This apparent contradiction with present results may reflect a different biological activity between the progestational agents or a species difference. SUMMARY AND CONCLUSIONS The menstrual cycle parameters of Macaca fascicularis determined by laparoscopy during this study were in agreement with previous determinations employing other methods, and suggested that the large number of laparoscopic examinations did not alter their cyclicity. The 14.0 r 1.1 day follicular phase length was not different from the 15.6 i 1.8 day luteal phase length determined by laparosc0py but suggested that the luteal phase might be longer and more variable than the follicular phase. Alteration of ouvlation between ovaries was not indicated and in fact, data on paired cycles and on longer consecutive series suggested a random selection of follicular site. Pre-ovulatory follicular morphology was photographed and categorized during consecutive menstrual cycles from individual animals. Two major types of development were described and follicular rupture was observed and photographed in three cases. Following observation of ovulation, laparotomy was performed and the cumulus mass was found to be closely adhered to the ovarian surface. The cumulus mass was removed from the ovarian surface and the ovum was found in its center confirming that ovulation had indeed just occurred. Clomiphene citrate was successful in causing ovulation in at least one cycle in 88% of the animals studied. Clomiphene-induced ovulations usually occurred mid-way in the rebound cycle and were 54 55 followed by menstruation within 20 days. Ovarian hyperstimulation and abnormally appearing ovulations were frequently observed during treatment. Administration of a progestational agent, megestrol acetate, during regular menstrual cycles did not inhibit ovulation or alter follicular deve10pment. LIST OF REFERENCES A-_ -:- .IY1‘-l LIST OF REFERENCES Allen, E. 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Presented at the American Institute of Biological Sciences Regional Meeting, Erie, Penn., 1971. Ovulation Morphology in Nonhuman Primates, by J.M.R. Rawson and W.R. Dukelow. Presented at the Michigan Academy of Science, Arts and Letters, East Lansing, Mich., March 23- 25, 1972. Comparative Ovulation in Nonhuman Primates, by W.R. Dukelow and J.M.R. Rawson. Presented at the Federation of American Societies for Experimental Biology Meeting, Atlantic City, N.J., April 1972. Natural and Artificial Control of Ovulation in Nonhuman Primates, by W.R. Dukelow, R.M. Harrison, J.M.R. Rawson, and M.P. Johnson. Presented at the Third Conference on Experimental Medicine and Surgery in Primates, Lyon, France, June 21-23, 1972. Follicular Development and Ovulation in Macaca fascicularis, Saimiri sciureus, and Galago senegalensis, by W.R. Dukelow, D.A. Jewett, and J.M.R. Rawson. Presented at the Fourth International Congress of Primatology, Portland, Oregon, August 15-18, 1972. 62 APPENDIX B ABSTRACTS COMPARATIVE OVULATION IN NONHUMAN PRIMATES1 W.R. Dukelow and J.M.R. Rawson2 Michigan State University Previous studies in our laboratory have demonstrated the use of photolaparography to depict finite changes in follicular morphology during the reproductive cycle of Macaca fascicularis. In recent studies we have attempted to depict the comparative morphology of follicular growth to the time of ovulation and through corpus luteum formation in Saimiri sciureus and Galago senegalensis as well as M. fascicularis. Vascular patterns appear on the follicular surface in all three species within 30 hr of ovulation. The vessels cause a distinct division of the follicle into hemispheres in approximately 80% of the cases with the subsequent development of two translucent areas of the follicle, one of which will represent the point of rupture (stigma). After ovulation the corpus hemorrhagicum is of variable morpholOgy ranging from a discrete single ovulation point to the classical, blood-filled, collapsed follicle seen in rodents. In M: fascicularis the majority of ovulations occur without rupture of the follicular vessels and these persist throughout the establishment of CL. In S: sciureus the CH is more hemorrhagic in appearance than M. fascicularis. 1Presented at the Fed. of Amer. Soc. for Exp. Biol. Meeting, Atlantic City, N.J., April 1972. 2Endocrine Research Unit (Ctr. Lab. Animal Res., Depts. An Hus. 8 Physiol.). 63 NATURAL AND ARTIFICIAL CONTROL OF OVULATION IN NONHUMAN PRIMATES1 W. Richard Dukelow, R.M. Harrison, J.M.R. Rawson and M.P. Johnson2 Michigan State University Ovulation occurs in response to endocrine changes induced by the reproductive cycle, the environment, or both. The present studies outline the progressive development of the follicle in Macaca fascicularis as the moment of ovulation approaches and characterizes morphological variation observed in the follicular vascularity and the formation of the corpora hemorrhagicum. Comparative follicular morphology has been studied by photolaparOgraphic techniques in M: fascicularis, Saimiri sciureus, and Galago senegalensis. Using laparoscopic techniques to predict the time of ovulation, and precise- mating techniques for a limited period of time, pregnancies have been obtained. Because of the increased use of S: sciureus in biomedical research and the strong contrast between native and captive environ- ments (equatorial and temperate) we have studied the effectiveness of an induced ovulation scheme on animals in captivity from 2 weeks to 2 years, as well as the effectiveness of a progestational compound on blocking ovulatory response. Ovulation in M: fascicularis is characterized by distinct changes occurring 24-36 hr before ovulation. These involve bifurcation of the 1Presented at the Third Conference on Experimental Medicine and Surgery in Primates, Lyon, France, June 21-23, 1972. 2Endocrine Research Unit (Ctr. Lab. Animal Res., Depts. An. Hus. G Physiol.). 64 follicle by follicular blood vessels and (8 to 10 hr before ovulation) the appearance of stigmata. At the same time the fimbriated end of the oviduct positions itself adjacent to the deve10ping follicle to facilitate ovum pick-up. Vascular patterns vary from small diffuse veins (S: sciureus) to distinct vessels dividing the follicles into 2 hemispheres (M: fascicularis). The appearance of the corpora hemorrhagica can vary from precise ovulation points and blood-filled follicles to a discrete "patchy" appearance on the follicular wall. Vessel patterns persist through corpora luteum formation. Using the laparoscope and precise-mating techniques, 5 pregnancies have been obtained in M: fascicularis, which occurred between Days 11 and 15 of the cycle (or, on the basis of the luteal phase, 16 to 18 days before the next expected menses). The effectiveness of our ovulation regime in S: sciureus (J. Reprod. Fertil., 22:303, 1970), was found to vary by season of the year with a minimal number of animals ovulating in the summer months. At other times of the year, a good ovulatory response was obtained but could be effectively blocked with injections of 500 ug megestrol acetate (MA) daily. Lower doses inhibited ovulation to a lesser degree. MA could also be administered by subcutaneous silastic implant with a rate of release of approximately 50 ug/day. These studies have bearing on the reproductive physiology of captive nonhuman primates especially relative to their use in the testing of contraceptive compounds. 65 FOLLICULAR DEVELOPMENT AND OVULATION IN MACACA FASCICULARIS, SAIMIRI SCIUREUS AND GALAGO SENEGALENSISr W. Richard Dukelow, D.A. Jewett and J.M.R. Rawson2 Michigan State University During the 24 to 36 hrs before ovulation discrete changes occur in a fixed sequence which allow the prediction of ovulation in macaques. These changes can be observed using photolaparographic techniques we have described earlier (Lab. Animal Sci. 21:594, 1971; Folia Primat. 16:216, 1971) and involve bifurcation of the follicle by blood vessels and (8 to 10 hrs before ovulation) the appearance of stigmata. The fimbriated end of the oviduct positions itself adjacent to the follicle to facilitate ovum recovery. Vascular patterns on the follicular surface vary from stellate, complex patterns to single vessles and from small diffuse veins (8: sciureus) to pronounced vessles (ML fascicularis). Vascular patterns are also seen in G. senegalensis but are less pronounced than in M. fascicularis. The appearance of the corpora hemorrhagica can vary from precise ovulation points and blood-filled follicles to a discrete "patchy" appearance on the follicular wall. Vessel patterns can persist through corpus luteum formation and has been observed on 8 day old corpora lutea. Using laparoscopic prediction of ovulation time one can plan precise matings. Using 20 to 30 minute periods of exposure to the male (M: fascicularis) we have obtained five pregnancies in our colony of 22 females. The pregnancies occurred between days 11 and 15 of the cycle (or, 16 to 18 days before the next expected menses). 1Presented at the Fourth International Congress of Primatology, Portland, Oregon, August 15-18, 1972. 2Endocrine Research Unit (Ctr. Lab. Animal Res., Depts. An. Hus. 5 Physiol.). Name: Born: Birthplace: Formal Education: Degrees Received: VITA Jon M.R. Rawson May 8, 1949 Lansing, Michigan University High School, Carbondale, Illinois 1963-67 Southern Illinois University, Carbondale, Illinois 1967-68 Youngstown State University, Youngstown, Ohio 1968-71 Michigan State University East Lansing, Michigan 1971-present Bachelor of Science, Youngstown State University, 1971 67