Preovulatory Graafian Follicles Research Articles

Characterization of the Management of Select Comorbid Conditions Amongst Cancer Patients

In rabbits, pigs, cows and humans, pre-ovulatory Graafian follicles may be more than 1.0 °C cooler than ovarian stroma and both these ovarian compartments are cooler than deep rectal temperatures. This study examines the effect of follicular cooling on the incidence of pregnancy in dairy cows.Follicular measurements were compiled for cows with one ovulatory follicle (monovular) and cows with one ovulatory follicle per ovary (bi-ovular) and their corresponding uterine horn contents. The study sample consisted of 80 pre-ovulatory follicles in which antral temperatures were measured using a fine thermistor probe.Mean (± S.D.) follicular fluid temperature of the ovulating follicles was 1.12 ± 0.86 °C significantly cooler (P < 0.0001) than rectal temperatures. No significant differences in temperatures were found for non-ovulating follicles. In follicles undergoing cooling (n = 58), a one-tenth of a degree drop in temperature with reference to control rectal temperature gave rise to a 3.6-fold increase (odds ratio) in the pregnancy rate (P = 0.003). The follicle-rectum temperature differential giving rise to pregnancy (n = 18; 1.51 ± 1.15 °C) was significantly greater (P = 0.004) than the differential recorded in cooling follicles at that did not result in a subsequent pregnancy (n = 40; 0.83 ± 0.57 °C).Follicular cooling is needed to trigger ovulation and correlates positively with the potential for pregnancy in cows. This finding has interesting implications for human reproductive medicine.

Pre-ovulatory follicular cooling correlates positively with the potential for pregnancy in dairy cows: Implications for human IVF

IntroductionIn rabbits, pigs, cows and humans, pre-ovulatory Graafian follicles may be more than 1.0 °C cooler than ovarian stroma and both these ovarian compartments are cooler than deep rectal temperatures. This study examines the effect of follicular cooling on the incidence of pregnancy in dairy cows. Material and methodsFollicular measurements were compiled for cows with one ovulatory follicle (monovular) and cows with one ovulatory follicle per ovary (bi-ovular) and their corresponding uterine horn contents. The study sample consisted of 80 pre-ovulatory follicles in which antral temperatures were measured using a fine thermistor probe. ResultsMean (± S.D.) follicular fluid temperature of the ovulating follicles was 1.12 ± 0.86 °C significantly cooler (P < 0.0001) than rectal temperatures. No significant differences in temperatures were found for non-ovulating follicles. In follicles undergoing cooling (n = 58), a one-tenth of a degree drop in temperature with reference to control rectal temperature gave rise to a 3.6-fold increase (odds ratio) in the pregnancy rate (P = 0.003). The follicle-rectum temperature differential giving rise to pregnancy (n = 18; 1.51 ± 1.15 °C) was significantly greater (P = 0.004) than the differential recorded in cooling follicles at that did not result in a subsequent pregnancy (n = 40; 0.83 ± 0.57 °C). ConclusionFollicular cooling is needed to trigger ovulation and correlates positively with the potential for pregnancy in cows. This finding has interesting implications for human reproductive medicine.

Relationship between the monoamine and gonadotropin content in follicular fluid of preovulatory graafian follicles after superovulation treatment.

Noradrenaline (NA), serotonin (5HT), dopamine (DA), FSH, LH and prolactin (PRL) content was determined in 104 preovulatory follicular fluids obtained from 44 patients undergoing in vitro fertilization. The patients were given human menopausal gonadotropin (HMG) for ovarium stimulation, ovulation was induced with 10000 IU human chorionic gonadotropin (HCG) 34-36 hours prior to the follicular aspiration by vaginal ultrasound. Classification of the oocytes was performed by direct microscopic evaluation differentiating three groups of oocytes: Group I.: prophase I; Group II.: metaphase I; Group III.: metaphase II. There was no significant difference in monoamine and FSH content of follicular fluid at different stage of the oocyte maturation. LH and PRL significantly increased parallel with oocyte maturation (38.9; 48.8; 56.7 IU/l and 1324; 2382; 3134 IU/l). significant negative correlation was observed in Group I. between 5HT-LH (r = -0.64); in Group II. between NA-LH (r = -0.62) and NA-PRL (r = -0.51). Significant positive correlation were found in Group I. between FSH-LH (r = 0.63), in Group II. between LH-PRL (r = 0.56), in Group III. between NA-5HT (r = 0.66), NA-DA (r = 0.80) and 5HT-DA (r = 0.66). These observations suggest that action of LH and PRL may be negatively modulated by 5HT and NA in the final stage of oocyte maturation.

Inhibin may be involved in negative feedback in the prairie dog (Cynomys ludovicianus)

The changes in inhibin immunostaining in the gonads during the annual reproductive cycle of both sexes of the prairie dog are described. No inhibin immunostaining was found in primary or secondary follicles of the ovary. Theca and granulosa cells of preovulatory Graafian follicles found in January and February stained for inhibin. Corpora lutea of both pregnant and non-pregnant females stain more densely for inhibin than follicles. Inhibin staining is present in luteal cells for at least 4 months during regression, longer than detectable progesterone is secreted. Sertoli cells in the testes do not have inhibin immunostaining during recrudescence. These cells show light immunostain for inhibin during peak spermatogenic activity in January and February but stain more deeply during early regression of the testis. Stain is gradually lost in the next 4-5 months as the tubules close. Leydig cells and germ cells do not stain for inhibin at any stage of the annual cycle but interstitial cells and tunic cells stain during the breeding phase. The presence of immunochemical staining for inhibin in prairie dog gonads during regression suggests that inhibin is part of a negative feedback complex that includes progesterone in the female and testosterone or another androgen in the male. Negative feedback during regression may also cause gonadal inactivity.

Physiological routes from intra-uterine seminal contents to advancement of ovulation

Whole boar semen or seminal plasma has been demonstrated to advance the time of ovulation in gilts. As a means of clarifying this influence, the contribution of uterine lymphatics and their white cell populations has been examined. After duct visualisation with Evan's blue, lymph was sampled from a mesometrial vessel in eight pre-ovulatory gilts whose uterine lumen was infused simultaneously with whole semen in one ligated horn and saline in the contralateral ligated horn. Lymph was collected from cannulated vessels for periods of up to four hours under general anaesthesia. Thereafter, mesometrial lymph nodes, utero-tubal junction and uterine wall tissues were sampled. The proportion of nucleated cells in the sampled lymph increased towards the end of the collection period, but erythrocytes were found in all instances preventing a meaningful differentiation and identification of leukocytes. Prominent uterine lymph nodes were present in the mesometrium on both sides of the reproductive tract in 7 of 10 gilts. Differences in cellular contents were demonstrated between the side of the tract infused with semen and that infused with saline control. Two of 4 gilts had lower values for CD4 (Cluster Differentiation) and 3 of 6 gilts higher values for MHC II (Major Histocompatibility Complex) markers on the side challenged with semen. In contrast, values remained constant for CD8 but ranged widely for CD18. Immunohistochemical analysis of uterine tissue samples for MHC II+ cells revealed significant differences (P < 0.05) between the control and semen-treated ligated portions of the horns, as well as between the tissue sample of uterine wall and that from the utero-tubal junction, but there were no significant differences for CD4+ cells. It therefore remains plausible that semen-induced cytokines in the uterine lymph undergo counter-current transfer to the ipsilateral ovary and accelerate the final maturation of pre-ovulatory Graafian follicles.

Ovarian follicular development in cattle selected for twin ovulations and births.

Comparisons of numbers of antral ovarian follicles and corpora lutea (CL), of blood hormone concentrations, and of follicular fluid steroid concentrations and IGFBP activity were conducted between cows selected (twinner) and unselected (control) for twin births to elucidate genetic differences in the regulation of ovarian follicular development. Ovarian follicular development was synchronized among cows by a single i.m. injection of PGF2alpha on d 18 of the estrous cycle; six cows per population were slaughtered at 0, 24, 48, and 72 h after PGF2alpha. Jugular vein blood was collected from each animal at PGF2alpha injection and at 24-h intervals until slaughter. Ovaries of twinner cows contained more small (< or = 5 mm in diameter, P < 0.05), medium (5.1 to 9.9 mm, P < 0.05), and large (> or = 10.0 mm, P < 0.01) follicles and more (P < 0.01) CL than ovaries of controls. Follicular fluid concentrations of estradiol, androstenedione, testosterone, and progesterone reflected the stage of follicular development and were similar for twinner and control follicles at the same stage. Earlier initiation of follicular development and/or selection of twin-dominant follicles in some twinner cows resulted in greater concentrations of estradiol in plasma at 0, 24, and 48 h and of estradiol, androstenedione, and testosterone in follicular fluid of large follicles at 0 h after PGF2alpha for twinner vs. control cows (follicular status x time x population, P < 0.01). Binding activities of IGFBP-5 and -4 were absent or reduced (P < 0.01) in follicular fluid of developing medium and large estro-gen-active (estradiol:progesterone ratio > 1) follicles but increased with atresia. Only preovulatory Graafian follicles lacked IGFBP-2 binding, suggesting a possible role for IGFBP-2 in selection of the dominant follicle. Concentrations of IGF-I were twofold greater (P < 0.01), but GH (P = 0.10) and cholesterol (P < 0.05) were less in blood of twinners. Three generations of selection of cattle for twin ovulations and births enhanced ovarian follicular development as manifested by increased numbers of follicles within a follicular wave and subsequent selection of twin dominant follicles. Because gonadotropin secretion and ovarian steroidogenesis were similar for control and twinner cattle, enhanced follicular development in twinners may result from decreased inhibition by the dominant follicle(s), increased ovarian sensitivity to gonadotropins, and/or increased intragonadal stimulation, possibly by increased IGF-I.

Ultrastructure and morphometry of ovarian follicles in the armadillo Chaetophractus villosus (Mammalia, Dasypodidae).

Ultrastructural and morphometric changes in oocyte and surrounding granulosa cells during armadillo follicular growth are described. Primordial, intermediary, early and late primary, secondary, tertiary and preovulatory Graafian follicles were found. From primordial to Graafian follicle, granulosa cells increase in height, become multilayered while fluid-filled spaces arise among them in the tertiary follicle stage. As the follicle expands the oocyte is located eccentrically and grows in size. From primordial to secondary stage organelles develop in number and complexity and many vesicles start arising in the cytoplasm at the secondary stage. From tertiary follicle stage the number of mitochondria decreases. In follicular cells, secreting-related organelles expand, but mitochondrial number decreases according to the follicle expansion. Intercellular junctions between both cell types increase. The morphometrical study shows a biphasic pattern of oocyte and follicle growth. The morphological baseline here provided will be essential for further comprehension of the reproductive biology in armadillos.

Pre-ovulatory graafian follicles are cooler than neighbouring stroma in pig ovaries.

Using an infra-red camera, domestic pig ovaries were thermo-imaged almost instantaneously at laparotomy or within a closed abdomen by endoscopy. Rectal and jugular vein temperatures were recorded using thermo-probes. Graafian follicles (7-10 mm diameter) were cooler than ovarian stroma in all experimental models examined, and both compartments were cooler than rectal and jugular temperatures. The mean difference between follicles and stroma in 73 observations was 1.3 +/- 0.1 degrees C. When thermo-imaged under the fimbriated extremity of the Fallopian tube, follicles and stroma could still be distinguished. Follicles cooled slightly more rapidly than adjoining stroma during the first 10 s of a 60 s recording interval, after which curves for the two tissues remained parallel. Arresting ovarian blood supply for 5 min had a negligible influence on the temperature differentials. Endoscopy in three models recorded mean differentials between follicles and stroma of 0.6 +/- 0.1 degrees C to 1.1 +/- 0.1 degrees C. It is concluded that temperature gradients do exist in the ovarian tissues of mature animals, and that these are generated at least in part as a consequence of endothermic reactions within Graafian follicles.

Ovarian control of very low sperm/egg ratios at the commencement of mammalian fertilisation to avoid polyspermy

This essay considers the means whereby sperm/egg ratios close to unity are generated during the initial stages of fertilisation in placental mammals. Pre-ovulatory graafian follicles and their contents are seen to be key structures orchestrating the events of sperm progression and coordinating the subsequent meeting of male and female gametes. Three levels of control over the numbers of spermatozoa activated and released from the functional reservoir in the caudal region of the fallopian tube isthmus are proposed. A primary control would be obtained by means of a countercurrent transfer of ovarian follicular progesterone from the ovarian vein into the tubal branch of the ovarian artery. The concentration of progesterone so transferred would be proportional to the number of preovulatory follicles, and thus to the number of eggs to be shed, and would act progressively to reduce sperm binding to the endosalpinx of the caudal isthmus. Differential timing of the release from epithelial binding may be a crucial means of achieving the initial low sperm/egg ratios. a secondary regulation of the release of graded numbers of viable spermatozoa towards the ampullary-isthmic junction of the fallopian tubes would be by means of molecular messages derived from the mucified oocyte-cumulus complex shortly before and after the time of ovulation. Third would be reorientation of sperm trajectories by molecular gradients within the cumulus cell mass to direct competent spermatozoa to those oocytes as yet unpenetrated. Together these differing levels of control would impose low sperm/egg ratios during the initial stages of fertilisation, such strict quantitative regulation of male gametes lasting at least until the block to polyspermy is fully established and the vitellus is no longer at risk from further sperm penetration.

Ovarian control of very low sperm/egg ratios at the commencement of mammalian fertilisation to avoid polyspermy

This essay considers the means whereby sperm/egg ratios close to unity are generated during the initial stages of fertilisation in placental mammals. Pre-ovulatory graafian follicles and their contents are seen to be key structures orchestrating the events of sperm progression and coordinating the subsequent meeting of male and female gametes. Three levels of control over the numbers of spermatozoa activated and released from the functional reservoir in the caudal region of the fallopian tube isthmus are proposed. A primary control would be obtained by means of a countercurrent transfer of ovarian follicular progesterone from the ovarian vein into the tubal branch of the ovarian artery. The concentration of progesterone so transferred would be proportional to the number of preovulatory follicles, and thus to the number of eggs to be shed, and would act progressively to reduce sperm binding to the endosalpinx of the caudal isthmus. Differential timing of the release from epithelial binding may be a crucial means of achieving the initial low sperm/egg ratios. A secondary regulation of the release of graded numbers of viable spermatozoa towards the ampullary-isthmic junction of the fallopian tubes would be by means of molecular messages derived from the mucified oocyte-cumulus complex shortly before and after the time of ovulation. Third would be reorientation of sperm trajectories by molecular gradients within the cumulus cell mass to direct competent spermatozoa to those oocytes as yet unpenetrated. Together these differing levels of control would impose low sperm/egg ratios during the initial stages of fertilisation, such strict quantitative regulation of male gametes lasting at least until the block to polyspermy is fully established and the vitellus is no longer at risk from further sperm penetration. © 1996 Wiley-Liss, Inc.

Relationship between the monoamine, progesterone and estradiol content in follicular fluid of preovulatory graafian follicles after superovulation treatment.

We measured the progesterone, estradiol (E2), serotonin (5-HT), noradrenaline (NA) and dopamine (DA) contents of follicular fluid (FF) samples obtained from 35 patients undergoing in vitro fertilization and embryo transfer. Progesterone and E2 were determined by radioimmunoassay, and monoamines were measured by a spectrofluorimetric method. Significantly higher progesterone and NA levels were found (p < 0.01) in FF from cycles in which the oocyte cleaved and resulted in pregnancy compared to FF containing uncleaved oocytes. NA and DA contents were significantly higher in FF from cycles resulting in pregnancy than in FF containing cleaved oocytes not resulting in pregnancy (p < 0.01). There were significantly lower NA and 5-HT levels in FF containing uncleaved oocytes compared to the latter group (p < 0.05). Significant positive correlations were found between progesterone and 5-HT (r = 0.67) and between NA and DA (r = 0.93) in the pregnant group; between progesterone and E2 (r = 0.84) in FF containing uncleaved oocytes; between progesterone and E2 (r = 0.71) and between NA and DA (r = 0.62) in FF from cycles in which the oocyte cleaved but did not result in pregnancy. The results suggest that the follicular hormonal changes associated with oocyte maturation may be locally modulated by monoamines.

Immunohistochemical studies on the localization of aromatase and 17 alpha-hydroxylase/C17-20 lyase (17 alpha-lyase) in estrous cycling and pregnant hamster ovaries.

In order to clarify periodic changes in the localization of enzymes engaged in estrogen biosynthesis during the estrous cycle, immunohistochemical and fine structural studies were performed using estrous cycling and pregnant hamster ovaries. Results showed that ovulation takes place at midnight between Day 4 and Day 1 in the regular 4 day-cycle hamster. Immunoreactivity for aromatase is localized in the granulosa cells of the secondary follicle and granulosa lutein cells during the morning (10:00 am) of Day 1 to the evening (5:00 pm) of Day 4; in the night (9:00 pm) of Day 4, only the granulosa cells of the Graafian follicle showed a strong immunoreaction. As for 17 alpha-lyase, theca interna cells of the secondary follicle are immunopositive throughout Day 2 to the morning (10:00 am) of Day 4. Only a few cells in the theca interna of the Graafian follicles are immunopositive in the evening (5:00 pm) of Day 4. No positive cells for this enzyme were detected in the night (9:00 pm) of Day 4 or morning (10:00 am) of Day 1. The rapid decrease of estrogen biosynthesis occurring just before ovulation is considered to be due to the disappearance of 17 alpha-lyase in the theca interna cells of the ovary. On Day 10 of pregnancy, the granulosa cells of the secondary follicles and both the granulosa and theca lutein cells of the corpora lutea are immunostained with the aromatase antibody, while the theca interna cells of the secondary follicles reacted positively to the 17 alpha-lyase antibody. Only the granulosa and theca interna cells from the large preovulatory Graafian follicle of Day 4 (proestrus) which are positively stained for aromatase as well as 17 alpha-lyase show ultrastructural features typical of steroid secretory cells.

Rat ovarian apolipoprotein E: localization and gonadotropic control of messenger RNA.

Apolipoprotein E (apo E) is a 35-kDa protein found in association with various lipoproteins. It is synthesized by a wide variety of tissues, including the ovary. It can serve several functions, such as 1) transport of excess cholesterol from peripheral tissue to the liver; 2) directed movement of cholesterol from areas of high to low cholesterol concentration within tissue or organs; and 3) inhibition of the conversion of theca progesterone to androgen, thus acting as an autocrine or paracrine factor within the ovary. To better understand the physiological role of ovarian apo E, we employed the technique of in situ hybridization utilizing 35S-labeled apo E riboprobes to identify cells containing E mRNA. We studied ovaries of hypophysectomized immature rats administered various regimens of gonadotropins because of the uniform, predictable stimulation of follicular granulosa and theca development, ovulation, and corpus luteum formation. Apo E mRNA was localized predominantly in the theca, with an increase associated with theca hypertrophy. Apo E mRNA increased in granulosa cells with the development of preovulatory Graafian follicles, but decreased to baseline following ovulation and corpus luteum formation. These data are consistent with two roles for apo E in the ovary: 1) directing cholesterol to cells needing cholesterol as substrate for cell proliferation and steroidogenesis, and 2) acting as an autocrine regulatory factor to reduce theca androgen substrate for follicle estrogen production.

Characterization of ovarian follicular fluids of sheep, pigs and cows using proton nuclear magnetic resonance spectroscopy

Proton NMR spectra were produced for Graafian follicular fluids obtained by aspiration from sheep, pig and cow ovaries. The following low molecular mass, non-protein-bound metabolites were detected at concentrations exceeding 0.1 mM: acetate, alanine, creatinine/creatine, glycine, D-3-hydroxybutyrate, lactate, valine. Glucose was difficult to quantify and N-acetyl sugars gave a broad resonance at 2.06 ppm, presumably representing side-chains of glycoproteins. Ethanol was detected at up to millimolar concentrations in some specimens, though the physiological significance of this finding was not clear. The concentrations of all metabolites were comparable to those of plasma. These results have therefore shown that NMR spectroscopy is useful for gaining a broad and semiquantitative impression of the more abundant metabolites in the fluids of preovulatory Graafian follicles.

Variations in the morphology and Ca2+-ATPase activity of the ovarian surface epithelium in rats.

The ultrastructure and cytochemical localization of Ca2+-activated adenosine triphosphatase (Ca2+-ATPase) activity of the ovarian surface epithelium in immature rats and in adult rats during the estrous cycle and pregnancy were investigated at both light and electron microscopical levels. In the adult rat ovaries at all stages of the estrous cycle, the intensity of Ca2+-ATPase activity and ultrastructure of the surface epithelium were considerably varied in the different regions on the same ovary. The variations in the morphology and Ca2+-ATPase activity likely provide evidence for the differences in the functional activity of the surface epithelium, which are related to the topographical connection with adjacent structures of ovarian cortex, i.e., corpus luteum, preovulatory Graafian follicles, interstitium containing small follicles. In the adult rat ovaries at all reproductive stages, the cuboidal or columnar surface epithelial cells were found on the interstitium containing small follicles. In some places, the columnar surface epithelial cells, forming crypts of the epithelium, invaginated into the ovarian proper. This type of surface epithelial cell had well-developed microvilli on its apical membrane, and showed intense Ca2+-ATPase activity on its apical and lateral membranes. The squamous surface epithelial cells were found on the surface of corpus luteum and preovulatory Graafian follicles. These cells had a low number of microvilli and less activity of Ca2+-ATPase on the apical membrane. In the immature rat ovaries, the surface epithelial cells were in part squamous, in part cuboidal. Both types of surface epithelial cells had few microvilli. Ca2+-ATPase activity on the apical membrane was considerably less on all surfaces of the ovaries. It seems that the transport function of the surface epithelium in the immature ovary is not active as compared with the mature ovary.These results suggest that the surface epithelium may have an active function of local absorption and/or transepithelial transport between pelvic cavity and ovarian proper, and its functional activity is associated with the reproductive function of the ovary.

Ovarian lipoxygenase activity and its regulation by gonadotropin in the rat

In our previous study of a dose-dependent blockage of follicular rupture at ovulation by inhibitors of lipoxygenase was demonstrated. Here the presence of 5-lipoxygenase activity in the whole ovary and in the Graafian follicle is estimated by a chemiluminescence assay using unlabeled arachidonic acid as susbtrate in the presence of luminol and by conversion of 14C-arachidonic acid into lipoxygenase products as separated by HPLC. Both approaches demonstrated lipoxygenase activity in whole ovarian homogenates and in homogenates of preovulatory Graafian follicles. Furthermore, within 6 h after stimulation in vivo with hCG, lipoxygenase activity was increased by 2-fold in the whole ovarian homogenate and by 5-fold in the follicular homogenate. These results confirm the presence of lipoxygenase in rat ovaries, and its stimulation by gonadotropin and thus corroborate the suggested involvement of lipoxygenase products in follicular rupture at ovulation.

Oestradiol in follicular fluid and in utero-ovarian venous and peripheral plasma during parturition and postpartum oestrus in the tammar, Macropus eugenii.

Oestradiol-17 beta was measured in plasma and follicular fluid by a radioimmunoassay validated for the tammar. Plasma was sampled from the peripheral circulation and both utero-ovarian veins to assess the contribution of oestradiol from the corpus luteum in one ovary and from the Graafian follicle in the other during late pregnancy, parturition, post-partum oestrus and ovulation. Concentrations of oestradiol in the peripheral plasma and in the vein draining the CL remained less than 5 pg/ml on all days except the day of parturition, when they were 12.7 pg/ml and 14.3 pg/ml respectively. Mean oestradiol concentration in the vein draining the Graafian follicle was significantly higher than the mean peripheral concentration on Days 24-27. The highest average concentration (36.5 pg/ml) occurred on Day 27 coincident with parturition and before oestrus. These latter concentrations were significantly correlated with the concentration and total oestradiol in the fluid of the preovulatory Graafian follicle (r = 0.73 and r = 0.82 respectively), thus providing strong evidence that the Graafian follicle is the main source of oestradiol in the peripheral circulation at this time.

Progesterone secretion by highly differentiated human granulosa cells isolated from preovulatory Graafian follicles induced by exogenous gonadotropins and human chorionic gonadotropin.

Human granulosa-luteal cells were harvested from preovulatory Graafian follicles at the time of oocyte retrieval for in vitro fertilization after induction of follicle maturation by sequential injections of menopausal gonadotropins and hCG. Such highly differentiated granulosa cells produced large quantities of progesterone basally (6.8 pg/cell X 2 days) in monolayer culture. Human LH significantly increased progesterone biosynthesis after 6, 12, 48, 96, or 144 h in culture, with a maximal increase of 8- to 20-fold occurring at 96 h. The stimulatory effect of LH could be observed under serum-free conditions and was maximal in the presence of 4% serum. Human granulosa-luteal cells also exhibited significant stimulatory responses to hCG, prostaglandin E2, or the cAMP effectors 8-bromo cAMP, choleratoxin, or forskolin in serum-free incubations. Concentrations of 17 beta-estradiol that are attained physiologically in ovarian follicles in vivo markedly suppressed basal and LH (or cAMP)-stimulated progesterone production in vitro (maximal suppression, greater than 90%). The nonaromatizable androgen 5 alpha-dihydrotestosterone also inhibited progesterone production, but by no more than 45-50% even at supraphysiological concentrations. Estradiol's blockade of progesterone synthesis was associated with a corresponding increase in pregnenolone accumulation. The present studies indicate that human granulosa-luteal cells isolated from preovulatory follicles induced with exogenous gonadotropins and hCG secrete large quantities of progesterone in vitro. Such cells retain stimulatory responses to human LH, hCG, prostaglandin E2, and classical cAMP effectors in serum-free incubations. Moreover, physiological concentrations of 17 beta-estradiol suppress progesterone production, probably by inhibiting cellular conversion of pregnenolone to progesterone. Thus, the present in vitro system permits an investigation of hormone action in well differentiated, human granulosa-luteal cells isolated from preovulatory Graafian follicles that have a defined endocrine history of prior gonadotropin exposure in vivo.

Epinephrine-sensitive adenylyl cyclase activity in rabbit ovarian tissues.

Epinephrine-sensitive adenylyl cyclase (AC) activity was investigated in homogenates of rabbit ovarian tissues. Optimal epinephrine-stimulated AC activity in corpora lutea homogenates was achieved with millimolar concentrations of ATP (1-4 mM) and Mg2+ (2-12 mM) in the absence of exogenously added Ca2+. Epinephrine stimulated the luteal AC under these conditions in a dose-dependent manner, with an ED50 of 0.45 micrograms/ml. The AC of preovulatory follicle homogenates was found to be unresponsive to epinephrine stimulation. Epinephrine responsiveness of the AC slowly appeared in late preovulatory and early postovulatory follicles in response to the gonadotropin surge induced by coitus. By 18 h after mating, epinephrine-stimulated AC activity had doubled, and maximal activities in the newly formed corpora lutea were reached within 72 h. Thereafter, epinephrine-stimulated AC activity gradually declined to a relatively stable value that was maintained throughout the remainder of gestation, except for two transient increases in activity on days 14 and 19 of pregnancy. With parturition, luteal epinephrine-stimulated AC activity fell to basal levels. In corpora lutea of pseudopregnancy, epinephrine-stimulated AC activity was very similar to that of corpora lutea of pregnancy during the first 16 days but fell to basal levels on day 17 with the termination of pseudopregnancy. Neither corpora albicans nor interstitial tissue exhibited any epinephrine-stimulated AC activity. The presence of epinephrine-stimulated AC activity only in corpora lutea of pseudopregnancy and pregnancy and not in preovulatory Graafian follicles, corpora albicans, or interstitial tissue suggests a functional to role for catecholamines in luteal tissue. Moreover, these data indicate that the presence of an epinephrine-sensitive AC may be used as a functional marker for luteinization, at least in rabbit ovarian tissues.

Female Reproductive Anatomy and Development of Ovarian Follicles inMiniopterus Fraterculus

The uterus of Miniopterus fraterculus (Thomas & Swan 1906) is bicornuate and asymmetrical, the right uterine horn being larger than the left in parous and nulliparous females. Ovarian and uterine function is asymmetrical; 94% of observed ovulations originated in the left ovary and all implantations occurred in the right uterine horn. Growth and development of ovarian follicles follow the typical eutherian mammalian pattern. The preovulatory Graafian follicle differs from that of most hibernating verspertilionids in that the cumulus oophorus is relatively small. Two types of follicular atresia occur. The first type, in which the stratum granulosum degenerates before degeneration of the oocyte, occurs in multilaminar follicles. The second type occurs in primary and early secondary follicles and is characterized by the oocyte showing the first signs of regression.