Quantitative ultrastructure of the luteal cell of the 16-day-pregnant rat and its relation to progesterone secretion

The ultrastructure of luteal cells of five Day-16 pregnant rats were examined morphometrically to determine the relationship between the quantity of steroidogenic organelles and membranes and reported rates of progesterone secretion (2.3 micrograms/h). Each rat had 11.8 +/- 1.0 corpora lutea (mean +/- s.e.m.) with an average volume of 4.5 +/- 0.1 microliter. There were 210 000 +/- 10 000 luteal cells per CL and the luteal cell cytoplasm was composed of smooth endoplasmic reticulum (18%), mitochondria (10.6%), lipid droplets (8.9%) and granules (0.6%). The surface area of the smooth endoplasmic reticulum was 192 cm2 per CL, and that of the outer and inner mitochondrial membranes was 20 and 34 cm2, respectively. For each square micrometre of these membranes, respectively, 62, 590 and 355 molecules of progesterone would have been secreted per second. The luteal cell appears to secrete its major steroid hormone at a rate 50 times greater than that reported for the Leydig cell of the testis when secretion is expressed in terms of molecules per unit mass of steroidogenic cell or area of steroidogenic membrane.     

Characterization of large luteal cells and their secretory granules during the estrous cycle of the cow.

Large steroidogenic cells of the bovine corpora lutea were evaluated for morphological changes on Days 3, 7, 11, 14, 17, and 19 of the estrous cycle. Large cells were readily identified by size (25-50 microns diameter), numerous mitochondria, and the presence of dense secretory granules (150-300 nm in diameter). These granules were found in a discrete cluster and were not dispersed throughout the cytoplasm. Only 3% of the large cells contained a cluster of granules on Day 3. The percentage was highest during midcycle (Day 7, 84%; Day 11, 64%), dropped on Day 14 (26%), and was lowest on Days 17 (16%) and 19 (8%). Electron microscopic immunocytochemistry showed that oxytocin and neurophysin were co-localized in these granules on all days evaluated. As early as Day 14, large cells were observed with characteristics typical of regressing corpora lutea, i.e., a reduction in cells with secretory granules, large cytoplasmic lipid droplets, and swollen mitochondria with dense inclusions. However, since this was a time of the cycle when plasma concentrations of progesterone were very high, this corpus luteum is referred to as involutive rather than regressive. Our results may be summarized as follows: 1) from Day 7 to Day 14 there was a 69% decline in the number of large cells containing oxytocin-laden secretory granules. This occurred prior to the rise in uterine oxytocin receptors and the large luteolytic pulses of prostaglandin that reportedly occur after Day 14. The role of this apparent early release of oxytocin is not known. 2) Large steroidogenic luteal cells of the estrous cycle have morphological characteristics similar to those of large luteal cells during pregnancy. However, large luteal cells of the estrous cycle contain oxytocin whereas those of pregnancy are devoid of oxytocin.     

Size distribution of luteal cells from pregnant and non-pregnant marmoset monkeys and a comparison of the morphology of marmoset luteal cells with those from the human corpus luteum

The size distribution of marmoset luteal cells was determined on Days 6, 14 and 20 after ovulation in non-pregnant cycles and in early pregnancy. Image analysis was used to estimate the cell diameter of dispersed cells prepared from the marmoset corpus luteum (CL). Steroidogenic cells showed a size distribution consistent with one population of cells. There was a significant increase in mean cell diameter (P less than 0.05) from Day 6 to Day 14 in pregnant and non-pregnant animals with no further increase on Day 20. Micrographs of marmoset luteal tissue showed cells of greater than 10 micron containing the organelles typical of steroid-producing cells, and smaller non-steroidogenic cells surrounding the steroid-producing cells. On the basis of microscopy, there were no areas within the CL where cell composition was noticeably different. In contrast, micrographs of human luteal tissue showed two types of steroidogenic cell; most cells were similar to those in the marmoset CL but a smaller population of smaller cells could be distinguished around the periphery and along vascular septa. It is likely that these smaller and larger types of steroidogenic cells are of theca and granulosa cell origin respectively, the two cell populations differing in the degree of electron density and amount of rough endoplasmic reticulum. A distinguishing feature between marmoset and human luteal cells was the shape of the mitochondrian which were considerably rounder in marmoset luteal cells. The origin of steroidogenic cells in the marmoset CL is unclear, although in marmosets and man the luteal cell types display morphological characteristics distinct from the large and small luteal cells described for CL of the domestic ungulates.     

Endocrine profiles during the estrous cycle and pregnancy in the Baird's tapir (Tapirus bairdii)

Serum samples were collected 1–3 times weekly from two Baird's tapirs (Tapirus bairdii) for 6 months in 1987–1988, and for more than 3 consecutive years beginning in 1989 to characterize hormone patterns during the estrous cycle and pregnancy. Based on serum progesterone concentrations, mean (±SEM) duration of the estrous cycle (n = 20) was 30.8 ± 2.6 days (range, 25–38 days) with a luteal phase length of 18.1 ± 0.4 days (range, 15–20 days). Mean peak serum progesterone concentrations during the luteal phase were 1.35 ± 0.16 ng/ml, and nadir concentrations were 0.19 ± 0.03 ng/ml during the interluteal period. Distinct surges of estradiol preceded luteal phase progesterone increases in most (14/20) cycles. Gestation length was 392 ± 4 days for three complete pregnancies. Mean serum progesterone concentrations increased throughout gestation and were 1.83 ± 0.13, 2.73 ± 0.13, and 4.30 ± 0.16 ng/ml during early, mid- and late gestation, respectively. Serum estradiol concentrations began to rise during mid-gestation, increasing dramatically during the last week of pregnancy. Patterns of serum estriol and estrone secretion during pregnancy were similar to that observed for estradiol. In contrast to progesterone and estrogens, serum cortisol concentrations were unchanged during pregnancy or parturition. Females resumed cycling 16.2 ± 2.0 days after parturition (n = 4) and, on two occasions, females became pregnant during the first postpartum estrus. These data suggest that the tapir cycles at approximately monthly intervals and that increases in serum progesterone are indicative of luteal activity. The interluteal period is relatively long, comprising approximately 40% of the estrous cycle. During gestation, progesterone concentrations are increased above luteal phase levels, and there is evidence of increased estrogen production during late gestation. The absence of increased cortisol secretion at the end of gestation suggests that this steroid does not play a major role in initiating parturition in this species. © 1994 Wiley-Liss, Inc.     

Functional differences between small and large luteal cells of the late-pregnant vs. nonpregnant cow

This paper describes an in vitro model for the study of two types of steroidogenic luteal cells from cows in different physiological states. Two different populations of enzymatically dispersed bovine luteal cells were separated on the basis of size in a Cel-Sep Sedimentation Chamber. The separated small (12.5-23 micron in diameter) and large (greater than 23 micron in diameter) luteal cells of late-pregnant cows (Days 190-280) contained the distinct morphological characteristics previously defined for these two populations of cells. Cells were evaluated for progesterone (P4) production during a 3-h incubation with and without bovine luteinizing hormone (bLH, 10 ng/ml). Both small and large luteal cells from the late-pregnant cow were found to contain equal levels of P4 at Time 0 and increased but equal levels of P4 after a 3-h incubation. Neither cell type showed an increase in P4 production in response to the addition of bLH (p greater than 0.05). Since these results differed from earlier reports for luteal cells of the nonpregnant cow, small and large luteal cells of the mid-cycle (Day 14) were incubated, and the levels of P4 production were compared with P4 levels from the late pregnant cow. In agreement with previous reports for nonpregnant cows, progesterone content at Time 0 was 7-fold higher in large cells than in small cells (p less than 0.05), and after 3 h of incubation, 13-fold higher (p less than 0.05). Although the small cells responded to the presence of bLH in the incubation medium with a 4-fold increase in P4 production, this increase was not significant (p greater than 0.05). The large cell did not respond to bLH. However, the large cell type continued to contain and produce more P4 than did the small cells treated with bLH. This study indicates that both the small and large luteal cells of late-pregnancy are able to produce P4. However, the large luteal cell of the estrous cycle produces greater quantities of P4 than does the small luteal cell or the large luteal cell of late pregnancy.     

Changes in activities of superoxide dismutase, nitric oxide synthase, glutathione-dependent enzymes and the incidence of apoptosis in sheep corpus luteum during the estrous cycle

Anti-oxidative enzymes play a role in protecting cells from oxidative stress-induced cell death. The present study was conducted to evaluate whether the anti-oxidant and pro-oxidant enzymatic capacities of the sheep corpus luteum (CL) are correlated with steroidogenic and structural status of the gland during the estrous cycle. Steroidogenic activity, apoptosis and superoxide dismutase (SOD1 and SOD2), nitric oxide synthase (NOS), glutathione peroxidase (GPX), glutathione reductase (GSR) and glutathione S-transferase (GST) activities were determined in the CL at specific developmental stages of the luteal phase. The intensity of apoptotic DNA fragmentation, characteristic of physiological cell death, was much greater in CL at late luteal phase than at early and mid-luteal phase, concomitantly with the diminution in the plasma progesterone concentrations from mid-to late luteal phase. SOD1 and GPX activities increased from early to mid-luteal phase, and increased further at late luteal phase. SOD2 and GST activities were not different between early and mid-luteal phase, but increased at late luteal phase. GSR activity was not different between any luteal phase examined. NOS activity decreased from early to mid- and late luteal phase. These results show that the activities of SOD1, SOD2, NOS, GPX, GSR and GST in the sheep CL are subject to major changes during the estrous cycle, and that the anti-oxidant and pro-oxidant enzymatic capacities of luteal cells are not correlated with cell steroidogenic status and integrity during the late luteal phase.     

Immunocytochemical localization of relaxin in human corpora lutea: cellular and subcellular distribution and dependence on reproductive state

Relaxin is one of the hormones present during pregnancy and it is synthesized primarily by corpora lutea (CL). Other reproductive tissues including CL of the menstrual cycle may also synthesize this hormone. Very little is known, however, about the cellular and subcellular distribution of relaxin in human CL and dependence of luteal relaxin on the reproductive state. The light and electron microscope immunocytochemical studies described here were undertaken to obtain this information using antisera to porcine and human relaxin. Immunostaining was found in large luteal cells (17-30 microns) but not in small luteal cells (7-16 microns) or in nonluteal cells in any of the reproductive states or in human hepatocytes. Luteal immunostaining was low in early luteal phase; it increased progressively, reaching the highest level in late luteal phase, and then decreased greatly in corpora albicantia. Term pregnancy CL contained similar immunostaining as early luteal phase CL. Mid luteal phase CL contained more immunostained cells than late luteal phase CL, but the late luteal phase CL contained a greater amount of immunostaining per cell than mid luteal phase CL. The immunogold particles due to relaxin were primarily present in secretory granules and to a small extent in rough endoplasmic reticulum. Quantitation revealed that secretory granules contained a much higher number of gold particles than did rough endoplasmic reticulum. These two organelles from late luteal phase CL contained greater numbers of gold particles than those from mid luteal phase.(ABSTRACT TRUNCATED AT 250 WORDS)     

Changes in the distribution of sizes of ovine luteal cells during the estrous cycle

The ovine corpus luteum is composed of two types of steroidogenic cells, which are referred to as small and large luteal cells. In this study, the size and number of steroidogenic cells were determined in corpora lutea collected on Days 4, 8, 12, and 16 of the estrous cycle. Corpora lutea were dissociated into single-cell suspensions that were stained for 3 beta-hydroxysteroid dehydrogenase (3 beta-HSD) activity, a marker for steroidogenic cells. The size of 3 beta-HSD-positive cells was measured with a Zeiss Videoplan Image Analyzer. On Day 4, most of the 3 beta-HSD-positive cells were less than 18 microns in diameter, the median being 11.2 microns. By Day 8, the number of 3 beta-HSD-positive cells increased 3-fold, and the median diameter increased to 12.8 microns. Although the number of 3 beta-HSD-positive cells was reduced by approximately 50% on Day 16, the median size on Days 12 and 16 was 14.6 and 16.8 microns, respectively. The ratio of large (greater than 18 microns) to small (less than 18 microns) luteal cells was 0.11 +/- 0.03 on Day 4; the ratio increased linearly to 0.67 +/- 0.09 by Day 16. This increase between Days 4 and 12 was attributable to an overall increase in the size of the cells; the increase between Days 12 and 16, however, was due to a loss of small luteal cells. When the experiment was conducted near the end of the breeding season, before animals became anestrous, the median size of the luteal cells did not change at different times of the estrous cycle but remained constant throughout. These data suggest that development of the corpus luteum is associated with an increase in the size and number of steroidogenic luteal cells, and that luteolysis is associated with a preferential loss of small luteal cells.     

Secretory Pattern of Inhibin During Estrous Cycle and Pregnancy in African (Loxodonta africana) and Asian (Elephas maximus) Elephants

The ovary of female elephants has multiple corpora lutea (CL) during the estrous cycle and gestation. The previous reports clearly demonstrated that inhibin was secreted from lutein cells as well as granulosa cells of antral follicles in cyclic Asian elephants. The aim of this study is to investigate the inhibin secretion during the pregnancy in African and Asian elephants. Two African elephants and two Asian elephants were subjected to this study. Circulating levels of immunoreactive (ir‐) inhibin and progesterone were measured by radioimmunoassay. Four pregnant periods of an African elephant and three pregnant periods of an Asian elephant were analyzed in this study. Circulating levels of ir‐inhibin started to increase at 1 or 2 week before the ovulation and reached the peak level 3 or 4 weeks earlier than progesterone during the estrous cycle in both African and Asian elephants. After last luteal phase, the serum levels of ir‐inhibin remained low throughout pregnancy in both an African and an Asian elephant. The mean levels of ir‐inhibin during the pregnancy were lower than the luteal phase in the estrous cycle despite high progesterone levels were maintained throughout the pregnancy. These results strongly suggest that CL secrete a large amount of progesterone but not inhibin during the pregnancy in elephants. Zoo Biol 31:511‐522, 2012. © 2011 Wiley Periodicals, Inc.     

The preovulatory rise of ovarian ornithine decarboxylase is required for progesterone secretion by the corpus luteum

Ovarian progesterone secretion during the diestrus stage of the estrous cycle is produced by luteal cells derived from granulosa and thecal cells after the differentiation process that follows ovulation. Our results show that blockade of the preovulatory rise of ovarian ornithine decarboxylase (ODC), a key enzyme in polyamine biosynthesis, by treatment with the specific inhibitor alpha-difluoromethylornithine (DFMO) leads to a significant decrease in the ovarian progesterone content and a dramatic fall in the plasma levels of this hormone during the following diestrus. The same inhibition was produced in spite of the fact that both luteinizing and follicle stimulating hormones were given concomitantly with DFMO. On the other hand, the acute rise in the plasma progesterone levels observed after administration of human chorionic gonadotropin to mice at different periods of the estrous cycle was not affected by DFMO administration. Our results indicate that although elevated levels of ODC are not required for acute ovarian steroidogenesis, the preovulatory peak of ovarian ODC activity observed in the evening of proestrus may be critical for the establishment of a constitutive steroidogenic pathway and progesterone secretion by the corpus luteum during the diestrus stage of the murine estrous cycle.     

Progesterone profiles around the time of insemination do not show clear differences between of pregnant and not pregnant dairy cows

In this study, features of progesterone profiles were examined in relation to the outcome of insemination. Three groups of estrous cycles were analyzed: resulting in pregnancy, not resulting in pregnancy and resulting in lost pregnancy. The aim of the study was to identify a complex of progesterone profile features associated with successful insemination. The features used were (1) from the estrous cycle preceding the artificial insemination: estrus progesterone concentration, post-estrus maximum rate of increase in progesterone, luteal phase peak, pre-estrus maximum rate of decline in progesterone and the length of follicular and luteal phase and (2) from the estrous cycle following insemination: estrus progesterone concentration, post-estrus maximum rate of increase in progesterone and days from estrus to post-estrus maximum rate of increase in progesterone. A discriminant analysis did not reveal clear differences between the groups. However, the analysis correctly classified 75% of true pregnant cows. Conversely, only 60% of not pregnant animals were classified as such by the discriminate analysis. Individual analysis of progesterone profile features in pregnant and not pregnant groups of estrous cycles showed that a shorter follicular phase preceding insemination is associated with proper timing of post-ovulatory luteinisation and therefore is more likely to result in pregnancy.     

Changes in mitochondrial and microsomal 3 beta-hydroxysteroid dehydrogenase activity in mouse ovary over the course of the estrous cycle.

3 beta-Hydroxysteroid dehydrogenase (HSD) is located in the endoplasmic reticulum and mitochondria. To determine whether the separate enzymes play different roles in steroidogenesis, the specific activity (SA) of both were measured at four different stages of the mouse estrous cycle. Microsomal HSD activity changed little throughout, averaging 8.7 +/- 0.7 nmol progesterone/min/mg protein. In contrast, mitochondrial HSD activity changed dramatically at diestrus, increasing to 14.4 nmol progesterone/min/mg protein. When measured at proestrus, estrus, and metestrus, mitochondrial HSD activity was 5.5, 7.4, and 4.5 nmol progesterone/min/mg protein, respectively. To ascertain whether the increase in mitochondrial HSD activity at diestrus could be due to a preferential induction of enzyme, its SA and the SA of a mitochondrial inner membrane enzyme, cytochrome C oxidase, were compared to the SA of a mitochondrial outer membrane enzyme, rotenone-insensitive NADH cytochrome C reductase. The SA of all three enzymes changed proportionally at diestrus, suggesting that the increase in mitochondrial HSD activity was not due to its preferential induction. Rather, we believe that the HSD activity in the mitochondrial fraction, as measured at the four stages of the estrous cycle, is a reflection of the combined contributions from an ever changing population of ovarian cells. Mitochondria from luteal cells have the highest HSD activity, and are very likely responsible for the major synthesis of progesterone during the luteal phase.     

Expression and localization of progesterone receptor membrane component 1 and 2 and serpine mRNA binding protein 1 in the bovine corpus luteum during the estrous cycle and the first trimester of pregnancy

The aim of this study was to evaluate the mRNA and protein expression and the localization of progesterone receptor membrane component 1 (PGRMC1), PGRMC2, and the PGRMC1 partner serpine mRNA binding protein 1 (SERBP1) in the bovine CL on Days 2 to 5, 6 to 10, 11 to 16, and 17 to 20 of the estrous cycle as well as during Weeks 3 to 5, 6 to 8, and 9 to 12 of pregnancy (n = 5–6 per each period). The highest levels of PGRMC1 and PGRMC2 mRNA expression were found on Days 6 to 16 (P < 0.05) and 11 to 16, respectively, of the estrous cycle and during pregnancy (P < 0.001). The level of PGRMC1 protein was the highest (P < 0.05) on Days 11 to 16 of the estrous cycle compared with the other stages of the estrous cycle and pregnancy, whereas PGRMC2 protein expression (P < 0.001) was the highest on Days 17 to 20 and also during pregnancy. The mRNA expression of SERBP1 was increased (P < 0.05) on Days 11 to 16, whereas the level of its protein product was decreased (P < 0.05) on Days 6 to 10 of the estrous cycle and was at its lowest (P < 0.001) on Days 17 to 20. In pregnant cows, the patterns of SERBP1 mRNA and protein expression remained constant and were comparable with those observed during the estrous cycle. Progesterone receptor membrane component 1 and PGRMC2 localized to both large and small luteal cells, whereas SERBP1 was observed mainly in small luteal cells and much less frequently in large luteal cells. All proteins were also localized in the endothelial cells of blood vessels. The data obtained indicate the variable expression of PGRMC1, PGRMC2, and SERBP1 mRNA and protein in the bovine CL and suggest that progesterone may regulate CL function via its membrane receptors during both the estrous cycle and pregnancy.     

Gap junctional intercellular communication of bovine luteal cells from several stages of the estrous cycle: Effects of prostaglandin F2α, protein kinase C and calcium

Cellular interactions mediated by both contact-dependent and contact-independent mechanisms are probably important to maintain luteal function. The present studies were performed to evaluate the effects of luteotropic and luteolytic hormones, and also intracellular regulators, on contact-dependent gap junctional intercellular communication (GJIC) of bovine luteal cells from several stages of luteal development. Bovine corpora lutea (CL) from the early, mid and late luteal phases of the estrous cycle were dispersed with collagenase and incubated with no treatment, LH, PGF or LH + PGF (Experiment 1), or with no treatment, or agonists or antagonists of protein kinase C (TPA or H-7) or calcium (A23187 or EGTA; Experiment 2). After incubation, media were collected for determination of progesterone concentrations. Then the rate of GJIC was evaluated for small luteal cells in contact with small luteal cells, and large luteal cells in contact with small luteal cells by using the fluorescence recovery after photobleaching technique and laser cytometry. Luteal cells from each stage of the estrous cycle exhibited GJIC, but the rate of GJIC was least (P<0.05) for luteal cells from the late luteal phase. LH increased (P<0.05) GJIC between small luteal cells from the mid and late but not the early luteal phase. PGF increased (P<0.05) GjIC between small luteal cells from the mid luteal phase and diminished (P<0.05) LH-stimulatory effects on GjIC between small luteal cells from the late luteal phase. Throughout the estrous cycle, TPA decreased (P<0.05) the rate of GjIC between large and small, and between small luteal cells, and A23187 decreased (P<0.05) the rate of GJIC between large and small luteal cells. LH and LH + PGF, but not PGF alone increased (P<0.05) progesterone secretion by luteal cells from the mid and late luteal phases. Agonists or antagonists of PKC or calcium did not affect progesterone secretion by luteal cells. These data demonstrate that both luteal cell types communicate with small luteal cells, and the rate of communication depends on the stage of luteal development. LH and PGF affect GjIC between small luteal cells during the fully differentiated (mid-luteal) and regressing (late luteal) stages of the estrous cycle. In contrast, at all stages of luteal development, activation of PKC decreases GjIC between small and between large and small luteal cells, whereas calcium ionophore decreases GjIC only between large and small luteal cells. Luteotropic and luteolytic hormones, and intracellular regulators, may be involved in regulation of cellular interactions within bovine CL which likely is an important mechanism for coordination of luteal function.     

Progesterone is a cell death suppressor that downregulates Fas expression in rat corpus luteum

In female rats, apoptotic cell death in the corpus luteum is induced by the prolactin (PRL) surge occurring in the proestrous afternoon during the estrous cycle. We have previously shown that this luteolytic action of PRL is mediated by the Fas/Fas ligand (FasL) system. During pregnancy or pseudopregnancy, apoptosis does not occur in the corpus luteum. Progesterone (P4), a steroid hormone secreted from luteal steroidogenic cells, attenuated PRL-induced apoptosis in cultured luteal cells in a dose-dependent manner. P4 significantly decreased the expression of mRNA of Fas, but not FasL, in cultured luteal cells prepared from both proestrous and mid-pseudopregnant rats. These data indicate that P4 suppresses PRL-induced luteal cell apoptosis via reduction of the expression level of Fas mRNA in the corpus luteum, suggesting that P4 acts as an important factor that can change the sensitivity of corpus luteum to PRL.     

Effects of second messengers on gap junctional intercellular communication of ovine luteal cells throughout the estrous cycle

Corpora lutea (CL) from Days 5, 10, and 15 after superovulation were enzymatically dispersed, and a portion of the cells were elutriated to obtain fractions enriched with small or large luteal cells. Mixed, small, and large luteal cell fractions were incubated with no treatment or with agonists or antagonists of cAMP (dbcAMP or Rp-cAMPS), protein kinase C (PKC; TPA or H-7), or calcium (A23187, EGTA, or A23187 + EGTA). The rate of contact-dependent gap junctional intercellular communication (GJIC) was evaluated by laser cytometry. Media were collected for progesterone (P(4)) radioimmunoassay, and luteal cells cultured with no treatment were fixed for immunocytochemistry or frozen for Western blot analysis. Luteal cells from each stage of the estrous cycle exhibited GJIC. The dbcAMP increased (P < 0.05) GJIC for all cell types across the estrous cycle. The Rp-cAMPS decreased (P < 0.05) GJIC for small luteal cells on Day 5 and for all cell types on Days 10 and 15. The TPA inhibited (P < 0.01), but H-7 did not affect, GJIC for all cell types across the estrous cycle. The A23187 decreased (P < 0.05) GJIC for large luteal cells touching only small or only large luteal cells, whereas A23187 + EGTA decreased (P < 0.05) GJIC for all cell types across the estrous cycle. For the mixed and large luteal cell fractions, dbcAMP increased (P < 0.05), but TPA and A23187 + EGTA decreased (P < 0.05), P(4) secretion. The A23187 alone decreased (P < 0.05) P(4) secretion by large, but not by mixed, luteal cells. For all days and cell types, the rate of GJIC and P(4) secretion were correlated (r = 0.113-0.249; P < 0.01). Connexin 43 was detected in cultured luteal cells by immunofluorescence and Western immunoblotting. Thus, intracellular regulators like cAMP, PKC, or calcium appear to regulate GJIC, which probably is an important mechanism for coordinating function of the ovine CL.     

Morphometric quantification of mitochondria in the two steroidogenic ovine luteal cell types

Progesterone secretion is regulated by different mechanisms in large and small steroidogenic ovine luteal cells. Large cells secrete approximately 7-fold more progesterone in an unstimulated state than small cells. Since cholesterol side-chain cleavage, which is catalyzed by an inner mitochondrial membrane enzyme complex, is a major rate-limiting step in progesterone synthesis, mitochondrial components were quantified in the two steroidogenic cell types throughout the estrous cycle. Corpora lutea collected on Days 4 (n = 4), 8 (n = 4), 12 (n = 5), and 16 (n = 6) of the estrous cycle were prepared for electron microscopy. Volume densities of cell types within corpora lutea and mitochondrial densities within cell types were estimated by point-counting; nuclear and cytoplasmic volume densities were estimated by planimetric analysis. A total of 570 micrographs (magnification 5300 X) were analyzed. Large cell volume density was unchanged during the cycle (35 +/- 1%) while small cell volume density increased (p less than 0.05) from 13 +/- 1% on Day 4 to 20 +/- 3% on Day 12. Large cell mitochondrial volume density increased (p less than 0.05) from 13 +/- 1% on Day 4 to 23 +/- 1% on Day 16 accompanied by an increase in cytoplasmic volume density such that nuclear to cytoplasmic ratio increased (p less than 0.05) from 1:14 to 1:34 between Days 4 and 16. Small cell mitochondrial volume density increased from 11 +/- 1% on Day 4 to 14 +/- 1% (p less than 0.05) for the rest of the cycle while the nuclear to cytoplasmic ratio remained at 1:14.(ABSTRACT TRUNCATED AT 250 WORDS)     

Regression of the corpus luteum of pregnancy following parturition in the ewe

Corpora lutea (CL) of pregnancy from single-lambing ewes were examined by light and electron microscopy within 24 h and at 8, 15, 23, 31 and 41 days after parturition (2 ewes per stage). Within 24 h of parturition the structure of the CL was well preserved and both large and small luteal cells, characteristic of this species, were present in substantial numbers. However, both types of luteal cell contained numerous cytoplasmic lipid droplets, and smooth endoplasmic reticulum and secretory granules in large luteal cells were less prominent than in normal functional CL of cyclic ewes. Leucocytic infiltration, and death of some luteal and endothelial cells, were also observed at this stage. Further regression of the CL progressed slowly, and lipid-rich large luteal cells were still readily recognisable 15 days after parturition. The size of the CL declined progressively, and the proportion of tissue occupied by intercellular substances increased. Corpora albicantia approximately equal to 3-4 mm in diameter were still recognisable 41 days after parturition. It was concluded that luteal regression post partum progresses much less rapidly than at the end of the oestrous cycle.     

Investigations on spermicidal activity in the sheep vagina.

This study was conducted to elucidate some of the effects of a synthetic progestagen and natural ovarian hormones on spermicidal activity in the sheep vagina. In the first experiment, parous ewes were treated for 17 days either intravaginally with medroxyprogesterone acetate (MAP) or subcutaneously with progesterone. They were inseminated artificially either on the last day of progestagen treatment or during estrus after progestagen withdrawal. Their vulvovaginal junctions were ligated to prevent the loss of sperm cells by drainage to the exterior. Untreated control ewes were inseminated during either estrus or the luteal phase of the estrous cycle. The ewes were killed 22 hr. after insemination, their vaginas flushed, and intact sperm cells and tailless sperm heads counted. In the second and third experiments, some of the ewes were bilaterally ovariectomized and inseminated several weeks later. Other ewes were ovariectomized and given subcutaneous injections of estradiol, progesterone, or both hormones.In the first experiment, most sperm cells were recovered intact from estrous or luteal phase control ewes. The intravaginal administration of MAP increased both the breakage of sperm cells into heads and tails and the disappearance of sperm cells. The spermicidal effects of MAP were just as great in ewes inseminated on the last day of treatment. as in those inseminated during the ensuing estrus; these results indicated that the peak estrogen secretion that occurs near the beginning of estrus was not necessary for the intensification of spermicidal activity.In the second experiment, ovariectomized ewes were compared to estrous and luteal phase ewes in regard to vaginal spermicidal activity. Sperm breakage and disappearance occurred least in estrous ewes, to a somewhat greater degree in luteal phase ewes, and to the greatest extent in ovariectomized ewes. The results suggested that endogenous ovarian hormones, particularly those in estrous ewes, suppress spermicidal mechanisms in the vagina.In the third experiment, the administration of estradiol and progesterone to ovariectomized ewes prevented the increase in sperm cell disappearance. Neither hormone alone prevented the increase.