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In mammals, hormonal regulation during gestation is crucial for embryo implantation and pregnancy success. This regulation is controlled through the level of progesterone (P4) that blocks the activity of the hypothalamic-hypophyseal-gonadal (HHG) axis. Previous studies in the pregnant South American plains vizcacha, Lagostomus maximus, have shown that the HHG axis activates around mid-gestation, promoting pre-ovulatory follicle formation. However, the characterization of the hormonal dynamics throughout gestation and its ovarian correlation has not been studied in depth. We studied the ovarian dynamics of L. maximus and its correlation with the hormonal profile during gestation, analyzing serum levels of P4, 17β-estradiol (E2), 4Δ-androstenedione (A4), luteinizing hormone (LH) and follicle stimulating hormone (FSH) as well as the ovarian distribution and expression of their receptors. Additionally, we have analyzed the folliculogenesis and accessory corpora lutea (ACL) formation. P4 showed two concentration peaks reaching its highest level at mid-gestation decreasing at 91-100days post-coitum. P4 decrease is followed by an increase of circulating levels of A4, E2, FSH and LH and with an elevated number of antral/pre-ovulatory follicles which express PGR, ESR1, ESR2, AR, LHR and FSHR. In addition, ACL with oocyte retention and cytoplasmic lipid droplets in luteal cells were detected at this time point. These results show that in L. maximus the decrease of P4 level from mid-gestation enables follicular recruitment until pre-ovulatory stage and the development of functional ACL., (Copyright © 2017 Elsevier Inc. All rights reserved.)

The major characteristic of cell death by apoptosis is the loss of nuclear DNA integrity by endonucleases, resulting in the formation of small DNA fragments. The application of confocal imaging to in vivo monitoring of dynamic cellular events, like apoptosis, within internal organs and tissues has been limited by the accessibility to these sites. Therefore, the aim of the present study was to test the feasibility of fibered confocal fluorescence microscopy (FCFM) to image in situ apoptotic DNA fragmentation in surgically exteriorized sheep corpus luteum in the living animal. Following intra-luteal administration of a fluorescent DNA-staining dye, YO-PRO-1, DNA cleavage within nuclei of apoptotic cells was serially imaged at the single-cell level by FCFM. This imaging technology is sufficiently simple and rapid to allow time series in situ detection and visualization of cells undergoing apoptosis in the intact animal. Combined with endoscope, this approach can be used for minimally invasive detection of fluorescent signals and visualization of cellular events within internal organs and tissues and thereby provides the opportunity to study biological processes in the natural physiological environment of the cell in living animals.

Structural luteolysis shows striking interspecies differences. Morphological changes in the corpus luteum (CL) of the cyclic hamster have been studied alongside the potential involvement of known luteolytic hormones. Ovaries from intact Syrian golden hamsters killed at 1100 h on days 1 and 2 and at 1100 and 1700 h on days 3 and 4 of the estrous cycle were dissected for histological study. The day of ovulation, the day of estrus, was arbitrarily designated day 1 of the estrous cycle. Steroidogenic cells in the CL were scarcely luteinized on day 1 and reached full luteinization on day 2. On the morning of day 3, initial regressive changes (accumulation of lipid droplets, invasion by neutrophils, and accumulation of phagocytic cells) were observed. These regressive changes increased progressively and apoptotic cells as well as phagocytic cells containing phagocytized apoptotic cells were abundant on the evening of day 3. On the morning of day 4, apoptotic cells/bodies and phagocytic cells containing phagocytized material were extremely abundant throughout the CL. However, steroidogenic cells with intact nuclei and well-preserved blood vessels were also found. Surviving cells in the CL showed progressive morphological changes. These cells showed morphological features intermediate between luteal and interstitial cells in the evening of day 4 and were virtually indistinguishable from interstitial cells on day 1 of the following cycle. Additional animals were injected at 1100 h on day 2 with: (a) the dopaminergic agonist CB154 (0.4 mg) to block prolactin secretion, (b) the anti-estrogen LY117018 (1.6 mg) or the anti-androgen Flutamide (3 mg) to block estrogen or androgen receptors, respectively, and (c) progesterone (2 mg) to prevent the fall in serum progesterone concentrations. Ovaries from these animals were collected at 1700 h on day 3 and at 1000 h on day 4. The luteolytic process was not affected by any treatment. These data indicate that, in contrast to its close relatives (e.g., the rat), structural luteolysis in the hamster is independent of the apoptotic inducing luteolytic hormones. In addition, differences in the cellular mechanisms responsible for CL elimination were also present. In the hamster, part of the luteal cells do not undergo apoptosis and seemed to progress through another developmental path giving rise to interstitial-like cells., (Copyright 2001 Academic Press.)

The relationship between plasma levels of arginine vasotocin (AVT), progesterone, and corpus luteum formation and degeneration was studied in the viviparous lizard Tiliqua rugosa. Hormone levels were monitored in free-ranging, pregnant females which were located for sampling by means of attached radio transmitters. There was an increase in plasma AVT levels in the 30 days immediately prior to parturition. Concurrent with this event was a decline in plasma progesterone levels from relatively high levels in mid-term to basal levels prior to parturition. This is associated with degenerative changes in the corpus luteum which include pyknosis of the nuclei of the cells of the cell mass and increasing prevalence of intercellular spaces, while the thecal layer became increasingly compacted. Ovariectomy experiments indicate that the major source of progesterone during pregnancy in T. rugosa is ovarian.

The effect of exogenous prostaglandin F2 alpha (PGF) on luteal function and morphology was examined in the lizard, Anolis carolinensis. Reproductively active females were injected with 1.0 micrograms PGF, 0.1 micrograms PGF, or 0.05 ml saline. Animals were killed 2, 6, or 24 hr after injection. Prostaglandin treatment induced luteolysis as evidenced by significant decreases in plasma progesterone concentration in animals sacrificed 6 and 24 hr after treatment. Both doses of PGF were effective. However, 0.1 micrograms PGF induced luteolysis in less than 50% of the treatment group. Luteal degradation became apparent at the cellular level 24 hr after PGF treatment whereas no significant changes were observed in corpora lutea of the 2- or 6-hr postinjection specimens. These data suggest that PGF is a luteolytic agent in reptiles.

Changes in the gonadotropin-binding sites in plasma membranes and several intracellular organelles of bovine corpora lutea of days 3, 13 and 19 of the cycle were investigated. These three times represent periods of rapid luteal growth (early luteal phase), maturity (mid luteal phase) and the onset of regression (late luteal phase), respectively. The 5'-nucleotidase activity was highest in the fraction possessing a predominance of plasma membranes. It was undetectable in nuclear fractions and detectable to a varying extent in fractions enriched with mitochondria-lysosomes, rough endoplasmic reticulum and Golgi. The gonadotropin-binding sites, as measured by 125I-human choriogonadotropin (hCG) specific binding, were found in all the subcellular organelles. Whereas the affinities remained about the same, the total number of available gonadotropin-binding sites in all the organelles increased from day 3 to 13 and then declined by day 19 of the cycle. Occupancy of binding sites by endogenous luteinizing hormone was not detectable and therefore was unlikely to be responsible for the changes in total number of available binding sites. Thus, binding site changes observed in all the organelles of early, mid and late luteal phase corpora lutea probably reflect actual changes in the steady-state turnover of binding sites. Morphometrically determined relative membrane counts of various subcellular organelles varied with the luteal phase. The relative total gonadotropin-binding sites, calculated from the relative membrane counts and the total number of available binding sites, increased in all the organelles from early to mid and then declined by late luteal phase. Plasma membranes of all three luteal phases contained greater relative total gonadotropin-binding sites than any other single intracellular organelle. However, all the intracellular organelles combined contained 59% of the total luteal cell gonadotropin-binding sites in early luteal phase which decreased to 43 and 28% by mid and late luteal phases respectively.

Luteal cells of immature female rats treated with gonadotropins contain microtubules with a number of interesting features. Many of the microtubules of these cells are arranged in bundles in which they are separated one from another by strands of material (i-MT bands) of unknown composition. The microtubules within the bundles assume a hexagonal packing pattern with i-MT bands between any two microtubules. The bundle microtubules and their i-MT bands are further connected via crosslinking filaments: pattern obtained from densitometer scans (measuring the arrangement of the crosslinking filaments) suggest that the filaments may represent microtubule-associated proteins. The complex arrangement of the microtubules within a bundle does not appear to extend for the entire length of the individual microtubules, and occasionally one sees profiles of single microtubules fanning out from the ends of the bundle: whether the same microtubules are regrouped at some other point in the cell is not known. Structures similar to the i-MT band and the crosslinking filaments have also been observed connecting microtubules to segments of the luteal cell plasma membrane: in these instances the i-MT-like band is found between the longitudinally sectioned microtubule and the membrane, with filaments connecting the two structures via the intermediate band. It is of interest that the microtubules of these luteal cells are not sensitive to treatment with antimicrotubule drugs and we suggest that the complex bundling arrangement provides their unusual stability.

The administration of arginine vasotocin (AVT) to gravid snapping turtles with steroidogenically active corpora lutea and high plasma progesterone concentration (1480 +/- 155 pg/ml) did not trigger oviposition, whereas 12 days after ovulation when luteolysis occurred and plasma progesterone concentration was low (570 +/- 78 pg/ml), treatment with AVT caused oviposition. Controls with high plasma progesterone concentration (1605 +/- 185 pg/ml) oviposited 15-23 days after ovulation when plasma progesterone concentration dropped to 201 +/- 35 pg/ml. Deluteinization-induced oviposition was initiated 15 hr after surgery and was completed by 30 hr. Oviposition of complete clutches occurred and was correlated with a significant drop in plasma progesterone. Sham-operated turtles did not exhibit oviposition and no significant change in progesterone concentration was observed. A single injection of prostaglandin F2 alpha (PGF) in recently ovulated turtles induced early luteolysis and a significant decrease in plasma progesterone concentration after 24-30 hr. A single administration of PGF caused the disappearance of steroidogenic features such as the smooth endoplasmic reticulum and mitochondria with tubular cristae 48 hr later. Also PGF triggered the invasion of a hyaline-like material from the luteal theca into the luteal cell mass which eventually induced luteolysis. The role of AVT, PGF, and progesterone in relation to egg retention and oviposition is discussed.