Rodney A. Mead

Effects of progesterone and estrogen on the histology of the oviduct of the garter snake, Thamnophis elegans

Abstract Silastic capsules packed with estradiol and/or progesterone were implanted in ovariectomized viviparous garter snakes ( Thamnophis elegans ) in order to define the role of these steroids in inducing histological recrudescence in the uterine segment of the oviduct. During the preovulatory period, the uterine tubuloalveolar glands and tall columnar, luminal epithelium of intact snakes were exceedingly well-developed and possessed numerous PAS-positive granules. Plasma estrogen concentrations were quite high (1833 ± 167 pg/ml) at this time and progesterone levels were variable (3.3 ± 2.3 ng/ml). Ovariectomy significantly reduced plasma levels of estrogen (79 ± 48 pg) and progesterone (1.1 ± 0.3 ng/ml) as well as the height and secretory condition of the luminal and glandular epithelium of the uterus. Administration of estradiol to ovariectomized garter snakes for 10 days elevated plasma estrogen levels to 758 ± 351 pg/ml and partially restored epithelial cell height and glandular activity. Administration of progesterone elevated plasma levels to physiological concentrations (4.3 ± 2.6 ng/ml) but had little or no stimulatory effect on uterine histology. Simultaneous administration of estradiol and progesterone for 10 days failed to induce additional stimulatory effects on uterine histology over and above those produced by estradiol alone. Results of this study indicate that estradiol is the most important ovarian hormone in regulating development of the uterus. However, additional hormones, other than progesterone, may possibly be required for full restoration of the garter snake uterus to the preovulatory condition.

Sources and levels of progesterone during pregnancy in the garter snake, Thamnophis elegans

Abstract Although it is commonly assumed that corpora lutea (CL) in live-bearing reptiles function as endocrine glands during pregnancy, it remains to be shown that CL of snakes or other live-bearing reptiles contain and secrete progesterone at this time. To test the latter possibilities, progesterone concentrations in CL of 66 viviparous garter snakes ( Thamnophis elegans ) were measured throughout pregnancy using gas phase chromatography and electron capture detection. In addition, 101 viviparous garter snakes ( Thamnophis elegans ) were bled throughout pregnancy or before and after luteectomy, sham luteectomy, ovariectomy, placentectomy, or adrenalectomy; and plasma progesterone levels were quantitated by radioimmunoassay. Concentrations of progesterone in the CL and plasma were correlated with luteal histology. Average concentrations of 1.1–2.0 ng of progesterone/mg of CL and 1.8–2.8 ng/ml of plasma were recorded during the first trimester of pregnancy, at which time the thecal layers were prominent and the luteal tissue was invaded by blood vessels. Lutein cell nuclei were characteristically round or oval and their cytoplasm vesicular. Progesterone concentrations were highest during the second trimester of pregnancy in the CL (2.6 ng/mg) and plasma (6.2 ng/ml), at which time the thecal layers began compaction. The luteal tissue was conspicuously vascular, lutein cell cytoplasm vesicular, and nuclei of lutein cells were irregular in shape. Progesterone concentrations in the CL fell during the third trimester of pregnancy and were nondetectable 1 wk postpartum. During this time the plasma progesterone levels declined steadily to a mean of 1.0 ng/ml at parturition, and remained low (approx 1.2 ng/ml) for 2 wk postpartum. Nuclei of lutein cells appeared darker and pronounced vacuolation of lutein cell cytoplasm was often observed during this time. Following luteectomy, plasma progesterone levels were approximately equal to those of nonpregnant snakes (1.1 ng/ml) but fell to nondetectable levels following ovariectomy. Progesterone levels were nondetectable after adrenalectomy, but ovariectomy of nonpregnant females produced only a modest decline. Placentectomy or sham surgical treatments did not significantly alter plasma progesterone levels. We conclude that CL are the major source of plasma progesterone during pregnancy, but that nonluteal portions of the ovary also contribute small quantities of progesterone at this time.

Mechanisms of Chromosomal Changes in Mammalian Speciation

Ultimately, biologists should be able to describe each species (or subdivisions thereof) in terms of base sequences of all its genes and the order in which the genes are aligned. Evolution can then be studied by comparing the base sequences and arrangement of genes in various taxa. This naturally is not possible at the present time, so that only cruder criteria, phenotypes such as morphology, anatomy and behavior, have been used to elucidate species relationships, population structure, and evolutionary trends.

Effects of suprachiasmatic nuclear ablation and melatonin on delayed implantation in the spotted skunk.

The suprachiasmatic nuclei (SCN) have been implicated as neural timers of reproductive events and as possible sites of action for melatonin. We tested the hypothesis that ablation of the SCN (SCNx) would counteract the inhibitory effect of exogenous melatonin in blastocyst implantation in the spotted skunk by removing a possible site of action. Thirty-eight pregnant females with unimplanted blastocysts were treated as follows: 4 served as untreated controls, 6 received empty Silastic capsules, 5 received Silastic capsules containing melatonin, 10 received sham lesions in the SCN, 7 received lesions in the SCN and Silastic capsules containing melatonin, and 6 received lesions in the SCN and empty Silastic capsules. All surgical treatments were completed by February 15. The skunks were subjected to a natural photoperiod, and the duration of preimplantation was measured. The lesions destroyed an average of 87 +/- 10% of the SCN and a small amount of the surrounding hypothalamus. SCNx had no significant effect on duration of preimplantation (200 +/- 25.6 days) when compared to sham SCNx (205 +/- 21.5 days). Melatonin significantly (p less than 0.05) lengthened the duration of preimplantation in both intact (277 +/- 59 days) and SCNx (265 +/- 64.7 days) skunks when compared to all other groups. These data are not consistent with the hypothesis that the SCN are required for melatonin to exert its influence on timing of implantation in the spotted skunk.

Progression of trophoblast into the endometrium during implantation in the western spotted skunk.

In mustelid carnivores, several blastocysts become implanted either ∼12 days following fertilization or after a delay of implantation. In the western spotted skunk, implantation occurs following a long period of delayed implantation and a brief activation stage. Within each implantation chamber, a large number of trophoblastic plaques form, and the syncytial trophoblast of these plaques adheres to and penetrates into the uterine luminal epithelium. The presence of multiple attachment sites was used to analyse the way in which trophoblast adheres to, penetrates, and removes uterine epithelium and its subsequent association with the subepithelial vascular plexus.

Function of corpora lutea of pregnancy in the viviparous garter snake, Thamnophis elegans.

Although corpora lutea of the viviparous garter snake, Thamnophis elegans, are the major source of plasma progesterone during pregnancy, a specific function for this steroid or any other luteal factor has yet to be shown. Thirty pregnant snakes weighing 80–200 g were divided into three equal groups so that all groups contained snakes of comparable size. Equal numbers of snakes were luteectomized, sham luteectomized, or received no treatment during early pregnancy and were then allowed to complete gestation. Duration of pregnancy was significantly (P < 0.01) longer in luteectomized snakes than in either control group. Luteectomized snakes gave birth to a lower percentage of living young and fewer birth products, which also weighed significantly less. However, luteectomy did not prevent normal embryonic development and did not appear to affect the rate of embryonic development. There were no significant differences between intact and sham luteectomized snakes in any of the above parameters. We conclude that CL in the viviparous garter snake, Thamnophis elegans, function through the mediation of a hormone which ensures that parturition occurs at the normal time and with the delivery of a normal number of young. The mechanism or the compound responsible for this action remains unknown.

Changes in Uterine Expression of Leukemia Inhibitory Factor Receptor Gene During Pregnancy and Its Up-Regulation by Prolactin in the Western Spotted Skunk

Abstract The multifunctional cytokine leukemia inhibitory factor (LIF) is presumed to participate in preparing the uterus for blastocyst implantation. Increased production of LIF is positively correlated with termination of embryonic diapause and preparation for implantation in the spotted skunk. This study examined changes in the expression, localization, and hormonal regulation of LIF receptor (LIFRβ) gene expression in the uterus of the skunk. Changes in the uterine concentration of LIFRβ mRNA during pregnancy or in response to hormones after ovariectomy were determined by Northern hybridization analysis and reverse-transcriptase polymerase chain reaction. The skunk uterus produces two LIFRβ transcripts, the levels of which increase in concentration when the blastocysts resume their development but then decline somewhat during the latter stage of blastocyst activation. Ovariectomy significantly reduced uterine LIFRβ expression. Estradiol and/or progesterone failed to significantly elevate LIFRβ mRNA levels in ovariectomized animals. Prolactin significantly increased uterine concentrations of LIFRβ mRNA to greater than those of ovariectomized controls, but these levels were not comparable to those observed during preimplantation. The LIFRβ mRNA was predominately localized to stromal cells surrounding the uterine glands and in yolk sac endoderm, syncytiotrophoblast, and cytotrophoblast of postimplantation embryos.

Autoradiographic localization of binding sites for 2-[125I]iodomelatonin in the pars tuberalis of the western spotted skunk (Spilogale putorius latifrons)

Pineal melatonin secretion mediates photoperiodic regulation of implantation of blastocysts in the female western spotted skunk. Autoradiography studies demonstrated that high affinity 2-[125I]iodomelatonin binding sites are present in the pars tuberalis but not in any other hypophyseal region or in the hypothalamus or thalamus of this species. This restricted localization of 2-[125I]iodomelatonin binding, which was characteristic of spotted skunks sacrificed at various times during their annual reproductive cycles, suggests that the pars tuberalis mediates photoperiodic responses in this species.

Hormonal and gestational evidence for delayed implantation in the striped skunk, Mephitis mephitis

Two lines of evidence are presented for a brief, obligate delay in implantation in the striped skunk. First, a relationship was found between the date of mating and gestation and is described by a weighted least-squares model Y = 59.85 + 204.94 e−0.0519X. Matings occurred between 21 February and 5 April with gestations ranging from 59 to 77 days. Longer gestations usually were associated with matings occurring early in the breeding season. The second line of evidence involves differences in progesterone levels in females mated early (2 March) and late (22 March) in the breeding season. Females mated early in the breeding season had progesterone levels that required longer for their initial increase (4–7 days) and did not peak (32.3 ng/ml) until 27 days postcoitus. Females bred later in the breeding season had initial increases in progesterone after 2–4 days postcoitus and a peak in progesterone (31.1 ng/ml) at 19 days postcoitus. Progesterone levels in mated females that failed to give birth were significantly lower on Days 47 and 56 than hormone levels for pregnant skunks. Unmated controls that were samples throughout the year had negligible amounts of progesterone (<1 ng/ml) present. Therefore, striped skunks mated early in the breeding season had longer gestations, lower initial progesterone levels, and later peaks in progesterone secretion, suggesting that a brief, obligate delay in implantation occurred.

Effects of Hypophysectomy on Blastocyst Survival, Progesterone Secretion and Nidation in the Spotted Skunk

12 female skunks were hypophysectomized during the 7-month preimplantation period. Sham operations were performed on 7 skunks by drilling through the basisphenoid but without exposing the pituitary. 22 skunks remained as controls. Complete hypophysectomy significantly reduced peripheral plasma progesterone levels within 40 days while there was a tendency for values to increase in sham hypophysectomized skunks. The only obvious effect hypophysectomy had on ovarian histology during the preimplantation period was the abolition of all antral follicles; corpora lutea remained unchanged for up to 90 days. Blastocysts from hypophysectomized skunks were undergoing a slow attrition of cells after Day 34, and there was some indication that hypophysectomy may have a deleterious effect as early as Day 22. Hypophysectomy significantly prolonged the preimplantation period in 6 out of 7 skunks. It was concluded that the pituitary is essential for initiation of nidation.