Insulin gene expression pattern in rat pancreas during the estrous cycle.

Sex steroid hormones influence insulin homeostasis and glucose metabolism, estradiol (E2) and progesterone (P4) induce changes in both fasting and postprandial insulinemia in rodents, however, insulin gene expression during estrous cycle is unknown. The aim of the present study was to determine an insulin gene expression pattern during the estrous cycle in the rat. Groups of 6 adult rats in each day of the estrous cycle were used. Serum P4, E2, testosterone (T) and insulin concentrations were determined by radioimmunoassay (RIA). A Northern blot analysis was performed to assess insulin gene expression in pancreatic tissue. We found a marked variation in insulin gene expression during the estrous cycle. The highest insulin expression was observed during the proestrus day. Interestingly, E2 and P4 but not T levels were correlated with changes in insulin mRNA content. The variations in serum insulin during the cycle were correlated with its mRNA content in pancreas. The overall results showed variations in serum insulin and insulin gene expression during estrous cycle of the rat that correlated with circulating E2 and P4 levels.     

Changes in the content and distribution of microtubule associated protein 2 in the hippocampus of the rat during the estrous cycle

The molecular mechanisms involved in the regulation of synaptic plasticity in the hippocampus during the estrous cycle of the rat are not completely understood. Because this process implicates changes in neuronal cytoskeleton organization, we analyzed the content of microtubule associated protein 2 (MAP2) and Tau in the hippocampus and the frontal cortex of the rat by Western blot, as well as the hippocampal distribution of MAP2 during the estrous cycle by immunohistochemistry. In the hippocampus the lowest content of MAP2 was found on diestrus day, and it significantly increased at proestrus. This increase was maintained on estrus and metestrus days. In the frontal cortex MAP2 content did not significantly change during the estrous cycle. In contrast, the content of Tau did not vary during the estrous cycle in either the hippocampus or the frontal cortex. The immunohistochemical analysis showed an increase in dendrite thickness and in dendritic branching in the CA1 region on proestrus day, as well as an aggregation of MAP2 in apical dendrites near to pyramidal somata on this day in comparison with diestrus. We suggest that changes in the content and neuronal distribution of MAP2 are involved in the structural changes that occur in the hippocampus of the rat during the estrous cycle, and that these variations are related to changes in estradiol and progesterone levels.     

Variations in free secretory component levels in mucosal secretions of the rat

The present studies were conducted to compare the levels of free secretory component (SC) in a number of rat mucosal secretions and to determine whether SC content varies significantly during the four stages of the estrous cycle. Levels of SC, as measured by radioimmunoassay, were markedly different in various external secretions. Bile contained the highest amount, irrespective of whether SC was normalized to volume or protein. Concentrations of SC in saliva or uterine fluid from intact rats were approximately 20- to 30-fold less than measured in bile. When SC levels were normalized to protein, the SC to protein ratios in uterine, vaginal, and respiratory secretions were six to 18 times greater than values calculated in salivary and small intestinal fluids. Analysis of SC levels in mucosal secretions during the estrous cycle indicated significant variations occur in uterine and vaginal samples, but not in saliva or small intestinal secretions. In the uterine lumen, SC levels were highest at proestrus, partially elevated at estrus, and lowest at both days of diestrus. In contrast, vaginal SC levels were maximal at estrus and reduced at all other stages of the cycle. Immunoglobulin A content was also measured in uterine and vaginal secretions during the estrous cycle. Significant changes in IgA levels were found and these coincided with the changing pattern of SC. These results suggest hormones may modulate SC levels in reproductive tissues. In addition, our findings indicate variations in SC during the estrous cycle may direct the movement of IgA from tissue to lumen.     

Changes in tissue histamine during the estrous cycle, pregnancy and pseudopregnancy in the golden hamster

The histamine content of reproductive tissues and skeletal muscle was determined in the golden hamster during the estrous cycle, pregnancy, and pseudopregnancy. Histidine decarboxylase activity was measured in uterine implantation sites and intersites from Day 4 to Day 10 of pregnancy. Histidine decarboxylase was also measured in mesometria and placentas on selected days of gestation. During the estrous cycle, uterine and skeletal muscle histamine levels were highest on Day 2 and lowest on Day 4 of the cycle. The ovarian histamine content did not change significantly among the different stages of the cycle. While the histamine content of uterine implantation sites of attachment was high on Days 4 and measurable on Days 5 and 6 of pregnancy, the levels were below the limits of detection by Day 7. On the other hand, the highest levels of histamine were in the uterine interimplantation sites on Days 8 and 9. The ovarian levels of histamine were highest on Day 13 of pregnancy. Histamine in skeletal muscle did not change significantly during pregnancy. The histidine decarboxylase activity in the implantation sites began rising on Day 9 and increased dramatically on Day 10. Placental histidine decarboxylase activity was very high on Days 13 and 15. Overall, we observed changes in uterine and skeletal muscle histamine during the estrous cycle that may be explainable in light of previously reported changes in mast cell numbers and circulating estrogens. During pregnancy, histamine levels of implantation sites and implantation intersites varied, as did the histamine content of ovarian tissue. Histidine decarboxylase activity rises in the uterus and placental tissue after the formation of the placenta.     

Autophagic and proteolytic processes in the Harderian gland are modulated during the estrous cycle

The Syrian hamster Harderian gland (HG) is an organ that undergoes physiological autophagy in response to oxidative stress induced by porphyrin production. Porphyrin production in the HG has marked sex differences and is closely linked to reproductive function. In the present study, we observed that the estrous cycle and associated estrogen variations may affect oxidative-stress-induced proteolytic processes. In particular, significant changes in autophagic activity were detected during the estrous cycle. Notably, increased activation of macroautophagy as well as chaperone-mediated autophagy in the estrus phase coincided with a minimal antioxidant capability and the highest protein damage levels. By contrast, autophagic machinery was found to be blocked in the diestrus phase, likely due to mammalian target of rapamycin activation, which could be corroborated by the subsequent pS6K activation. Analogous results were observed regarding proteasome activity, which also showed maximal activity in the estrus phase. Interestingly, all these mechanisms were associated with important morphological changes in the HG during the estrous cycle. We observed statistically significant increases in Type II cells, which may be related to extensive autophagy in the estrus phase. Physiologically, this would result in a significant release of porphyrins specifically when females are more receptive. These data support the role of porphyrins as pheromones, as other authors have previously suggested, thus making the HG a scent organ. In addition, these results suggest a porphyrin-based approach to the treatment of porphyria during pregnancy, a condition for which no treatment is currently known.     

3α-Hydroxysteroid dehydrogenase and 3β-hydroxysteroid dehydrogenase in the ovary of young Mongolian gerbils

Summary The ovaries of sexually mature, pregnant mare serum gonadotropin (PMSG) stimulated, 12 week old Mongolian gerbils were investigated morphologically and enzyme histochemically for the appearance of the 3-hydroxysteroid and the 3-hydroxysteroid dehydrogenase during the estrous cycle. Up to ovulation, on day 3 of the estrous cycle, the number of vesicular follicles increases continuously. Primarily atretic follicles can be seen on day 4. On day 5 corpora lutea appear, but they degenerate already by day 6.During the entire estrous cycle, 3-hydroxysteroid dehydrogenase and 3-hydroxysteroid dehydrogenase activity can be found in the theca of tertiary follicles and in the interstitial cells, whereas the theca of secondary follicles and the granulosa of healthy follicles do not exhibit any enzyme activity. The activity decreases from day 1 till day 6. The granulosa of atretic follicles and the cells of corpora lutea show only weak activity. It may be significant that the intensity of enzyme activity in the ovary and the estrogen level in the plasma are differently correlated to the estrous cycle.This investigation was supported by the Deutsche Forschungsgemeinschaft     

A quantitative method for assessing stages of the rat estrous cycle

The impact of gender and/or hormone variations on a wide variety of neural functions makes the choice between studying males or females (or both) of a given species difficult. Although female rats are widely used experimentally, few studies control for the stage of estrus. More detailed information about how to distinguish the various stages of the estrous cycle is needed. For the present study, vaginal smears were obtained once a day and stained using an adaptation of the Papanicolaou (PAP) procedure. Images are provided of unstained “wet” samples and the corresponding PAP stained smears illustrating the cellular profile for each stage of the cycle as well as post-ovariectomy. The different cell populations across the cycle were quantified and ratios determined to show trends between the predominant and other cell types in each stage of the estrous cycle. Both stained and unstained images and cell quantification data provide valuable guidelines for distinguishing the stages of the estrous cycle.     

A quantitative method for assessing stages of the rat estrous cycle

The impact of gender and/or hormone variations on a wide variety of neural functions makes the choice between studying males or females (or both) of a given species difficult. Although female rats are widely used experimentally, few studies control for the stage of estrus. More detailed information about how to distinguish the various stages of the estrous cycle is needed. For the present study, vaginal smears were obtained once a day and stained using an adaptation of the Papanicolaou (PAP) procedure. Images are provided of unstained “wet” samples and the corresponding PAP stained smears illustrating the cellular profile for each stage of the cycle as well as post-ovariectomy. The different cell populations across the cycle were quantified and ratios determined to show trends between the predominant and other cell types in each stage of the estrous cycle. Both stained and unstained images and cell quantification data provide valuable guidelines for distinguishing the stages of the estrous cycle.     

Prostaglandin metabolism in the ovine corpus luteum: catabolism of prostaglandin F(2alpha) (PGF(2alpha)) coincides with resistance of the corpus luteum to PGF(2alpha)

To examine possible mechanisms involved in resistance of the ovine corpus luteum to the luteolytic activity of prostaglandin (PG)F(2alpha), the enzymatic activity of 15-hydroxyprostaglandin dehydrogenase (PGDH) and the quantity of mRNA encoding PGDH and cyclooxygenase (COX-2) were determined in ovine corpora lutea on Days 4 and 13 of the estrous cycle and Day 13 of pregnancy. The corpus luteum is resistant to the action of PGF(2alpha) on Days 4 of the estrous cycle and 13 of pregnancy while on Day 13 of the estrous cycle the corpus luteum is sensitive to the actions PGF(2alpha). Enzymatic activity of PGDH, measured by rate of conversion of PGF(2alpha) to PGFM, was greater in corpora lutea on Day 4 of the estrous cycle (P < 0.05) and Day 13 of pregnancy (P < 0.05) than on Day 13 of the estrous cycle. Levels of mRNA encoding PGDH were also greater in corpora lutea on Day 4 of the estrous cycle (P < 0. 01) and Day 13 of pregnancy (P < 0.01) than on Day 13 of the estrous cycle. Thus, during the early estrous cycle and early pregnancy, the corpus luteum has a greater capacity to catabolize PGF, which may play a role in the resistance of the corpus luteum to the actions of this hormone. Levels of mRNA encoding COX-2 were undetectable in corpora lutea collected on Day 13 of the estrous cycle but were 11 +/- 4 and 44 +/- 28 amol/microgram poly(A)(+) RNA in corpora lutea collected on Day 4 of the estrous cycle and Day 13 of pregnancy, respectively. These data suggest that there is a greater capacity to synthesize PGF(2alpha), early in the estrous cycle and early in pregnancy than on Day 13 of the estrous cycle. In conclusion, enzymatic activity of PGDH may play an important role in the mechanism involved in luteal resistance to the luteolytic effects of PGF(2alpha).     

Release of norepinephrine from the rat ovary: local modulation of gonadotropins.

Experiments were undertaken to define the role of gonadotropins in the release of norepinephrine and the relationship with beta-receptors of the ovary. Rat ovaries were removed at different stages of the estrous cycle and incubated in [3H]norepinephrine. Subsequently, ovaries were electrically stimulated and the release of [3H]norepinephrine was recorded. There were no changes in the norepinephrine content during the estrous cycle. The ovary exhibited cyclical variation in norepinephrine-induced release during the estrous cycle. The lowest release of norepinephrine was found during diestrus; there was an increase during proestrus and estrus followed by a decline during metestrus. The release of norepinephrine changed in the opposite way to the beta-receptor number, suggesting a process involving down-regulation between norepinephrine release and beta-receptors of the ovary. Norepinephrine released from the ovary was locally regulated by gonadotropins. The presence of FSH in the superfusion medium stimulated the norepinephrine-induced release from the ovaries of rats in diestrus (by 20%) and estrus (by 40%), but no effect was found during proestrus. In addition, the presence of hCG stimulated (by 40%) norepinephrine-induced release during proestrus, but no changes were apparent during the other stages of the estrous cycle. These results suggest that the local action of gonadotropins on nerve terminals of the ovary might be one of the factors governing the changes in norepinephrine release through the estrous cycle. The changes in the norepinephrine released to the synaptic cleft might exert down-regulation on the beta-adrenergic receptor content of the ovary and in this way control the ovarian steroid secretory activity.     

Histamine content and mast cells distribution in mouse uterus: the effect of sexual hormones, gestation and labor

The histamine content of uteri from mice was analyzed in terms of both concentration and total amount per uterine horn a) at two stages of the estrous cycle (estrous and diestrous), b) under sex hormone treatment, c) during pregnancy and after delivery. Histamine concentration and mast cell density were greater during diestrous and in mice treated with progesterone (p less than 0.001). This effect was attributed to a reduction in uterine mass weight, since the amount of histamine per uterine horn remained constant throughout the estrous cycle. During pregnancy, both concentration and amount of histamine per uterine horn were increased, values were significantly higher than in estrous (p less than 0.001) from day 14-17 until day 21 when labor occurred. After six to eight hours post-partum an abrupt reduction on histamine content was observed. Mast cells were more abundant in myometrium than in endometrium, their density followed the same pattern as histamine concentration throughout the estrous cycle.     

Estrogen Modulates Neural Control of Muscle Glucose 6-Phosphate Dehydrogenase

Abstract: Exogenously administered estrogens can enhance the rate of increase of glucose 6-phosphate dehydrogenase activity in rat extensor digitorum longus muscles following denervation. In this communication, we report that the effect of denervation on glucose 6-phosphate dehydrogenase activity is modified by variations in endogenous estradiol during the estrous cycle.     

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Abstract

The Gastrin content has been estimated in the antral mucosae of male and female rats in different phases of the estrous cycle. Gastrin content appears to be highest in the rats at di‐estrous cycle end lowest in the rats at pro‐estrous. There is not much difference between the gastrin content at estrous and pro‐estrous. The gastrin content in the antral mucosa of male rats is higher than in rats a both estrous and pro‐estrous but it is about the same amount in rats at di‐estrous. The possible role of the circulating sex hormones on the storage of gastrin in the antral mucosa of rats has been discussed.     

Physiological variation in plasma total homocysteine concentrations in rats

Hyperhomocysteinemia was initially related to cardiovascular diseases; but homocysteine (Hcy) metabolism disturbances have more recently associated with a wide range of pathophysiological conditions including age-related diseases, disrupted circadian rhythms and gynaecological disorders. Since in many cases we do not know to what extent animal models are physiologically similar to human ones, this study aimed to track spontaneous variations in rat plasma Hcy concentrations during different physiological processes such as life cycle, 24 hours and estrous cycle. Plasma total Hcy concentrations were accessed by HPLC. Plasma Hcy concentration varied with age and newborns had the lowest values (2.94 +/- 0.47 micromol/L). Rats aged 10 days presented concentration similar to 3 month old animals (6.87 +/- 0.67 and 8.29 +/- 1.55 micromol/L respectively). Values decreased to 6.42 +/- 1.65 micromol/L at 6 months and 4.87 +/- 0.81 micromol/L at 28 months. Concerning circadian variations in Hcy concentration cosinor analysis showed acrophase in young rats at 1:09 pm, but no plasma Hcy circadian variations in aged rats. Female rats showed changes in Hcy concentration during the estrous cycle with higher values during the diestrous I (10.61 +/- 1.81 micromol/L) compared with the estrous (8.47 +/- 1.86 micromol/L) and diestrous II (7.68 +/- 1.58 micromol/L) phases. In conclusion, plasma Hcy concentration varied spontaneously with ontogenic development and during the estrous cycle and presented a circadian rhythm variation in young rats.     

Orienting behavior of female white rats in estrus cycle and its dependence on handling

Characteristics of orienting behavior of intact and handled female white rats were studied in the "open field" test. Experimental group of animals were formed according to stages of estrous cycle: (1) diestrus, (2) proestrus, (3) estrus, and (4) metestrus. The stage of the cycle was determined by vaginal smears. Over the period of 10 days rats were handled daily for 5 minutes. No behavioral changes over the course of estrous cycle were found in intact rats. Handling revealed some behavioral features related to the levels of steroid hormones in reproductive cycle. Most prominent changes in orienting behavior were observed at transition from estrous to metestrous. At the stage of estrous motor activity was maximal and grooming was minimal. The maximal contrast in the structure of orienting behavior was observed between estrous and diestrous stages.     

Concentrations of 13, 14-dihydro-15-keto-prostaglandin F(2)alpha, estradiol-17beta and progesterone during the peripubertal period in heifers

Twenty prepubertal Holstein heifers were utilized to assess plasma 13, 14-dihydro-15-keto-prostaglandin F(2)alpha (PGFM), serum progesterone (P(4)) and estradiol-17beta (E(2)) concentrations as well as the E(2):P(4) ratio during the onset of puberty in cattle. All animals were maintained as a group along with a sterile marker bull to assist in the detection of estrus. Upon detection of the first estrus (Day=O), daily blood samples were collected from a jugular vein until the heifers had completed 3 estrous cycles. The average body weight and age at first estrus were 247.6+/-4.8 kg and 304.0+/-7.5 days, respectively. Frequency of abnormal length estrous cycles was greater (P<0.02) during the first (40%) and second (35%) cycles than during the third estrous cycle (0%). All heifers had normal cycle lengths (18 to 24 days) by the third estrous cycle. Serum P(4) was greater during the third cycle (P<0.05) from Day 10 to Day 4 before the next estrus compared with the same period of the first estrous cycle. Serum E(2) did not peak until the day of estrus in the first cycle, whereas E(2) reached a maximal level 2 days before estrus in the third estrous cycle. Serum E(2) was higher (P<0.0001) 2 days before estrus in the third cycle than in the first estrous cycle. Plasma PGFM reached maximum concentrations 3 days before estrus in the third cycle compared with 1 day before estrus at the end of first estrous cycle. As estrus approached during the third cycle, PGFM rose 1 day before E(2) rose and P(4) declined, while the rise in PGFM and E(2) occurred simultaneously, with P(4) declining at the end of the first estrous cycle. During diestrus, the E(2):P(4) ratio was lower (P<0.07) in the third cycle than in the first, but it was higher (P<0.04) at estrus and 1 day before in the third estrous cycle. These data reveal a high incidence of abnormal length estrous cycles during the first two estrous cycles of the peripubertal period, and demonstrate anomalies in uterine and ovarian endocrine activity during the peripubertal period in cattle.     

The effect of the estrous cycle and estrogen on the release of immunoreactive alpha-melanocyte-stimulating hormone

Circulating levels and tissue content of alpha-MSH were measured on the morning of various days of the estrous cycle, and on the afternoon of proestrus in freely moving conscious rats. No surges of alpha-MSH were detected by RIA in the morning of various days of the cycle. The neurointermediate lobe content of alpha-MSH was slightly elevated on diestrus 1 as compared to the levels on diestrus 11 and proestrus but not to estrous levels. No changes in alpha-MSH content were detected in the anterior pituitary, the median eminence, mediobasal hypothalamus and the preoptic area at various stages of the estrous cycle. Plasma alpha-MSH levels were slightly elevated at 1500 hr of proestrus which was followed three hours later by a decline. This profile of plasma alpha-MSH on the afternoon of proestrus was reproduced by the SC administration of estradiol benzoate to long-term ovariectomized rats. These data suggest that, contrary to the results obtained by bioassay of alpha-MSH no surges of alpha-MSH occur on any day of the cycle, although a slight elevation on the afternoon of proestrus was detected. The altered pattern of release of this peptide on the afternoon of proestrus may be induced by estrogen.     

Distribution of γ-Aminobutyric Acid and Glutamate Decarboxylase in the Layers of Rat Oviduct

An enzymatic microassay method for glutamate decarboxylase (GAD) and gamma-aminobutyric acid (GABA) was improved to yield a high sensitivity and a low blank. The 20-microns thick freeze-dried sections (0.2-1.5 micrograms dry weight) were prepared from the oviduct and ovary of rat. The analysis of these microsamples by the improved method showed that, contrary to the previous observations, the rat ovary is devoid of GAD activity and contains a trace amount of GABA. Both are present abundantly in the oviduct. In the oviduct mucosa, significant GAD activity was found in the estrous phase, whereas the activity was nearly null during other phases of the estrous cycle. GABA concentration in the oviduct mucosa was 10-fold higher than in the cerebral cortex; its variation during the estrous cycle was not remarkable. In the muscle layer of oviduct, GAD activity had a low peak in the estrous phase and GABA concentration was almost constant during the estrous cycle. The denervation experiment showed that GAD is present in the nerve terminals innervating the oviduct.