Cross-organ interactions between reproductive, gastrointestinal, and urinary tracts: modulation by estrous stage and involvement of the hypogastric nerve

Central nervous system neurons process information converging from the uterus, colon, and bladder, partly via the hypogastric nerve. This processing is influenced by the estrous cycle, suggesting the existence of an estrous-modifiable central nervous system substrate by which input from one pelvic organ can influence functioning of other pelvic organs. Here, we tested predictions from this hypothesis that acute inflammation of colon, uterine horn, or bladder would produce signs of inflammation in the other uninflamed organs (increase vascular permeability) and that cross-organ effects would vary with estrous and be eliminated by hypogastric neurectomy (HYPX). Under urethane anesthesia, the colon, uterine horn, or bladder of rats in proestrus or metestrus, with or without prior HYPX, was treated with mustard oil or saline. Two hours later, Evans Blue dye extravasation was measured to assess vascular permeability. Extravasation was increased in all inflamed organs, regardless of estrous stage. For rats in proestrus, but not metestrus, either colon or uterine horn inflammation significantly increased extravasation in the uninflamed bladder. Much smaller cross-organ effects were seen in colon and uterine horn. HYPX reduced extravasation in the inflamed colon and inflamed uterine horn, but not the inflamed bladder. HYPX eliminated the colon-to-bladder and uterine horn-to-bladder effects. These results demonstrate that inflaming one pelvic organ can produce estrous-modifiable signs of inflammation in other pelvic organs, particularly bladder, and suggest that the cross-organ effects involve the hypogastric nerve and are at least partly centrally mediated. Such effects could contribute to co-occurrence and cyclicity of distressing pelvic disorders in women.     

Exercise pressor reflex function in female rats fluctuates with the estrous cycle

In women, sympathoexcitation during static handgrip exercise is reduced during the follicular phase of the ovarian cycle compared with the menstrual phase. Previous animal studies have demonstrated that estrogen modulates the exercise pressor reflex, a sympathoexcitatory mechanism originating in contracting skeletal muscle. The present study was conducted in female rats to determine whether skeletal muscle contraction-evoked reflex sympathoexcitation fluctuates with the estrous cycle. The estrous cycle was judged by vaginal smear. Plasma concentrations of estrogen were significantly (P < 0.05) higher in rats during the proestrus phase of the estrus cycle than those during the diestrus phase. In decerebrate rats, either electrically induced 30-s continuous static contraction of the hindlimb muscle or 30-s passive stretch of Achilles tendon (a maneuver that selectively stimulates mechanically sensitive muscle afferents) evoked less renal sympathoexcitatory and pressor responses in the proestrus animals than in the diestrus animals. Renal sympathoexcitatory response to 1-min intermittent (1- to 4-s stimulation to relaxation) bouts of static contraction was also significantly less in the proestrus rats than that in the diestrus rats. In ovariectomized female rats, 17β-estradiol applied into a well covering the dorsal surface of the lumbar spinal cord significantly reduced skeletal muscle contraction-evoked responses. These observations demonstrate that the exercise pressor reflex function and its mechanical component fluctuate with the estrous cycle in rats. Estrogen may cause these fluctuations through its attenuating effects on the spinal component of the reflex arc.     

Distribution and changes with age of calcitonin gene-related peptide- and substance P-immunoreactive nerves of the rat urinary bladder and lumbosacral sensory neurons

In the distal parts of the urinary tract, nerves containing calcitonin gene-related peptide (CGRP) or substance P (SP) are sensory with their cell bodies located in lumbosacral dorsal root ganglia. These two neuropeptides are recognised as being present in pelvic sensory nerves, and may be involved in the mediation of pain, stretch and/or vasodilatation. We have used indirect immunohistochemical techniques to examine the distribution and regional variation of nerves immunoreactive (-ir) for CGRP and SP in the urinary bladder and in neurons in lumbosacral dorsal root ganglia (L1-L2 & L6-S1) of young adult (3 months) and aged (24 months) male rats. Semi-quantitative estimations of nerve densities were made for CGRP-ir and SP-ir fibres innervating the dome, body and base of the urinary bladder. Quantitative studies were also used to examine the effects of age on the percentage of dorsal root ganglion neurons immunoreactive for CGRP and SP. There were very few immunoreactive axons in the dome and the overall density of innervation increased progressively towards the base of the bladder. The density of innervation in the aged rats revealed a slight reduction in CGRP and SP innervation of the detrusor muscle but was otherwise comparable to the young group. However, immunostaining of the lumbosacral dorsal root ganglia revealed that the percentage of CGRP- and SP-ir neuronal profiles showed a significant (P < 0.05) reduction from (mean +/- S.D) 44.5 +/- 2; 23.3 +/- 2 in young adult to 25.0 +/- 2.9; 14.8 +/- 1.6 in aged rats, respectively. These findings suggest that the involvement of CGRP and SP in urinary bladder innervation is relatively unchanged in old age, but their expression in dorsal root ganglion neurons is affected by age. The afferent micturition pathway from the pelvic region via these lumbosacral ganglia may be perturbed as a result.     

大鼠动情周期雌激素水平变化对晕动病易感性的影响

目的：众多的流行病学研究和动物实验表明,晕动病存在明显的性别差异,特别是雌激素对晕动病易感性可能存在某些易化的调节作用,本研究为探讨“异食癖”模型上大鼠动情周期雌激素水平的变化对晕动病易感性的影响。方法：大鼠在不同的动情周期,给予足够的旋转刺激以后,通过摄取高岭土量的变化评价大鼠的晕动病反应,同时测定血浆雌激素（E2和P）水平,观察雌激素水平的变化对晕动病易感性的影响。结果：大鼠体内的雌性激素（E2和P）水平随着动情周期而发生波动,在动情期时,E2水平达到最高,而在动情前期则达到最低。P水平在动情间期和动情前期较高而在动情期与动情后期较低。足够的旋转刺激之后,大鼠的摄取高岭土量显著增加,并且呈现与大鼠动情周期雌激素水平波动的一致性,即动情期时摄取高岭土量最多。结论：大鼠动情周期雌激素水平的升高可能在一定程度上会加重大鼠的晕动病反应。可为进一步探讨雌激素水平与晕动病易感性之间的关系提供参考,从而也可能为发现晕动病新病因的研究打下基础,还可能为晕动病预防策略和措施启发新的应用价值。     

Oxytocin secretion induced by osmotic stimulation in rats during the estrous cycle and after ovariectomy and hormone replacement therapy

Hyperosmolality is a potent stimulus for the secretion of oxytocin. Oxytocinergic neurons are modulated by estrogen and oxytocin secretion in rats varies according to the phase of the estrous cycle, with higher activity during proestrus. We investigated the oxytocin secretion induced by an osmotic stimulus (0.5 M NaCl) in female rats. Plasma oxytocin and the oxytocin contents in the neurohypophysis and the paraventricular and supraoptic nuclei were determined during the morning (8-9 h) and afternoon (17-18 h) of the estrous cycle and after ovariectomy followed or not by hormone replacement. Plasma oxytocin peaked in control animals during proestrus. Oxytocin content decreased in the paraventricular and supraoptic nuclei during proestrus and estrus compared to diestrus and increased in the neurohypophysis during proestrus morning. No significant difference was observed in the oxytocin content of the neurohypophysis, nuclei or plasma between ovariectomized animals and ovariectomized animals treated with estrogen or estrogen plus progesterone. Therefore, any ovarian factor other than estrogen or progesterone seems to play a direct or indirect role in the increase in oxytocin secretion. The osmotic stimulus caused an increase in plasma oxytocin throughout the estrous cycle. A reduction in oxytocin content during diestrus and an increase during proestrus were observed in the paraventricular nuclei. In ovariectomized animals, the treatment with estrogen potentiated the response of oxytocin to the osmotic stimulus, with the response being even stronger in the case of estrogen plus progesterone. In conclusion, the ovarian steroids estrogen plus progesterone could modulate the osmoreceptor mechanisms related to oxytocin secretion.     

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.     

Estrous cycle variation of TRPV1-mediated cross-organ sensitization between uterus and NMDA-dependent pelvic-urethra reflex activity

Cross-organ sensitization between the uterus and the lower urinary tract (LUT) underlies the high concurrence of pelvic pain syndrome and LUT dysfunctions, and yet the role of gonadal steroids is still unknown. We tested the hypothesis that cross-organ sensitization on pelvic-urethra reflex activity caused by uterine capsaicin instillation is estrous cycle dependent. When compared with the baseline reflex activity (1.00 +/- 0.00 spikes/stimulation), uterine capsaicin instillation significantly increased reflex activity (45.42 +/- 9.13 spikes/stimulation, P < 0.01, n = 7) that was corroborated by an increase in phosphorylated NMDA NR2B (P < 0.05, n = 4) but not NR2A subunit (P > 0.05, n = 4) expression. Both intrauterine pretreatment with capsazepine (5.02 +/- 2.11 spikes/stimulation, P < 0.01, n = 7) and an intrathecal injection of AP5 (3.21 +/- 0.83 spikes/stimulation, P < 0.01, n = 7) abolished the capsaicin-induced cross-organ sensitization and the increment in the phosphorylated NR2B level (P < 0.05, n = 4). The degrees of the cross-organ sensitization increased in a dose-dependent manner with the concentration of instilled capsaicin from 100 to 300 microM in both the proestrus and metestrus stages, whereas they weakened when the concentrations were higher than 1,000 microM. Moreover, the cross-organ sensitization caused by the uterine capsaicin instillation increased significantly in the rats during the proestrus stage when compared with the metestrus stage (P < 0.01, n = 7). These results suggest that estrogen levels might modulate the cross-organ sensitization between the uterus and the urethra and underlie the high concurrence of pelvic pain syndrome and LUT dysfunctions.     

Effects of pregnancy and female sex steroid hormones on calcitonin gene-related peptide content of mesenteric artery in rats

Calcitonin gene-related peptide (CGRP) levels in plasma and the dorsal root ganglia (DRG) are increased during pregnancy and in ovariectomized rats injected with ovarian hormones. Vasodilatory responses to CGRP are also increased in these animals. In the present study, we hypothesized that pregnancy and ovarian hormones elevate the contents of CGRP in perivascular nerves. We assessed CGRP-dependent mesenteric vascular relaxation induced by electrical field stimulation (EFS) and arterial content of CGRP. Because the mesenteric artery represents resistance vessels, segments of mesenteric arteries collected from female rats at different stages of the estrous cycle, pregnancy, or postpartum and from male rats were used in this study. The EFS-induced relaxation in the presence and absence of CGRP(8-37), an antagonist of CGRP, was used to measure CGRP-dependent relaxation, and radioimmunoassay (RIA) of tissue homogenates was used to assess changes in CGRP content in mesenteric branch arteries. The results show that CGRP-dependent, EFS-induced relaxation response was lower in female rats at diestrus and proestrus than in male rats, and no statistically significant differences were observed between Gestational Day 20 and Postpartum Day 2. The RIA revealed significantly lower mesenteric artery CGRP levels in female rats at proestrus, gestation, and postpartum than in female rats at diestrus or in male rats. We conclude that no correlation exists between CGRP-dependent, EFS-induced relaxation and CGRP content in the mesenteric arteries of these animal groups. Because both CGRP levels in DRG and serum are reported to be elevated, the reduced CGRP content in the vasculature during pregnancy and proestrus implicate enhanced basal release of CGRP at the nerve terminal in these animals.     

The influence of estrogen on the hyperglycemic action of alloxan in female rats

The purposes of this study were to investigate (1) the relationship between estrogen and glucose levels in the circulation during an estrous cycle of the rat and (2) the effect of estrogen on the hyperglycemic action of alloxan. Three-month old Long Evans strain rats were used. The present study established for the first time the correlation between estrogen and glucose levels in the circulation during an estrous cycle. A significant (p less than 0.01) negative correlation was observed with the lowest glucose level at the proestrus stage where estrogen is at peak. The severity of hyperglycemia induced by alloxan was positively correlated with the circulating level of endogenous estrogen, but was negatively correlated with the fasting glucose level. A similar pattern was observed in ovariectomized female rats that were followed by estradiol implantation to achieve a constant level of circulating estrogen. It may be concluded from the present study that estrogen, under physiological conditions, participates in the regulation of glucose metabolism during various stages of the estrous cycle in the rats, and that glucose is likely able to protect the pancreatic beta cells of the animals against the hyperglycemic action of alloxan.     

Organ cross talk modulates pelvic pain

Interstitial cystitis (IC) is a chronic bladder inflammatory disease of unknown etiology that is often regarded as a neurogenic cystitis. IC is associated with urothelial lesions, voiding dysfunction, and pain in the pelvic/perineal area, and diet can exacerbate IC symptoms. In this study, we used a murine neurogenic cystitis model to investigate the development of pelvic pain behavior. Neurogenic cystitis was induced by the injection of Bartha's strain of pseudorabies virus (PRV) into the abductor caudalis dorsalis tail base muscle of female C57BL/6J mice. Infectious PRV virions were isolated only from the spinal cord, confirming the centrally mediated nature of this neurogenic cystitis model. Pelvic pain was assessed using von Frey filament stimulation to the pelvic region, and mice infected with PRV developed progressive pelvic pain. Pelvic pain was alleviated by 2% lidocaine instillation into either the bladder or the colon but not following lidocaine instillation into the uterus. The bladders of PRV-infected mice showed markers of inflammation and increased vascular permeability compared with controls. In contrast, colon histology was normal and vascular permeability was unchanged, suggesting that development of pelvic pain was due only to bladder inflammation. Bladder-induced pelvic pain was also exacerbated by colonic administration of a subthreshold dose of capsaicin. These data indicate organ cross talk in pelvic pain and modulation of pain responses by visceral inputs distinct from the inflamed site. Furthermore, these data suggest a mechanism by which dietary modification benefits pelvic pain symptoms.     

Gender and estrous cycle influences on behavioral and neurochemical alterations in adult rats neonatally administered ketamine

Neonatal N‐methyl‐D‐aspartate (NMDA) receptor blockade in rodents triggers schizophrenia (SCZ)‐like alterations during adult life. SCZ is influenced by gender in age of onset, premorbid functioning, and course. Estrogen, the hormone potentially driving the gender differences in SCZ, is known to present neuroprotective effects such as regulate oxidative pathways and the expression of brain‐derived neurotrophic factor (BDNF). Thus, the aim of this study was to verify if differences in gender and/or estrous cycle phase during adulthood would influence the development of behavioral and neurochemical alterations in animals neonatally administered ketamine. The results showed that ketamine‐treated male (KT‐male) and female‐in‐diestrus (KTF‐diestrus, the low estrogen phase) presented significant deficits in prepulse inhibition of the startle reflex and spatial working memory, two behavioral SCZ endophenotypes. On the contrary, female ketamine‐treated rats during proestrus (KTF‐proestrus, the high estradiol phase) had no behavioral alterations. This correlated with an oxidative imbalance in the hippocampus (HC) of both male and KTF‐diestrus female rats, that is, decreased levels of GSH and increased levels of lipid peroxidation and nitrite. Similarly, BDNF was decreased in the KTF‐diestrus rats while no alterations were observed in KTF‐proestrus and male animals. The changes in the HC were in contrast to those in the prefrontal cortex in which only increased levels of nitrite in all groups studied were observed. Thus, there is a gender difference in the adult rat HC in response to ketamine neonatal administration, which is based on the estrous cycle. This is discussed in relation to neuropsychiatric conditions and in particular SCZ. © 2015 Wiley Periodicals, Inc. Develop Neurobiol 76: 519–532, 2016     

Acute and chronic estrogen supplementation decreases uterine sympathetic innervation in ovariectomized adult virgin rats

Uterine innervation undergoes substantial reorganization associated with changes in reproductive status. Nerves innervating the uterus are decreased in pregnancy and puberty, and even the normal rodent estrous cycle is characterized by fluctuations in numbers of myometrial nerve fibers. During the follicular (proestrus/estrous) phase of the estrous cycle, intact nerves are rapidly depleted and then return over the next 2-3 days in the luteal (metestrus/diestrus) phase. We hypothesize that uterine nerve depletion is initiated by increased circulating estrogen in the follicular phase. However, studies have not shown whether estrogen can reduce uterine innervation and, if so, whether the time course is compatible with the rapid changes observed in the estrous cycle. These questions were addressed in the present study. Mature ovariectomized virgin rats received 17-beta-estradiol as a single injection (10 microg/kg s.c.) or chronically from timed-release pellets (0.1 microg/pellet for 3 weeks sustained release). Total (protein gene-product 9.5-immunoreactive) and sympathetic (dopamine beta-hydroxylase-immunoreactive) uterine innervation was assessed quantitatively. Both total and sympathetic innervation was abundant in uterine longitudinal smooth muscle of ovariectomized rats. However, following acute or chronic estrogen administration, total and sympathetic fiber numbers were markedly decreased. This was not due to altered uterine size, as reductions persisted after correcting for size differences. Our results indicate that sympathetic nerves are lost from uterine smooth muscle after estradiol treatment in a manner similar to that seen in the intact animal during estrus and pregnancy. This suggests that the rise in estradiol prior to estrus is sufficient to deplete uterine sympathetic innervation.     

Amygdala activation by corticosterone alters visceral and somatic pain in cycling female rats

Irritable bowel syndrome (IBS) is often seen in women, and symptom severity is known to vary over the menstrual cycle. In addition, activation of the hypothalamic-pituitary-adrenal (HPA) axis enhances symptomology and patients with IBS have increased activation of the amygdala, a brain region known to facilitate HPA output. However, little is known about the effects of amygdala activation during different stages of the menstrual cycle. We therefore investigated the effects of amygdala activation on somatic and visceral pain perception over the rat estrous cycle. Female Wistar rats were implanted with either corticosterone (Cort) or cholesterol as a control onto the dorsal margin of the central amygdala. Visceral sensitivity was quantified by recording the visceromotor response (VMR) to colorectal distension (CRD) and somatic sensitivity was assessed via the Von Frey test. In cholesterol controls, both visceral and somatic sensitivity varied over the estrous cycle. Rats in proestrus/estrus responded to CRD with an increased VMR compared with rats in metestrus/diestrus. Somatic sensitivity followed a similar pattern with enhanced sensitivity during proestrus/estrus compared with metestrus/diestrus. Elevated amygdala Cort induced visceral hypersensitivity during metestrus/diestrus but had no effect during proestrus/estrus. In contrast, elevated amygdala Cort increased somatic sensitivity during both metestrus/diestrus and proestrus/estrous. These results suggests that amygdala activation by Cort eliminates spontaneously occurring differences in visceral and somatic pain perception, which could explain the lowered pain thresholds and higher incidence of somatic pain observed in women with IBS.     

Striatal D-2 dopamine agonist binding sites fluctuate during the rat estrous cycle

Striatal D-2 dopamine (DA) antagonist and agonist binding sites were measured during the rat estrous cycle and compared to ovariectomized (OVX) rats. Dopaminergic D-2 antagonist binding sites were constant during the estrous cycle while agonist binding sites show a rapid and significant decrease of the ratio of high to low D-2 agonist binding sites from proestrus AM (PAM) to diestrus 1 (D1) and return to OVX value in diestrus 2 (DII). Thus, physiological fluctuations of hormones as occur during the estrous cycle can modulate extrahypothalamic biogenic amine activity, namely striatal DA systems which are not involved in the control of hormone secretion.     

Estrous cycle-dependent changes in the expression of aromatic hydrocarbon receptor (AHR) and AHR-nuclear translocator (ARNT) mRNAs in the rat ovary and liver

The aromatic hydrocarbon receptor (AHR) and AHR nuclear translocator protein (ARNT) mediate the toxic effects of a wide variety of halogenated and polycyclic aromatic hydrocarbons. While it can be assumed that AHR has an endogenous function, its role in reproduction is currently undefined. The present study seeks to examine the regulation of AHR and ARNT mRNAs in liver and ovarian tissues across the rat estrous cycle. Message for hepatic AHR was increased significantly on the morning of proestrus, and decreased dramatically by the evening of proestrus; while hepatic ARNT mRNA was significantly decreased between diestrus and the morning of proestrus, and between the evening of proestrus and the morning of estrus. Ovarian AHR mRNA was unchanged from diestrus to proestrus, and was decreased on the evening of proestrus. Changes in the expression of ARNT mRNA mirrored changes in the liver. To assess interaction between the AHR- and estrogen-receptor (ER)-signaling pathways and to test the hypothesis that estrogen regulates AHR mRNA, 25-day-old female rats were injected with either 17beta-estradiol, the ER antagonist ICI 182 780, or with vehicle, and hepatic AHR mRNA was measured. Treatment with estrogen or the estrogen antagonist did not alter the abundance of AHR mRNA in the liver. These data suggest that while estrogen may not be the key regulator of AHR mRNA expression, a factor associated with the rat reproductive cycle may be important in regulating the expression of both the AHR and ARNT genes in the ovary and liver.     

Calcitonin gene-related peptide, neurokinin A and substance P: effects on nociception and neurogenic inflammation in human skin and temporal muscle.

Calcitonin gene-related peptide (CGRP) was injected alone and in combination with substance P (SP) or neurokinin A (NKA) into the forearm skin and temporal muscle of human volunteers. In the skin, 50 pmol of CGRP induced a wheal response and a delayed erythema. No pain was recorded. No interaction between CGRP and SP or NKA was observed. In the temporal muscle, 200 pmol of CGRP alone did not induce pain or tenderness but, in combination with SP or NKA, CGRP elicited a significant pain sensation. It is concluded that CGRP may be involved in neurogenic inflammation and that only SP, of the three peptides present in nociceptive C fibers, seems to be of major importance in relation to cutaneous nociception. Simultaneous neurogenic release of CGRP and other neuropeptides in skeletal muscle may induce myofascial pain.     

The effects of paradoxical sleep deprivation on estrous cycles of the female rats

The present purpose was to examine how sleep deprivation affects the estrous cycle of the female rat. Paradoxical sleep-deprived (PSD) adult female Wistar rats were compared to home-cage control (CTRL) on their estrous cyclicity. Forty-four PSD and forty-four CTRL female rats were distributed into 4 subgroups of 11 animals each according to the phase of estrous cycle and were subjected to sleep deprivation for 96 h by the multiple platform technique. After PSD period, vaginal estrous cycles were taken for an additional 9 days. Animals that were submitted to PSD in diestrus phase (PSD-diestrus) had their estrous cycles disrupted during the recovery period by showing a constant diestrus during the first week. As for hormone alterations, progesterone concentrations were statistically higher in PSD-diestrus compared to respective phase control and to PSD-proestrus and PSD-estrus rats while CTRL-metestrus had higher levels than CTRL-proestrus and estrus groups. Testosterone was significantly decreased in PSD-estrus in relation to PSD-proestrus and PSD-diestrus groups and was lower in CTRL-diestrus rats than in home-cage rats in proestrus. In addition, PSD-diestrus phase exhibited higher concentrations of corticosterone and lower estrogen than the respective control rats. These data indicate that PSD may modulate the ovarian hormone release through alterations in hormonal-neurochemical mechanisms.     

Increased LH pulse frequency and estrogen secretion associated with termination of anestrus followed by enhancement of uterine estrogen receptor gene expression in the beagle bitch

The relationships among pulsatile LH secretion pattern, estrogen secretion, and expression of the uterine estrogen receptor gene were examined throughout the estrous cycle in beagle bitches. In Experiment 1, blood samples were collected from 30 bitches every 10 min for 8 h from a cephalic vein during different phases of the estrous cycle. An increase in the mean plasma levels of LH occurred from mid to late anestrus (P < 0.01). The LH pulse frequency increased (P < 0.01) from late anestrus to proestrus, and was strongly correlated (r = 0.96, P < 0.001) with the mean plasma level of estradiol-17 beta (E2). In Experiment 2, middle uterine samples, including the myometrium and endometrium, from 18 bitches were taken at 6 stages of the estrous cycle. The total number of estrogen receptors and nuclear estrogen receptor and its mRNA levels in the uterus also increased (P < 0.01) from late anestrus to proestrus. Mean plasma E2 level and the number of uterine estrogen receptor were positively correlated (r = 0.81, P < 0.05). In Experiment 3, nine bitches were ovariectomized in mid anestrus. Two weeks later they received a single injection of 10 or 50 micrograms/kg, i.m., estradiol benzoate. The number of uterine estrogen receptor and their mRNA levels for ovariectomized bitches were low, but increased (P < 0.05) after treatment with a low dose of estradiol benzoate. These results suggest that increases in LH pulse frequency and estrogen secretion are associated with termination of anestrus and that subsequent enhancement of uterine estrogen receptor expression may be up-regulated by estradiol.     

Effects of thymulin and GnRH on the release of gonadotropins by in vitro pituitary cells obtained from rats in each day of estrous cycle

The effects of thymulin and GnRH on FSH and LH release were studied in suspension cultures of anterior pituitary cells from female adult rats sacrificed on each day of the estrous cycle. The spontaneous release of gonadotropins by pituitaries, as well as their response to GnRH or thymulin addition, fluctuated during the estrous cycle. Adding thymulin to pituitary cells from rats in diestrus 1 increased the concentration of FSH; while in cells from rats in estrus, FSH level decreased. Thymulin had a stimulatory effect on the basal concentration of LH during most days of the estrous cycle. Adding GnRH increased FSH release in cells from rats in diestrus 1, diestrus 2, or proestrus, and resulted in higher LH levels in cells obtained from rats in all days of the estrous cycle. Compared to the GnRH treatment, the simultaneous addition of thymulin and GnRH to cells from rats in diestrus 1, diestrus 2, or proestrus resulted in lower FSH concentrations. Similar results were observed in the LH release by cells from rats in diestrus 1, while in cells from rats in proestrus or estrus, LH concentrations increased. A directly proportional relation between progesterone serum levels and the effects of thymulin on FSH release was observed. These data suggest that thymulin plays a dual role in the release of gonadotropins, and that its effects depend on the hormonal status of the donor's pituitary.