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Serum luteinizing hormone, prolactin, and thyrotropin and their pituitary subunit mRNA levels during proestrus in the Syrian hamster.

mRNA levels for alpha, luteinizing hormone beta (LH beta), and prolactin (Prl) were examined during the hamster estrous cycle, with sampling most frequent (1-hour intervals) on the afternoon of proestrus. These transcripts encode the peptide subunits for the pituitary hormones LH and Prl which are necessary for reproductive function. Serum hormone levels of LH and Prl, analyzed by 24-hour periodic regression, exhibited a 24-hour periodicity on proestrus characterized by a large surge peaking at about 18.00 h. Combining the data for non-proestrous days of the cycle disclosed a rhythm with similar timing for LH and Prl. Thyroid-stimulating hormone (TSH) and TSH beta RNA profiles during hamster proestrus are reported for the first time. Serum TSH exhibited a pronounced peak coincident with that of the other hormones on proestrus. Because of variations at other times on the day of proestrus, however, a 24-hour periodicity was not manifested by regressional analysis. Combined non-proestrous serum TSH data also revealed no consistently timed regressional 24-hour periodicity. During proestrus, pituitary mRNA values for alpha, LH beta, and Prl simultaneously exhibited a rise from the lowest to the highest of all proestrous values in the 3-5 h prior to the time of the pre-ovulatory peak of circulating hormone concentrations. RNA for TSH beta exhibited an earlier, broader peak on proestrus. Periodic regression indicated a significant 24-hour rhythm for alpha mRNA in data pooled from non-proestrous days (acrophase 05.00 h) and for TSH beta mRNA on proestrus (acrophase 04.54 h).(ABSTRACT TRUNCATED AT 250 WORDS)

Analysis of Variance

Ovarian steroid regulation of basal pulsatile luteinizing hormone release between the mornings of proestrus and estrus in the rat.

The aim of this study was to examine the regulation of basal pulsatile LH release by ovarian estradiol (E2) and progesterone (P) during the interval between the mornings of proestrus and estrus in the rat estrous cycle. Pulsatile LH release was studied in six groups of rats bled continuously through jugular venous cannulae between 0930-1230 h at a rate of 50 microliter whole blood/5 min: 1) bled on proestrus; 2) sham ovariectomy (OVX) at 0900-1000 h on proestrus and bled on estrus; 3-6) OVX at 0900-1000 h on proestrus, implanted with either empty or E2-, P-, or E2- plus P-containing Silastic capsules, and bled 24 h after OVX. In our colony, plasma E2 levels peaked at 1300 h, remained high through 1730 h, and then declined. Plasma P values increased between 1300 and 1730 h, peaked at 2000 h, and were rapidly declining by 2400 h. To reproduce the magnitude as well as the temporal pattern for these changes in plasma E2 and P levels, E2 capsules were inserted at the time of OVX on proestrus and removed at 1830 h. P capsules were inserted at 1400 h and removed at 2300 h. Groups of ovariectomized or sham-ovariectomized control animals had empty capsules implanted and removed at comparable times. Capsules producing basal E2 and P levels were not inserted after the removal of the original implant, since mean blood LH levels, pulse amplitude, and frequency were the same in rats sham ovariectomized or ovariectomized at 1830 h on proestrus and bled the next morning between 0930-1230 h. Mean blood LH levels decreased between the mornings of proestrus and estrus due to a reduction in LH pulse frequency as pulse amplitude remained stable. OVX at 0900-1000 h on proestrus increased mean blood LH levels 2.5-fold compared to values on estrus due to increases in both LH pulse frequency and amplitude. Restoration of physiological proestrous levels of only E2 returned LH pulse frequency to estrous values, but did not significantly affect LH pulse amplitude. P alone also had no significant effect on LH pulse amplitude, but slightly reduced pulse frequency, although, unlike E2, not to values seen on estrus. Replacing both E2 and P returned LH pulse amplitude to estrous levels and reduced LH pulse frequency.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals

Progesterone suppresses tyrosine hydroxylase messenger ribonucleic acid levels in the arcuate nucleus on proestrus.

This study examined the intracellular mechanisms for the regulation of tyrosine hydroxylase in the tuberoinfundibular dopaminergic neurons of cycling female rats. It also evaluated the hormonal influences that contribute to the control of this enzyme on proestrus. Tyrosine hydroxylase messenger RNA (mRNA) levels in the arcuate nucleus of the hypothalamus were assessed by in situ hybridization. Tyrosine hydroxylase activity in the stalk-median eminence was determined from the in vitro or in vivo rate of 3,4-dihydroxyphenylalanine (DOPA) accumulation after inhibiting DOPA decarboxylase with brocresine or m-hydroxybenzylhydrazine, respectively. Tyrosine hydroxylase mRNA levels and in vitro DOPA accumulation were similar on diestrous day 2 and proestrous mornings, but were reduced by 50% on estrus. Although circulating PRL concentrations were similar on the morning of each day of the estrous cycle, a broad preovulatory PRL surge was observed on the afternoon of proestrus. In vitro DOPA accumulation was similar at 1000 h before the PRL surge and at 1330 h during the peak phase of the PRL surge, but was reduced during the plateau phase of the PRL surge (1700 and 2200 h) coincident with the preovulatory progesterone rise and remained low on estrus. However, in vivo DOPA accumulation was transiently decreased only at 1700 h on proestrus. Tyrosine hydroxylase mRNA levels were similar at 1000, 1330, and 1700 h on proestrus, were reduced by 50% at 2200 h on proestrus subsequent to the decrease in enzyme activity, and remained low on the morning of estrus. Okadaic acid, a protein phosphatase-1 and -2A inhibitor, induced a similar increase in tyrosine hydroxylase activity in vitro at 1330 and 2200 h on proestrus and at 1100 h on estrus, indicating that tyrosine hydroxylase was capable of being activated in spite of decreased mRNA levels. Ovariectomy between 1100-1200 h on proestrus prevented the decrease in tyrosine hydroxylase mRNA levels and in vitro DOPA accumulation at 2200 h. The effects of ovariectomy were completely reversed by progesterone, whereas estradiol had no effect. Circulating PRL levels at 2200 h were suppressed to basal levels after ovariectomy, but were increased by progesterone treatment at 1530 h to levels similar to those in the plateau phase of the PRL surge in control rats. Administration of the progesterone antagonist RU486 at 1200 h on proestrus did not alter tyrosine hydroxylase activity, tyrosine hydroxylase mRNA levels, or circulating PRL concentrations at 2200 h.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals

The antiprogestins RU486 and ZK98299 affect follicle-stimulating hormone secretion differentially on estrus, but not on proestrus.

Previous in vivo studies from our laboratory indicated that administration of the antiprogestin RU486 on proestrus suppresses both the preovulatory gonadotropin surges and the secondary FSH surge, suggesting a role for the progesterone receptor (PR) in the generation of these surges. The present study was designed to test the effects of another antiprogestin, ZK98299, which has been reported to block the PR through a mechanism different from that of RU486, on gonadotropin secretion in vivo. RU486 and ZK98299 (2 and 6 mg/kg) were administered s.c. at 1230 h on proestrus; uterine intraluminal fluid content, serum gonadotropins, and gonadotropin subunit messenger RNAs (mRNAs) were determined at 1830 h on proestrus and at 0900 h on estrus. At 1830 h on proestrus, both RU486 and ZK98299 at both doses caused equal suppression of the preovulatory FSH surge and FSHbeta mRNA. Both antiprogestins also equally attenuated the preovulatory LH surge at this time, with the higher doses causing greater suppression. In contrast, at 0900 h on estrus, the antiprogestins affected serum FSH differentially; only RU486 suppressed the secondary FSH surge despite the fact that both drugs prevented the release of uterine intraluminal fluid, confirming blockade of progesterone action at the level of the uterus. Neither drug had a significant effect on FSHbeta mRNA at 0900 h on estrus. ZK98299 at the higher dose caused a small, but significant, increase in serum LH. In a subsequent experiment, we compared the effects of RU486 and ZK98299 (6 mg/kg, s.c.), administered at 1230 h on proestrus, on serum FSH raised above the natural secondary FSH surge on the morning of estrus by passive immunization with an antiserum to inhibin-alpha (anti-I) at 1700 h on proestrus. Consistent with the results of the first experiment, both antiprogestins blocked the release of uterine intraluminal fluid, but only RU486 lowered serum FSH in both the normal sheep serum-treated controls and anti-I-treated rats; in contrast, ZK98299 actually increased serum FSH in the normal sheep serum-treated control animals. ZK98299 also increased FSHbeta mRNA in the control group; RU486, on the other hand, reduced FSHbeta mRNA only in the anti-I group. The results demonstrate unequivocally that whereas the effects of the two antiprogestins on serum FSH and FSHbeta mRNA are similar on proestrus, they are divergent on estrus. The data suggest that the functional state of the PR/transcriptional activation complex in the gonadotrope on the morning of estrus is different from that on the evening of proestrus.

Animals

Effect of recombinant inhibin on gonadotropin secretion during proestrus and estrus in the rat.

This work investigated the ability of recombinant human (rh) inhibin A or the GnRH antagonist [Ac-D2Nal1, delta Cpa2, delta 3Pal3, Arg5 delta 5-(p-methoxyphenyl)5-oxo-2-aminopentanoic acid6, delta Ala10]GnRH to alter plasma immunoreactive LH and FSH levels in cycling female rats. In a first series of experiments, rh inhibin A (25 micrograms/kg) was injected iv between 0800 and 0830 h, or at 0800 and 1200 h, on proestrus, and plasma hormone levels were measured from 1200-1900 h. Both regimens of administration significantly (P less than or equal to 0.01) lowered mean plasma FSH levels but did not mask the primary FSH surge. This treatment also lowered the overall amount of LH released, although the difference between control and inhibin-treated rats only reached statistical significance (P less than or equal to 0.05) after two injections of the protein. Measurement of the number of tubal ova shed on the next estrus showed no difference between rats injected with the vehicle or inhibin at 0830 h. Finally, it was shown that administration of the GnRH antagonist (100 micrograms/kg) at 1200 h interfered with the primary surges of both LH and FSH. In a second series of experiments, rh inhibin A (25 micrograms/kg) was injected once at 2000 h on proestrus. In these animals, inhibin significantly (P less than or equal to 0.01) decreased mean plasma FSH levels between 2000 h on proestrus and 0500 h on estrus and totally suppressed the secondary FSH surge. LH secretion remained low in control animals, and no measurable changes were observed after inhibin treatment. Flushing of the ova 5 days later (i.e. during estrus of the subsequent cycle) showed no difference between control and inhibin-treated rats. Injection of the GnRH antagonist (100 micrograms/kg) at 2000 h on proestrus decreased the total amount of FSH secreted during late proestrus and early estrus. However, FSH secretion showed an increase at between 0100 and 0400 h of estrus despite blockade of GnRH receptors. These results indicate that administration of rh inhibin A interferes with both the primary and secondary FSH surges. In contrast to its inability to alter LH release by ovariectomized rats, inhibin also blunted LH secretion during the afternoon of proestrus. Whether these results represent differential effects of inhibin on LH secreted by intact and gonadectomized animals, or whether the documented changes in pituitary responsiveness to GnRH during proestrus are accompanied by an increased sensitivity to inhibin, needs further investigation.

Animals

Interleukin-1 beta inhibits the endogenous expression of the early gene c-fos located within the nucleus of LH-RH neurons and interferes with hypothalamic LH-RH release during proestrus in the rat.

The ability of central interleukin-1 beta (IL-1 beta) administration to modulate the hypothalamic LH-RH release as well as the endogenous expression of the c-fos protein located within the nucleus of LH-RH neurons was examined during the afternoon of proestrus in rats. In a first series of experiments, 50 or 100 ng IL-1 beta were infused into the lateral ventricle of the rat brain at either 08.30, 12.00, 14.30, or 17.00 h of proestrus. The animals were then perfused transcardially with a solution of 4% paraformaldehyde from 17.30 and 18.00 h. In a second series of experiments, the rats were equipped with an intracerebroventricular (i.c.v.) cannula in the lateral ventricle and a push-pull cannula into the median eminence (ME), and LH-RH secretion was measured during the afternoon of proestrus. The third experiment investigated the putative role of corticotropin-releasing factor (CRF) in modulating the inhibitory effect of IL-1 beta on LH secretion by infusing CRF antagonists before the i.c.v. administration of the cytokine to gonadectomized male and female rats. The central infusion of 50 or 100 ng IL-1 beta at 12.00 h completely blocked the spontaneous expression of c-fos protein which normally occurs in the nucleus of LH-RH neurons between 17.30 and 18.00 h on proestrus. In contrast, 50 ng IL-1 beta was less effective (P < 0.05) when administered at 08.30 h, and totally without effect when infused at 14.30 h. Infusion of 50 ng IL-1 beta also markedly suppressed the hypothalamic release of LH-RH in proestrus rats bearing a push-pull cannula into the ME, and significantly decreased plasma LH levels in both gonadectomized male and female rats. Finally, we observed that the central administration of CRF antagonists did not modify the inhibitory effects of the cytokine on the activity of the hypothalamic-pituitary-gonadal (HPG) axis. These results provide the first direct evidence that IL-1 beta is a potent inhibitor of LH-RH neuronal activity during the proestrus LH surge in intact cycling rats. As central administration of this cytokine completely inhibited the endogenous expression of c-fos protein within the nucleus of LH-RH neurons, our findings also suggest that IL-1 beta acts at the level of LH-RH perikarya.

Animals

GABAergic neuronal activity and mRNA levels for both forms of glutamic acid decarboxylase (GAD65 and GAD67) are reduced in the diagonal band of Broca during the afternoon of proestrus.

There is considerable evidence that GABAergic neurons play an important role in the regulation of gonadotropin-releasing hormone (GnRH) secretion, and that these neurons may mediate the feedback actions of gonadal steroids on GnRH neurons. The aim of the present study was to investigate whether endogenous changes in ovarian steroid secretion during the estrous cycle influenced GABAergic neuronal activity in the preoptic region of the hypothalamus, and in other steroid-sensitive brain regions. Intact, adult female rats were sacrificed at various times during the days of metestrus or proestrus. GABAergic neuronal activity was estimated by measuring the rate of accumulation of GABA in microdissected brain regions after pharmacological inhibition of GABA degradation. Concentrations of mRNA for both forms of glutamic acid decarboxylase (GAD65 and GAD67) were quantified in microdissected brain regions by a microlysate ribonuclease protection assay. In the diagonal band of Broca at the level of the organum vasculosum of the lamina terminalis (DBB(ovlt)), GABAergic neuronal activity was significantly reduced during the afternoon of proestrus compared with the morning of either proestrus or metestrus. In the lateral septal nucleus, GABAergic neuronal activity was significantly increased in the afternoon of proestrus compared with the morning. There were no significant effects of time of day or day of estrous cycle in the medial preoptic nucleus, median eminence, ventromedial nucleus, suprachiasmatic nucleus, medial septal nucleus, hippocampus (CA1 region), or cingulate cortex. In the DBB(ovlt), mRNA levels for both GAD65 and GAD67 were significantly reduced in the afternoon of proestrus compared with the afternoon of metestrus. By contrast, there was no change in GAD65 and GAD67 mRNA levels in the cingulate cortex at any of the times examined. These results demonstrate that GABAergic neuronal activity, and mRNA levels for both GAD65 and GAD67, are reduced in the DBB(ovlt) during the afternoon of proestrus. These results support the hypothesis that decreased GABAergic neuronal activity in this region plays a major permissive role in the generation and maintenance of the estrogen-induced LH surge.

Animals

Concentrations of reproductive hormones in canine serum throughout late anestrus, proestrus and estrus.

Concentrations of estradiol, progesterone, luteinizing hormone (LH), follicle-stimulating hormone (FSH) and prolactin in serum from 6 bitches bled daily for at least 45 days before the onset of proestrus, during proestrus and estrus were determined by radioimmunoassay. Mean concentrations of estradiol in serum were high early in the sampling period (20 to 46 pg/ml) and appeared to decrease prior to the onset of proestrus (8 to 19 pg/ml). There were sporadic increases in serum concentrations of LH throughout the sampling period in each bitch. Five of the 6 bitches sampled had increased serum concentrations of LH following the low mean concentration of estradiol just before the onset of proestrus. Mean concentrations of FSH were highest during anestrus (240 to 294 ng/ml) and near the time of the preovulatory surge of LH (297 ng/ml) and were lowest during proestrus (131 to 200 ng/ml). The mean concentration of progesterone for the 6 bitches remained at less than 1.0 ng/ml throughout late anestrus, but increased to greater than 1.0 ng/ml the day of the maximum mean concentration of LH (preovulatory LH surge). Mean concentrations of prolactin were variable throughout the sampling period and demonstrated no consistent pattern among bitches. The results of the current investigation suggest that neither the canine ovary nor pituitary are quiescent during anestrus. The bitch appears to have sufficient FSH present during anestrus for follicular growth. Serum concentrations of LH appear to increase prior to the onset of proestrus when concentrations of estradiol are lowest, possibly inducing a new follicular phase. Progesterone and prolactin do not appear to be involved in the termination of anestrus in the bitch.

Anestrus

Ovarian steroid regulation of pulsatile luteinizing hormone release during the interval between the mornings of diestrus 2 and proestrus in the rat.

The object of this study was to determine the influence of ovarian steroids on pulsatile LH release in the interval between the mornings of diestrus 2 (D2) and proestrus in the rat. Four groups of rats were bled continuously for 3 h between 09.30-12.30 h at a rate of 75 microliters whole blood/6 min: bled on D2; sham ovariectomy (OVX) on D2 and bled on proestrus; OVX on D2, implanted with empty or oil-filled capsules, and bled 24 h later; and OVX on D2, implanted with estradiol (E2) capsules, and bled 24 h later. Between D2 and proestrus, plasma E2 levels increased from 13 +/- 1 to 42 +/- 9 pg/ml, and progesterone levels decreased from 27 +/- 3 to 13 +/- 2 ng/ml, the latter reflecting the decline of the corpus luteum early on D2. Between D2 and proestrus there was no change in mean blood LH levels, LH pulse amplitude, or pulse frequency. However OVX on D2 increased mean blood LH levels 2.5-fold over values on proestrus due to a 3.5-fold elevation in LH pulse amplitude and an 80% increase in pulse frequency. E2 levels fell in these rats to 8 +/- 1 pg/ml. Restoration of physiological proestrous levels of E2 (46 +/- 5 pg/ml) significantly reduced the increase in mean blood LH levels by lowering pulse frequency to proestrous values, and by causing a 50% reduction in pulse amplitude. However, LH pulse amplitude and therefore mean blood LH levels were still higher than values on proestrus.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals

Evidence that a decrease in opioid tone on proestrus changes the episodic pattern of luteinizing hormone (LH) secretion: implications in the preovulatory LH hypersecretion.

We have studied the LH secretion pattern evoked by diminution in the opioid tone produced by iv naloxone (NAL) infusion between 1100-1400 h on proestrus, the LH secretion pattern occurring spontaneously between 1430-1730 h on proestrus and the LH secretion pattern produced by exogenous LHRH administered either as a 10 ng/pulse at 20-, 30-, or 60-min intervals or infused continuously at a rate of 30 ng/h between 1200-1700 h in rats given pentobarbital at 1100 h on proestrus. Infusion of 0.5 ng NAL/h raised plasma NAL levels to 200-300 ng/ml and augmented LH secretion, as evident by increments in pulse amplitude and frequency discharge to one every 37.5 min from an average of one every 75 min in saline-infused control rats. A 4-fold increase in circulating NAL levels, produced by 2 mg/h NAL infusion, further augmented the frequency of LH episodes to 30-33 min and induced a surge-like LH secretion pattern which resembled that seen on the afternoon of proestrus. Further analysis of the secretory pattern of the preovulatory LH surge (n = 7) showed LH pulses of increased amplitude during the basal phase (n = 4), ascending phase (n = 2), and plateau and descending phases (n = 3); in two rats the LH rise was steep, and no LH pulses were identified. A LHRH pulse (10 ng/pulse) delivered at 20- or 30-min intervals or continuous infusion of LHRH at a rate of 30 ng/h produced LH surges, with peak levels reaching the range seen on the afternoon of proestrus. Further, despite the fact that 10 ng LHRH/pulse at 20-min intervals reproduced a proestrous-type LH surge, only 40% of the LHRH pulses were followed by identifiable LH pulses. Surprisingly, despite the observations that NAL evoked robust LH episodes, the basal pattern of FSH secretion in these rats was not altered. These findings show that a decrease in opioid tone on proestrus accelerates episodic LH discharge to the range that occurs after gonadectomy. A quantitative relationship between the degree of restraint on the opioid tone imposed by NAL and the magnitude of the LH response can be demonstrated. The evidence suggests that the preovulatory LH surge may occur in an episodic fashion and that it can be reproduced by LHRH delivered at a frequency rate of LH pulses seen in ovariectomized rats.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals

Ovary mediates the effects of RU486 given during proestrus on the diestrous secretion of luteinizing hormone in the rat.

The aim of these experiments was to study the action of proestrous afternoon follicular progesterone secretion on the preovulatory secretion of gonadotropins in the rat. Four-day-cycling rats were given 4 mg of the antiprogestagen RU486 in the morning of proestrus (Day 1), and its effects on the pituitary function during diestrus were compared with those of RU486 given in the morning of estrus (Day 2). The pituitary function was assessed by measuring basal secretion of LH and FSH as well as the pituitary response to either estradiol benzoate (EB) (3 mug/100 g BW at 1300 h on Day 3) or LHRH (100 ng/rat at 1200 h on Day 4). In all experiments, trunk blood was taken at 1300 h on Day 4 to measure serum gonadotropin concentrations. In rats receiving an injection of RU486 on estrus, the absence of only the diestrous progesterone actions increased basal serum concentrations of LH and decreased those of FSH, and as in vehicle-treated controls, EB inhibited and LHRH stimulated LH secretion. In contrast, the absence of both proestrous afternoon and diestrous progesterone actions (as characterized rats treated with RU486 on proestrus) antagonized the inhibitory effect of EB and sensitized the pituitary to LHRH. These effects of RU486 on proestrus are ovary-dependent and eliminated by ovariectomy on metestrus. The increased ovarian secretion of testosterone and estradiol-17 beta during diestrus does not mediate the effects of proestrus-administered RU486 on pituitary function: no differences were found in the serum concentrations of estradiol-17 beta in diestrus between the groups of rats treated with RU486, and administration of the antiandrogen flutamide (2 mg/rat at 0900 h on Days 2 and 3) did not reverse the effects of RU486 on proestrus. In conclusion, the results suggest that in the absence of proestrous afternoon progesterone action, the ovaries of the 4-day-cyclic rat keep the pituitary gland in a state of low sensitivity to the inhibitory effects of estradiol and high sensitivity to the stimulatory effects of LHRH. Moreover, the results suggest that the putative ovarian factors involved are factors other than progesterone, androgens, or estradiol-17 beta.

Androgen Antagonists

Serotonin, catecholamines and metabolites in discrete brain areas in relation to lordotic responding on proestrus.

Levels of serotonin (5-HT) and metabolite, 5-hydroxyindole-acetic acid (5-HIAA), dopamine and metabolites, dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA), and norepinephrine (NE) were measured in microdissected brain areas at 14.30 and 19.30 h on diestrus and proestrus in rats. Lordosis, sexual behavior, was exhibited only by proestrous females at 19.30 h. The content of monoamines and/or metabolites changed from afternoon to early evening of diestrus in a number of brain regions. On proestrus, during the time when females became sexually receptive, additional changes appeared and many diestrous changes were reversed or amplified. Proestrus-specific changes were found in areas providing the descending circuit for regulation of lordosis, the medial preoptic nucleus (mPOA), ventromedial nucleus of the hypothalamus (VMN) and midbrain central gray (MCG), and in areas outside of endocrine control centers such as parietal cortex (Ctx) and dorsal raphe nucleus (DR). Thus, these data are consistent with previous studies showing functional changes in monoaminergic transmitters during the day/night cycle and during hormonal induction of neuroendocrine events. NE levels increased between 14.30 and 19.30 h on proestrus in most brain areas, ranging from 20% increases in mPOA and Ctx to a 220% increase in the DR. NE levels decreased on proestrus in the VMN. The direction of proestrous changes in NE were reversed in some, but not all of the areas, between the afternoon and evening of diestrus. Dopamine was detectable in all areas sampled, and the metabolites HVA and DOPAC were detectable in some but not all areas.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals

Alterations in steroid and gonadotropin release resulting from surgical stress during the morning of proestrus in 5-day cyclic rats.

The purpose of this experiment was to determine whether surgical stress on the morning of proestrus would elicit an early release of gonadotropin from the pituitary. Animals exhibiting 5-day estrous cycles underwent bilateral sham-ovariectomy under ether anesthesia at 0800 h of proestrus. These animals had high levels of progesterone and estradiol following the surgery. These steroids were thought to be adrenal in origin, since animals adrenalectomized at 0800 h of proestrus had low progesterone levels and estradiol comparable to unoperated controls. Subsequently, the sham-operated animals showed high FSH but not LH values at 1300 h, prior to the normal critical period for gonadotropin release. By 1400, the LH surge had begun, and progesterone was again being released. Adrenalectomized and unoperated controls showed no increase in any steroid or gonadotropin measured before 1400 h. These findings suggest that stress-induced release of adrenal estradiol and progesterone, rather than some other consequence of the surgical procedure, during the morning of proestrus, can advance the onset of release of FSH, prior to LH. Ovariectomy at 0800 h proestrus led to a rapid and dramatic increase in FSH but not LH secretion by 4 h after surgery. By 6 h after ovariectomy FSH had increased to six times control values and LH had increased to twice control values. Estradiol remained at control values for 6 h following surgery but 20alpha-hydroxypreg-4-en-3-one (20alpha-OHP) dropped quickly to baseline values. It is possible that a reduction in circulating 20 alpha-OHP may be responsible for the increases in FSH prior to LH in this group, but the absence of other negative feedback factors from the ovary or adrenal may also be involved.

20-alpha-Dihydroprogesterone

LTP varies across the estrous cycle: enhanced synaptic plasticity in proestrus rats.

Previous studies have shown that the number of dendritic spines and synapses in hippocampal CA1 stratum radiatum decreases more than 30% between the proestrus (high estrogen) and estrus (low estrogen) phases of the rat estrous cycle [10,27]. In the present study, we investigated whether hippocampal synaptic plasticity, as measured by long-term potentiation (LTP), might also vary across the estrous cycle of the female rat. Male rats, and female rats at each phase of the estrous cycle were tested in either the morning or afternoon. There were no significant group differences in the pre-LTP I/O curves. However, females examined during the afternoon of proestrus, the phase during which prior studies indicate synapse number to be highest, demonstrated the greatest degree of potentiation. Diestrus, proestrus and estrus females tested in the morning demonstrated similar amounts of potentiation. There were also significant differences in post-LTP I/O curves between the afternoon proestrus females and males tested in the afternoon. These results suggest that gonadal hormones, interacting with the time of day, may regulate neural processes underlying learning and memory.

Analysis of Variance

Changes in beta-endorphin content in discrete areas of the hypothalamus throughout proestrus and diestrus of the rat.

The aim of the present study is to investigate changes in beta-endorphin content in the hypothalamus during different stages of the estrous cycle. Groups of 9 to 10 Sprague-Dawley rats were sacrificed every two hours on proestrus from 8.00 to 18.00 h and groups of 7 to 8 rats were sacrificed on diestrus at 8.00, 12.00, 14.00 and 18.00 h. Preoptic suprachiasmatic region, posterior hypothalamus, arcuate nucleus and median eminence were dissected and assayed for beta-endorphin. A significant increase in beta-endorphin content was detected in the arcuate nucleus during proestrus (9.00 h: 1.76 +/- .31; 14.00 h: 4.10 +/- .85 microgram/g tissue wet weight). Levels did not change during diestrus (1.18 +/- .06 microgram/g). The increase caused significant differences in beta-endorphin values between both days at 12.00, 14.00 and 18.00 h, while the concentrations at 8.00 h were similar. The opposite pattern was observed in the median eminence with significantly higher proestrous beta-endorphin levels at 8.00 h (11.24 +/- 3.1 vs 3.52 +/- .64 microgram/g) and nonsignificant differences for the rest of the day. No significant change in beta-endorphin concentration was seen in the preoptic suprachiasmatic region over the day of proestrus (1.35 +/- .09 microgram/g). Diestrous beta-endorphin concentrations in this region were higher during the morning (2.60 +/- .65 microgram/g) and lower at 18.00 h (0.94 +/- .12 microgram/g) when compared to proestrous values. This pattern was caused by a 50% increase in beta-endorphin during the afternoon of diestrus. No changes were observed in the posterior hypothalamus on either day with comparable levels of beta-endorphin except at 18.00 h, when values were significantly higher on proestrus (1.66 +/- .30 vs 0.83 +/- .06 microgram/g).

Animals

Effect of calcitonin on the prolactin surge of proestrus.

The effect of calcitonin (CT) on the prolactin (PRL) surge of proestrus in rats was investigated under normal and perturbed lighting conditions. Salmon calcitonin (SCT) was injected i.p. on diestrus 2 or on proestrus, plasma PRL levels were measured by radioimmunoassay. SCT had no effect on the PRL surge under normal lighting conditions but it induced a small drop in PRL level measured on proestrus morning, 3 hours after CT injection. Animals submitted to perturbed light conditions had higher PRL levels than those kept under normal lighting. These data would indicate that for the female rats on proestrus the sensitivity to stress due to injection and blood sampling may be modulated by changing the photoperiod. SCT injection under these conditions may facilitate this destabilization in PRL level.

Animals

A relative depletion of luteinizing hormone-releasing hormone was observed in the median eminence of young but not middle-aged rats on the evening of proestrus.

Computer-assisted analysis was used to examine LHRH reaction product in the median eminence of young and middle-aged rats prior to and after the expected peak of the LH surge on proestrus. The area of LHRH reaction product was analyzed in 5 rostral-caudal levels (A-E) of the median eminence. The relative depletion of LHRH in the median eminence of young females on the evening compared to the afternoon of proestrus suggested LHRH neurosecretion in conjunction with the preovulatory LH surge. The pattern of depletion observed further suggested that LHRH release may occur preferentially from restricted regions of the median eminence or in a coordinated wave-like pattern. Four of the five levels of the median eminence exhibited a relative decrease in LHRH on the evening of proestrus in young females, and this time-related difference in LHRH reaction product was statistically significant in median eminence levels B and C. In contrast, little evidence of a relative depletion in LHRH reaction product from early to late proestrus was observed in the median eminence of aging animals. Moreover, the concentration of the densest LHRH reaction product appeared diminished in the median eminence of middle-aged compared to young females at the time points examined in the present study. The age-related differences observed in LHRH reaction product in the median eminence may contribute to the attenuated LH surge documented in middle-aged female rats. Whether these changes in LHRH immunoreactivity can be attributed to age-related alterations in afferents received by LHRH neuronal cell bodies or terminals or to intrinsic deficits in signalling mechanisms within LHRH neurons remains to be determined. Computer-assisted analysis of the immunocytochemical data enabled the assessment of relative changes in reaction product within specific elements of LHRH neurons in precise regions of the median eminence.

Aging

Cell proliferation in the dog (beagle) ovary during proestrus and early estrus.

Cell proliferation in the ovaries of 5 Beagle dogs was studied by autoradiography after pulse labelling with (3H)-thymidine during different periods of proestrus and early estrus. Indices of labelling and of necrosis were determined for different follicle types grouped according to their stages of atresia. Cell proliferation became most obvious during early proestrus in antral and in Graafian follicles, and during the periovulatory period in preantral follicles. By contrast, low labelling indices appeared in all follicle types during the middle of proestrus. All follicle types demonstrated a continuous decrease in labelling from early to late atresia with the exception of the granulosal layer of antral follicles during the periovulatory period. While many necrotic granulosal cells were found in antral follicles during the first 6 days of proestrus, there were few during the periovulatory period. Even Graafian follicles displayed necrotic granulosal cells now and then. It may be concluded that (1) intact ovarian follicles can show subtle changes indicating the beginning of atresia, and (2) during the proliferative phase of a long estrous cycle cell replication in the ovary is characterized by two bursts at the beginning and at the end of this period.

Animals