Ovarian steroid regulation of pulsatile luteinizing hormone release during early gestation in the rat.

Abstract The object of this study was to examine ovarian regulation of pulsatile luteinizing hormone (LH) secretion during early gestation. This was done primarily by analyzing pulsatile LH release in rats that were either sham ovariectomized (OVX) on Day 7 of pregnancy, implanted with empty Silastic capsules, and bled on Day 8, or OVX on Day 7, immediately implanted with Silastic capsules producing plasma levels of estradiol and/or progesterone characteristic of Day 7 to 8 of pregnancy, and bled on Day 8. In addition, the role of progesterone in regulating pulsatile LH secretion was also examined by administration of the progesterone receptor antagonist, RU486, on Day 7 and examining pulsatile LH release on Day 8 of pregnancy. OVX caused a marked increase in LH pulse amplitude and frequency within 24 h. Replacement with physiological plasma levels of estradiol or progesterone alone had no suppressive effect on this OVX-induced increase in pulsatile LH secretion. Restoration of physiological plasma levels of both estradiol and progesterone returned LH pulse amplitude to values seen in sham OVX controls, and prevented the OVX-induced increase in LH pulse frequency. The group mean LH pulse frequency tended to be less in estradiol + progesterone-treated rats than in sham OVX controls, but this difference was not statistically significant. RU486 blocked uterine progesterone receptors as evidenced by endometrial hemorrhaging. In agreement with the OVX + steroid replacement data, RU486 administration also resulted in increases in LH pulse amplitude and frequency. These data demonstrate that the frequency and amplitude of LH pulses on Day 8 of gestation are held in check by negative feedback signals coming from the ovary. Neither steroid alone exerts any suppressive influence over pulsatile LH secretion during early gestation, but both steroids acting together exert a prominent negative feedback regulation on the pulsatile LH release process.

Neurotransmitter involvement in naloxone-induced stimulation of pulsatile LH release on day 8 of pregnancy in the rat.

Naloxone, an endogenous opioid peptide (EOP) receptor antagonist, increases LH pulse frequency and amplitude in early gestation in the rat (7). The object of this study was to further explore the suppression of pulsatile LH release by EOPs on day 8 of pregnancy by examining whether inhibition of norepinephrine (NE) or epinephrine (EPIN) synthesis, or stimulation of gamma-aminobutyric acid (GABA)-B receptors, modified the ability of naloxone infusion (0.5 mg/kg/hr for 3.5 hr) to stimulate pulsatile LH secretion. Blood sampling (50 microliters whole blood/5 min) began 0.5 hr after the onset of infusion. Three studies were conducted. 1) LY 134046 (PNMT inhibitor, 50 mg/kg IP), given at -27, -20, and -3 hr relative to the onset of a 3-hr blood sampling period, produced no change in hypothalamic-preoptic area (HPOA) levels of NE, and a 76% decline in HPOA-EPIN levels, as determined by HPLC. Although basal LH pulse frequency was reduced, this treatment had no effect on the stimulatory action of naloxone on pulsatile LH release. 2) FLA-63 (DBH inhibitor, 25 mg/kg IP) given at -3 hr produced a 72% decline in HPOA-NE levels, a 44% decrease in HPOA-EPIN values, and blocked the stimulatory action of naloxone on LH pulse frequency. Since depletion of HPOA-EPIN by LY 134046 did not compromise the LH response to naloxone, this effect of FLA-63 is due to depletion of HPOA-NE levels. 3) While saline had no effect on the increased pulsatile LH release caused by naloxone, administration of baclofen (GABA-B receptor agonist, 6 mg/kg IV) suppressed the pulsatile LH secretory response to naloxone infusion.(ABSTRACT TRUNCATED AT 250 WORDS)

Ovarian regulation of pulsatile luteinizing hormone secretion during late gestation in the rat*.

Abstract The object of this study was to examine the influence of both estradiol (E(2)) and progesterone (P) alone or in combination on luteinizing hormone (LH) pulse amplitude and frequency during the interval between Days 21 and 22 of gestation. This was done by analyzing pulsatile LH release in rats bled on Days 21 and 22 of gestation, and in animals ovariectomized (OVX) on Day 21, implanted with silastic capsules producing plasma levels of E(2) and/or P characteristic of the Day 21 to 22 interval, and bled on Day 22 Pulsatile LH release increased between Days 21 and 22 due to an increase in pulse frequency and a small elevation in pulse amplitude. OVX produced no further increase in pulse frequency but markedly enhanced the small change in pulse amplitude. Preventing either the decline in plasma P that normally occurs between Days 21 and 22, or just the small additional decrease in plasma P levels produced by OVX, had no suppressive effect on pulse amplitude or frequency, although Day 22 levels of P alone augmented the normal increase in pulse frequency occurring between Days 21 and 22. Restoration of physiological plasma E(2) levels had no effect on the normal increase in pulse frequency, but partially attenuated the OVX-induced increase in pulse amplitude. Replacement of physiological Day 22 levels of both E(2) and P also decreased LH pulse amplitude, although amplitude was not significantly different from that seen following E(2) replacement alone, and was still greater than the normal Day 22 value. In contrast, restoration of physiological plasma levels of E(2)+ P caused a suppression of LH pulse frequency below that normally seen on Day 22. While E(2)+ P did not completely prevent the OVX-induced increase in pulse amplitude, administration of charcoal-extracted porcine follicular fluid to rats OVX on Day 21, and in which physiological plasma levels of E(2)+ P were restored, caused a further reduction in pulse amplitude. These data demonstrate that 1) marked increases in LH pulse amplitude are prevented from occurring between Days 21 and 22 of gestation by ovarian steroids, notably E(2), and that this suppression is enhanced by a non-steroidal factor present in porcine follicular fluid, 2) neither E(2) or P alone suppresses LH pulse frequency on Day 22 of gestation; LH pulse frequency increases on Day 22 because the plasma level of one of these steroids, P, markedly declines, and 3) restoration of physiological plasma levels of both steroids in the absence of the ovary produces an unphysiological suppression of pulse frequency, i.e. results in a lower pulse frequency than normally occurs in the presence of these same plasma steroid levels in animals with their ovaries intact. One hypothesis consistent with the latter observation is that at the end of gestation in the rat the ovary may produce a factor which 'protects' the frequency of the LH pulse generator from the negative feedback action of ovarian steroids. This allows an increase in LH pulse frequency and mean blood LH levels, and thereby facilitates ovarian follicular development and the normal progress of the first postpartum estrous cycle.

Adrenergic and noradrenergic regulation of pulsatile luteinizing hormone release.

Abstract The object of this study was to further define the roles of both norepinephrine (NE) and epinephrine (EPIN) in regulating pulsatile luteinizing hormone (LH) release in 4-day ovariectomized rats, in particular to examine the effect of decreasing NE synthesis on pulsatile LH secretion in animals with already greatly depleted levels of brain EPIN. Rats were injected ip with vehicle or drug at -27, -20, -5 and - 3 h relative to the onset of a 3-h blood sampling period. Hypothalamic-preoptic area (HPOA) levels of NE and EPIN were determined by high-performance liquid chromatography. Compared to controls, FLA-63 (25 mg/kg, a dopamine-ss- hydroxylase inhibitor), given at - 3 h, produced 50% and 22% declines in HPOA-NE and EPIN, respectively, and reductions in pulse amplitude and frequency. LY134046 (50 mg/kg, a phenylethanolamine N-methyltransferase inhibitor), given at - 27, - 20 and - 5 h, or -27, -20, -5 and -3 h, produced no change in NE, 88% and 86% declines in EPIN, respectively, and reductions in pulse frequency only. Each LY134046 treatment protocol produced the same decline in EPIN and pulse frequency. Thus, EPIN levels were maximally decreased by three LY134046 injections. When rats were given LY134046 at -27, -20 and -5 h, and FLA-63 at -3 h, compared to rats treated with LY134046 alone, there was no further decrease in HPOA-EPIN (82% decline), a 46% decline in NE, a further reduction in pulse frequency and a reduction in pulse amplitude. This further suppression of LH release must be due to a reduction in HPOA-NE levels since no further decrease in EPIN levels occurred. These data demonstrate within the same animal that NE and EPIN are both stimulatory to pulsatile LH release. NE stimulates the amplitude and frequency, and EPIN stimulates the frequency of pulsatile LH secretion.

Declining plasma progesterone levels eliminate endogenous opioid peptide suppression of LH pulse frequency on day 22 of gestation in the rat.

Endogenous opioid peptides (EOPs) suppress pulsatile LH release during pregnancy in the rat, but the stimulatory effect of the EOP receptor antagonist naloxone on LH pulse frequency is reduced or eliminated on day 22 of gestation. Plasma progesterone (P) levels are elevated through day 20 and fall by day 22. The aim of this study was to determine whether the decline in plasma P levels underlies the loss of EOP suppression of LH pulse frequency on day 22. Rats were bled on day 20 of pregnancy while being infused with 0.9% saline (0.5 ml/h) for 3 h, or implanted with empty or P-filled silastic capsules on day 20 and bled on day 22 while being infused first with saline for 3 h and then naloxone (0.5 mg/kg/h) for 3 h. Plasma P levels in the P-capsule group did not differ significantly from day 20 values, whereas P values in the empty capsule group were markedly decreased compared to day 20 levels and to values in the P-capsule group. Plasma estradiol values did not vary significantly between the two capsule-implanted groups. Mean blood LH levels increased between day 20 and day 22 due to an increase in LH pulse frequency and a small but significant increase in LH pulse amplitude. On day 22, mean blood LH levels, pulse amplitude and pulse frequency values during the saline infusion period in the P-capsule group were less than in the empty capsule group, and did not differ from values in the day 20 group.(ABSTRACT TRUNCATED AT 250 WORDS)

Analysis of estradiol-independent and -dependent endogenous opioid peptide suppression of pulsatile LH release between the mornings of diestrus 2 and proestrus in the rat estrous cycle.

The aim of this study was to analyze possible estradiol (E2)-independent and -dependent endogenous opioid peptide (EOP) suppression of pulsatile LH release between the mornings of diestrus 2 (D2) and proestrus by examining the LH response to naloxone infusions in the presence or absence of proestrous levels of E2. Pulsatile LH secretion remained unchanged between D2 and proestrus but mean blood LH levels, pulse amplitude and frequency increased within 24 hr following ovariectomy on D2. This increase was due in large part to the loss of E2 negative feedback, since restoration of physiological proestrous E2 levels returned LH pulse frequency to proestrous a.m. levels and greatly reduced pulse amplitude. In ovariectomized rats lacking E2 negative feedback, continuous infusion of the EOP receptor antagonist naloxone (0.5 and 2 mg/kg/hr) caused a further increase in pulse amplitude and frequency. This naloxone-induced increment in pulsatile LH release was exerted via centrally located EOP receptors since naloxone did not alter pituitary responsiveness to LHRH, and its stimulatory action on pulsatile release was diminished by simultaneous infusion with morphine. Naloxone also increased pulsatile LH release in E2-treated animals. The naloxone-induced increments in LH pulse amplitude were the same in the presence or absence of E2 negative feedback. Moreover, the increments in amplitude produced by naloxone in E2-treated rats were significantly less than those resulting from the combination of ovariectomy plus naloxone infusion in empty capsule-implanted rats. These data indicated that naloxone infusion in E2-implanted animals blocked an E2-independent EOP suppression of this parameter of pulsatile release.(ABSTRACT TRUNCATED AT 250 WORDS)

Influence of photoperiod and gonadal steroids on hibernation in the European hamster.

Torpor was monitored daily in adult male and female European hamsters (Cricetus cricetus) induced to hibernate by exposure to a cold environment (6 degrees C). The effect of photoperiodic manipulations or administration of exogenous gonadal steroids was examined in gonadectomized or intact hamsters. 1. Gonadal regression occurred in all short day, but only in some long day, cold-exposed hamsters. Entry into hibernation was not observed until reproductive regression had occurred. Thus, gonadal atrophy appears to be a necessary precondition for hibernation. 2. Castrated hamsters in the short day cold condition showed a significantly greater incidence of torpor than those in the long day cold condition. Hence, photoperiod affected torpor independently of its effect on the gonadal cycle. 3. Testosterone, when administered via silastic capsules at near physiological levels, completely inhibited torpor in gonadectomized male and female hamsters hibernating in the short day cold condition. 4. In ovariectomized females, torpor was unaffected by progesterone treatment, but partially inhibited by estradiol. A greater inhibition of torpor was observed when estradiol-primed females were administered both estradiol and progesterone simultaneously. Thus, the effect of both hormones may be functionally comparable to that of the single testicular hormone. 5. Estradiol inhibited torpor to a greater extent in intact and ovariectomized female hamsters hibernating in long days than those in short days, suggesting an effect of photoperiod on responsiveness to estradiol. These results indicate an inverse relationship between the gonadal and hibernation cycles, and a probable role for gonadal steroids to influence the timing of the hibernation season. However, non-gonadal factors must also be involved in controlling hibernation, since photoperiod affected the incidence of torpor in gonadectomized animals and because hamsters were able to terminate hibernation in the absence of gonadal hormones.

Absence of steroid-dependent, endogenous opioid peptide suppression of pulsatile luteinizing hormone release between diestrus 1 and diestrus 2 in the rat estrous cycle.

The objective of this study was to determine whether the negative feedback action of ovarian steroids on pulsatile luteinizing hormone (LH) release in the diestrous 1 (D1)-diestrous 2 (D2) interval of the rat estrous cycle is mediated by endogenous opioid peptides (EOPs), by examining the pulsatile LH release response to naloxone infusions in the presence or absence of D1-D2 levels of estradiol (E2) and progesterone (P). As plasma E2 and P levels increased between D1 and D2, mean blood LH levels decreased due solely to a decrease in LH pulse amplitude as frequency remained stable. However, ovariectomy increased both parameters of pulsatile LH release, indicating the effect of loss of ovarian steroid-negative feedback in this interval. Replacement of D1-D2 plasma levels of E2 and P restored D2 values for both parameters of pulsatile LH release, and E2 + P did not alter in vivo pituitary responsiveness to LH-releasing hormone (LHRH). In ovariectomized rats lacking the negative feedback provided by E2 + P in this cycle interval, continuous infusion of naloxone caused a further dose-dependent augmentation in both LH pulse amplitude and frequency. This stimulatory action of naloxone was prevented by simultaneous infusion with morphine, and was not associated with any change in in vivo pituitary responsiveness to LHRH, indicating that this was an action exerted through centrally located EOP receptors. Naloxone also increased both parameters of pulsatile LH release in E2 + P-treated rats. However, the magnitudes of the naloxone-induced increments in LH pulse amplitude and frequency in ovariectomized, steroid-treated rats were not greater than those seen in ovariectomized, nonsteroid-treated rats given naloxone versus saline. In addition, mean values for both parameters of pulsatile LH secretion during EOP receptor blockade in steroid-treated rats were reduced when compared to values in ovariectomized, nonsteroid-treated rats infused with naloxone. Thus the stimulatory effect of naloxone on pulsatile LH release was similar in the presence or absence of the negative feedback action of D1-D2 plasma levels of E2 + P. This indicates that the negative feedback effect of E2 + P on pulsatile LH release in this interval is not mediated by EOPs whose actions are blocked by naloxone.

Endogenous opioid peptide regulation of pulsatile luteinizing hormone secretion during pregnancy in the rat.

The objective of this study was to determine if endogenous opioid peptides (EOPs) influence the pattern of pulsatile luteinizing hormone (LH) secretion on days 6-8, 14-16 and 22 of gestation in the rat. Unanesthetized animals with two jugular cannulae were initially infused with 0.9% saline during which the control pattern of pulsatile LH release was determined. Possible EOP involvement was then determined by infusion of the EOP receptor antagonist naloxone. Plasma estradiol (E2) and progesterone (P) values increased between days 6-8 and 14-16. While plasma E2 values remained elevated through day 22, plasma P values declined by 90%. As previously reported, mean blood LH levels during the control period on day 22 were higher than on days 6-8 and 14-16 due to an increase in LH pulse frequency. At each stage of gestation naloxone infusion increased mean blood LH levels. This stimulatory action of naloxone was reduced in a dose-dependent fashion by simultaneous infusion with morphine, demonstrating that this effect is mediated via EOP receptors. There was no difference in the in vivo pituitary responsiveness to LH-releasing hormone (LHRH) between rats infused with saline or naloxone at any stage of pregnancy, demonstrating that the stimulatory effect of naloxone was not exerted at the pituitary level. Naloxone increased both the amplitude and frequency of pulsatile LH secretion on days 6-8, and stimulated frequency on days 14-16. The effect on amplitude could not be assessed on days 14-16 because too few rats exhibited pulsatile LH secretion prior to naloxone infusion. The increase in pulse frequency was similar on days 6-8 and 14-16. Although naloxone increased LH pulse amplitude and frequency on day 22, these increases were significantly less than those seen on days 6-8 and 14-16, respectively. Pituitary responsiveness to LHRH was less at all stages of pregnancy in comparison to responsiveness in ovariectomized rats, and progressively declined from days 6-8 through day 22. The lowest responsiveness to LHRH was seen on day 22 and contributed, at least in part, to the diminished increase in LH pulse amplitude in response to naloxone infusion on day 22 compared to days 6-8. The reduced naloxone-induced increment in LH pulse frequency on day 22, occurring coincident with a precipitous decline in plasma P levels, suggests a decreased EOP suppression of pulse frequency at this time.(ABSTRACT TRUNCATED AT 400 WORDS)

Steroid-independent endogenous opioid peptide suppression of pulsatile luteinizing hormone release between estrus and diestrus in the rat estrous cycle.

We have previously demonstrated an absence of ovarian steroid negative feedback on pulsatile luteinizing hormone (LH) release between estrus and early diestrus 1 (D1) in the rat estrous cycle. The object of the present study was to determine if there was a steroid-independent endogenous opioid peptide (EOP) suppression of pulsatile LH release in this same 24-h interval, and if so, which parameter(s) of pulsatile LH release were affected. Rats were bled on estrus, or 24 h following sham ovariectomy (OVX), or OVX at 08.30-10.00 h on estrus. At the time of bleeding all rats were infused i.v. for 4 h either with 0.9% saline (0.5 ml/h) or naloxone (0.005, 0.05, 0.5, or 2 mg/kg/h). At 1 h after the infusion began, rats were bled for 3 h (40 or 50 microliters whole blood/5 min) between 09.30 and 12.30 h. Mean blood LH levels increased between estrus and early D1 due to increases in LH pulse amplitude and frequency. OVX on estrus decreased plasma levels of estradiol and progesterone 24 h later, but did not augment the increase in pulsatile LH release. However, naloxone infusion augmented the increase in pulsatile LH secretion in sham ovariectomized rats in a dose-dependent fashion. While infusion of 0.005 or 0.05 mg/kg/h had no effect, 0.5 or 2 mg/kg/h increased blood LH levels by increasing both LH pulse amplitude and frequency. The stimulatory effect of naloxone on pulsatile LH release was blocked by simultaneous infusion of morphine, demonstrating that the effect was mediated by EOP receptors.(ABSTRACT TRUNCATED AT 250 WORDS)

The relationship between declining plasma progesterone levels and increasing luteinizing hormone pulse frequency in late gestation in the rat.

The object of this study was to determine whether the increase in LH pulse frequency and mean blood LH levels on day 22 of pregnancy in the rat is due to the precipitous fall in plasma progesterone (P) levels that occurs late in gestation. On day 20 of pregnancy two groups of animals with indwelling jugular cannulae were implanted sc with empty or P-containing Silastic capsules. Blood samples were withdrawn 0.5 h before and 5.5 h postimplantation on day 20 (0800 and 1400 h), at 1400 h on day 21, and at the end of the study between 1200-1300 h on day 22 to follow the time course of changes in plasma P levels over this 2-day period in both groups. These groups were bled on day 22 for 3 h between 0900-1200 h for analysis of pulsatile LH release. A third group not implanted with Silastic capsules was bled on day 20 for 3 h; plasma P levels in these rats bled on day 20 did not differ from the preimplantation values observed in either group of capsule-implanted rats. In empty capsule-implanted animals, plasma P values declined slightly from days 20 to 21 and were dramatically reduced between days 21 and 22. In contrast, after implantation of P capsules, plasma P levels were elevated on day 20 and remained elevated on day 21 compared with preimplantation values. Although these increased plasma P values declined between days 21 and 22, reflecting a decrease in endogenous P secretion, they were nonetheless comparable to day 20 values due to the presence of the P-containing capsules. Plasma estradiol values did not differ significantly between any of the experimental groups. In the empty capsule group bled on day 22, mean blood LH levels and LH pulse frequency were significantly higher compared to day 20 values, at a time when plasma P levels had fallen significantly from day 20 values. However, in the P capsule group, mean blood LH levels and LH pulse frequency on day 22 were significantly lower than values in the empty capsule group and were not different from the low values on day 20. Thus, preventing a decline in plasma P values to the low levels normally found on day 22 prevented the increase in LH pulse frequency and mean blood LH levels normally seen at this time of pregnancy.(ABSTRACT TRUNCATED AT 250 WORDS)

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)

Influence of estradiol and progesterone on pulsatile LH secretion in 8-day ovariectomized rats.

Previous studies in our laboratory [Endocrinology 114: 1605-1612 (1984); Neuroendocrinology 41: 252-257 (1985)] examined the influence of ovarian steroids on pulsatile luteinizing hormone (LH) release, and involved immediate replacement following ovariectomy (OVX) of estradiol (E2) and progesterone (P) for a 24-hour period within the physiological context of the estrous cycle. The present study investigated the effects of replacing E2 and/or P 1 week after OVX, and therefore examined whether the time elapsed following OVX influences the effects of ovarian steroids on pulsatile LH release. Immediately after jugular venous cannulation, rats were implanted with either empty silastic capsules or capsules capable of restoring physiological levels of E2 and/or P comparable to those found in intact rats between the intervals of diestrus 1 (D1) and diestrus 2 (D2), or D2 and proestrous morning. 24 h later, these rats were bled continuously at a rate of 50 microliters whole blood/6 min for 3 h for analysis of pulsatile LH secretion. Rats with empty capsules had decreased levels of E2 and P and elevated mean blood LH levels, pulse amplitudes and frequencies. Two groups of animals with E2 capsules had plasma E2 levels comparable to those seen either in the D1-D2 or D2-proestrous intervals, decreased levels of P, and in both cases significant decreases in LH pulse amplitude, but no change in LH pulse frequency or basal LH secretion. Since mean blood LH levels in 8-day ovariectomized rats are determined by LH pulse amplitude, frequency and basal LH secretion [Neuroendocrinology 37: 421-426 (1983)].(ABSTRACT TRUNCATED AT 250 WORDS)

Interaction between estradiol and a nonsteroidal factor in porcine follicular fluid in regulating LH pulse amplitude between the mornings of diestrus 2 and proestrus in the rat.

The object of this study was to examine the effect of porcine follicular fluid (PFF) alone or in combination with estradiol (E2) on pulsatile LH release during the interval between the mornings of diestrus 2 (D2) and proestrus in the rat. Steroids were removed from PFF by charcoal extraction. Preliminary studies indicated that 1 ml PFF given intraperitoneally suppressed FSH secretion for up to 15 h, with an onset of action between 3 and 4 h and maximal suppression between 6 and 9 h. In subsequent experiments, six groups of animals were bled continuously for 3 h between 07.30 and 10.30 h at a rate of 50 microliter whole blood/5 min: group 1 was bled on D2; group 2 was sham ovariectomized on D2 (08.30-09.30 h), immediately implanted with an empty capsule, given saline at 12.00 and 24.00 h, and bled on proestrous AM; groups 3-6 were ovariectomized on D2, implanted with an empty or E2 capsule, given 1 ml saline or PFF at 12.00 and 24.00 h, and bled 24 h following ovariectomy (OVX). Between D2 and proestrus plasma E2 levels increased, and there was no change in any parameter of pulsatile LH release. However, OVX on D2 reduced plasma E2 levels and increased mean blood LH levels above proestrous values due to increases in LH pulse amplitude and frequency. Restoration of physiological proestrous levels of E2 reduced the increase in mean blood LH levels, by lowering pulse frequency to proestrous values and by greatly reducing pulse amplitude. However, LH pulse amplitude and mean blood LH levels were still higher than values on proestrus. PFF alone produced no alteration in any parameter of pulsatile LH release compared with saline-treated animals.(ABSTRACT TRUNCATED AT 250 WORDS)

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)

Effect of alterations in pulsatile luteinizing hormone release on ovarian follicular atresia and steroid secretion on diestrus 1 in the rat estrous cycle.

This study examined the importance of pulsatile luteinizing hormone (LH) release on diestrus 1 (D1; metestrus) in the rat estrous cycle to ovarian follicular development and estradiol (E2) secretion. Single injections of a luteinizing hormone-releasing hormone (LHRH) antagonist given at -7.5 h prior to the onset of a 3-h blood sampling period on D1 reduced mean blood LH levels by decreasing LH pulse amplitude, while frequency was not altered. Sequential injections at -7.5 and -3.5 h completely eliminated pulsatile LH secretion. Neither treatment altered the total number of follicles/ovary greater than 150 mu in diameter, the number of follicles in any size group between 150 and 551 mu, or plasma E2, progesterone, or follicle-stimulating hormone (FSH) levels. However, both treatments with LHRH antagonist significantly increased the percentage of atretic follicles in the ovary. These data indicate that: 1) pulsatile LH release is an important factor in determining the rate at which follicles undergo atresia on D1; 2) reductions in LH pulse amplitude alone are sufficient to increase the rate of follicular atresia on D1; 3) an absence of pulsatile LH release for a period of up to 10 h on D1 is not sufficient to produce a decline in ovarian E2 secretion, most likely because the atretic process was in its early stages and had not yet affected a sufficient number of E2-secreting granulosa cells to reduce the follicle's capacity to secrete E2; and 4) suppression or elimination of pulsatile LH release on D1 is not associated with diminished FSH secretion.

Pulsatile luteinizing hormone release during pregnancy in the rat.

The present studies were designed to characterize LH release during pregnancy in the rat. Unanesthetized animals with jugular cannulae were bled for 3 h between 1000-1300 h on days 6-8, 14-16 or 22 of gestation (50 microliters whole blood/5 min). Plasma estradiol and progesterone values both increased from days 6-8 to days 14-16. However, while plasma estradiol levels increased further between days 14-16 and day 22, plasma P levels had declined 86%. The percent coefficients of variation obtained for alterations in blood LH levels at each stage of pregnancy were all significantly greater than intraassay variation, indicating that LH release was pulsatile at each stage. Although there were no significant differences in mean blood LH levels, pulse amplitude, or frequency between days 6-8 and 14-16, the individual patterns of LH release clearly varied between these 2 groups, and most notably within the 14-16 day group. Fifty-three percent (9 of 17) of the LH records in rats on days 14-16 were nonpulsatile compared to only 20% (3 of 15) on days 6-8. However, despite a trend toward an absence of pulsatile LH release on days 14-16, mean frequency at this time did not differ from days 6-8, since on days 14-16 the remaining 8 animals demonstrated 3.5 pulses/3 h, while on days 6-8 the other 12 rats averaged only 2.5 pulses/3 h. On day 22, there was a marked increase in mean blood LH levels compared with either days 6-8 or 14-16. This increase was due to an increase in mean LH pulse frequency. All 15 rats demonstrated pulsatile LH secretion, a significantly greater incidence of pulsatile LH release than on days 14-16 (100% vs. 47%). These data demonstrate that LH release is pulsatile during pregnancy in the rat, and changes in the characteristics of this secretion occur at different stages of gestation.

Lack of ovarian steroid negative feedback on pulsatile luteinizing hormone release between estrus and diestrous day 1 in the rat estrous cycle.

The object of this study was to determine the roles of ovarian estradiol (E2) and progesterone (P) in regulating pulsatile LH release between estrus and diestrous day 1 (D1) in the rat estrous cycle. Three groups of rats were bled at rates of 75 or 100 microliter whole blood/6 or 8 min, respectively, between 0930-1230 h on estrus or 24 h after either sham ovariectomy (OVX) on estrus (i.e. on D1) or OVX on estrus. There were no differences in plasma E2 and P levels in rats between estrus and early D1. However, after OVX on estrus, plasma levels of both steroids declined and were significantly lower 24 h later than values in D1 controls, indicating an active ovarian secretion of both hormones in this interval of the rat cycle. A significant increase in mean blood LH levels occurred between estrus and D1 due to an increase in LH pulse amplitude and frequency. After OVX on estrus, all parameters of pulsatile LH release also increased within 24 h, but mean blood LH levels as well as LH pulse amplitude and frequency were virtually identical to values in D1 controls, despite the decline in plasma E2 and P levels. Thus, OVX did not augment the increases in LH pulse amplitude and frequency that occur between estrus and D1. This demonstrates that the increase in pulsatile LH release from estrus to early D1 occurs in the absence of ovarian steroid negative feedback; the increases in LH pulse amplitude and frequency are not under negative feedback control by the low plasma levels of E2 and P present at this time. These data stand in direct contrast to the presence of prominent ovarian steroid negative feedback systems operative between D1 and diestrous day 2, and diestrous day 2 and proestrus.

Presence of a neurophysin-like precursor in the green turtle (Chelonia mydas).

A glycoprotein of neurohypophysial origin was found to have cofractionated with FSH prepared from pituitary glands of the green turtle, Chelonia mydas. Antiserum raised against this preparation contained high antibody titres and affinity for the neurohypophysial component and allowed development of a specific radioimmunoassay to monitor its purification and distribution in the brain. Immunocytochemistry revealed that the glycoprotein was concentrated in the pars nervosa and associated nerve tracts passing through the median eminence to the supraoptic and paraventricular nuclei; similar distributions were observed in turtles and rats. The antiserum to the turtle material bound radiolabelled rat vasopressin (VP)-neurophysin and precipitated precursors of this neurophysin, but it did not cross-react with rat oxytocin-neurophysin. An amino-terminal alanine was also consistent with the structure of rat VP-neurophysin, but the turtle molecule was larger than the corresponding rat molecule. Limited tryptic digests of the turtle glycoprotein contained two components, one of which bound to lysine VP. Both components contained carbohydrate, but only the one which bound to VP cross-reacted in a radioimmunoassay for rat VP-neurophysin. The apparent surge in plasma immuno-FSH at the time of oviposition previously described in the turtle probably represented release of a neurophysin-like 'carrier' molecule associated with secretion of the neurohypophysial hormone (e.g. arginine vasotocin; AVT) responsible for oviduct contractility. These data suggest that the neurohypophysial glycoprotein represents a partially processed AVT precursor and provide the first biochemical evidence of a mammalian-like biosynthetic pathway for neurohypophysial hormones in a non-mammalian species.