Immunocytochemical and physiological evidence of a synapse between dopamine- and luteinizing hormone releasing hormone-containing neurons in the ewe median eminence.

Immunocytochemical labeling revealed that the arcuate nucleus (ARN) of the ewe's hypothalamus contains numerous tyrosine hydroxylase (TH)-positive neurons, that appear to lack dopamine-beta-hydroxylase (DBH)-like immunoreactivity. Axons of these presumed dopaminergic neurons converge in the median eminence (ME) with Luteinizing Hormone Releasing Hormone (LHRH)-containing axons originating mostly from neurons situated in the medial preoptic area. Electron microscopic double labeling revealed synaptic contacts between TH-positive presynaptic profiles and LHRH-containing postsynaptic elements. Samples of ME, ARN, paraarcuate and lateral hypothalamus were dissected and incubated to assess LHRH release and tissue content. Only ME-LHRH release was significantly reduced in the presence of dopamine (DA). All other regions released equal amounts with and without DA. Thus, a presynaptic dopaminergic inhibition of LHRH-containing axons at the level of the ME might contribute to the regulation of LHRH release into the portal vessels.

Hyperprolactinemia-induced precocious puberty in the female rat: ovarian site of action.

Hyperprolactinemia induced in immature female rats by chronic treatment with sulpiride, a dopaminergic receptor blocker, resulted in advancement of the onset of puberty. While serum PRL levels increased promptly after initiation of the treatment and remained elevated throughout the entire period studied (day 22 to first diestrus after vaginal opening), gonadotropin levels did not appear to be altered and increased abruptly only at the time of the first preovulatory surge. Except for a decrease during the first diestrus, serum TSH and GH were not consistently altered by sulpiride. After 5 days of treatment, serum progesterone levels, but not those of androgens, were increased in hyperprolactinemic (HP) rats. Uterine weight, taken as an index of estrogen secretion, was unambiguously increased in HP rats, the first significant difference being observed 5 days after initiation of the sulpiride treatment (day 27). Ovarian estrogen and progesterone responsiveness of normal animals to gonadotropins, as determined by in vitro release of the steroids after incubation with hCG, increased with age. The response was dramatically enhanced in HP rats, its magnitude increasing markedly as the animals approached puberty. Androgen response to hCG was, however, similar in HP and control rats. In vitro release of adrenal progesterone, but not that of estrogen or androgens, was slightly enhanced in HP rats. When sulpiride treatment was terminated on the first day of diestrus after vaginal opening, the animals continued to show estrous cycles. By contrast, maintenance of the treatment for several days after vaginal opening resulted in a condition of constant diestrus. The results suggest that one of the mechanisms by which PRL induces precocious puberty in the female rat is by sensitizing the ovaries to the low circulating gonadotropin levels observed during the prepubertal period. In addition, PRL also seems to stimulate progesterone secretion by the adrenal glands.

Distribution of luteinizing hormone-releasing hormone (LHRH) content and total LHRH-degrading activity (LHRH-DA) in the hypothalamus of the ewe.

Hypothalamic enzymatic activities capable of degrading LHRH may play a physiological role in the neuroendocrine control of LHRH. There is increasing evidence, however, that these enzymes are not peptide specific. The present study in the ewe analyzed the possibility that the specificity of LHRH-degrading activity (LHRH-DA) could be conferred by the relative location of LHRH-DA with respect to that of the LHRH peptide itself. LHRH content was correlated with LHRH-DA in discrete hypothalamic samples containing LHRH-positive cell bodies and axons and in immediately adjacent areas apparently devoid of LHRH immunoreactivity. LHRH content was assessed by RIA, and LHRH-DA was determined by HPLC of the LHRH decapeptide and its degradation fragments. The sampling of discrete hypothalamic areas was designed after immunocytochemical localization of LHRH. LHRH-containing cell bodies were observed in the medial preoptic area, projecting LHRH-positive fibers to the infundibular region. At all hypothalamic levels, there was a tight correlation (r greater than 0.95; P less than 0.01) among the regional distribution of LHRH-DA, LHRH content, and the presence of LHRH-like immunoreactivity. LHRH-DA, in addition, was present in areas of low (e.g. lateral hypothalamus) or undetectable (e.g. cerebral cortex) LHRH content that were devoid of LHRH-like immunoreactivity. The appearance of LHRH degradation fragments suggests that the initial cleavage of LHRH by LHRH-DA occurs at the Tyr5-Gly6 bond at all hypothalamic levels studied. These findings indicate that part of the total hypothalamic LHRH-DA may be located within the LHRH hypophysiotropic pathway. This suggests an anatomical locus for a possible physiological interaction between LHRH and LHRH-DA.

Blockade of steroid-induced leuteinizing hormone release by selective depletion of anterior hypothalamic norepinephrine activity.

Female Sprague-Dawley rats, weighing 200--225 g, were ovariectomized and, 10 days later, were given a single sc injection of 10 micrograms estradiol benzoate/100 g BW. Three days after estradiol benzoate treatment, animals received 50 micrograms progesterone (P)/100 g BW, resulting in a surge in LH release 7 and 9 h later. To determine the locus of the noradrenergic component of the P-induced LH surge, 6-hydroxydopamine (6-OH-DA), a neutrotoxin, was implanted into either the suprachiasmatic region or the median eminence (ME) 24 h before P administration. An implant of 6-OH-DA in the suprachiasmatic region decreased anterior hypothalamic norepinephrine concentration by 83%, anterior hypothalamic dopamine concentration by 24%, and eliminated the P-induced LH surge. ME implant of 6-OH-DA decreased norepinephrine concentration by 57% without affecting dopamine concentration, but was unable to alter the P. induced LH surge. These results indicate that the anterior hypothalamic noradrenergic system is necessary for the P-induced LH surge in ovariectomized rats and that noradrenergic nerve terminals in the ME are not involved in this process.

Evidence that catecholaminergic and peptidergic (luteinizing hormone-releasing hormone) neurons in suprachiasmatic-medial preoptic, medial basal hypothalamus and median eminence are involved in estrogen-negative feedback.

A time-course study was performed to correlate serum gonadotropin changes after ovariectomy (OVX) or OVX plus acute estradiol (E2) treatment with LHRH content and norepinephrine (NE) and dopamine (DA) content and turnover in the suprachiasmatic-medial preoptic region (SCH-PO), the medial basal hypothalamus (MBH), and the median eminence (ME). Rats on diestrous day 1 (D1) were sham operated to serve as controls or were ovariectomized and killed 1, 3, 10, 30, or 60 days after OVX. Catecholamines (CA) were measured by a radioenzymatic isotopic assay. LHRH, LH, and FSH were measured by RIA. CA turnover estimates were obtained after blocking CA synthesis with alpha-methyl-p-tyrosine. Twenty-four hours after OVX, serum FSH rose 3-fold while LH remained unchanged. An abrupt rise in the LHRH content of the ME and MBH (P<0.05) was found, while the LHRH content of the SCH-PO was unaltered. The content of DA in the ME and MBH showed a significant rise at 24 h. A significant increase in DA turnover in the ME was also observed at this time. LH rose slightly 3 days after OVX and increased 7-fold by day 10. The LHRH content of the SCH-PO started to rise by day 3 and reached peak levels by day 10, while in the ME and MBH it declined by day 3 and was significantly lower than D1 levels 10 days after OVX. The DA content of the ME and MBH and the NE content of the ME, MBH, and SCH-PO increased significantly by day 3. NE turnover in the SCH-PO increased significantly by day 3 and stayed elevated at day 10. A similar increment in NE turnover in the ME was observed at day 10. By 30 and 60 days after OVX, serum LH and FSH plateaued, LHRH in the ME and MBH remained significantly lower, SCH-PO LHRH declined to D1 values, and CAs in all areas returned to control levels. The acute negative feedback effect of E2 was evaluated 10 days after OVX by injecting 20 microg E2 benzoate 3 h before sacrifice. After E2 treatment, LH was lowered by 75%, but FSH was unchanged. A significant reduction to D1 levels of the contents of both LHRH and NE in the SCH-PO and a significant decrease in NE turnover in the SCH-PO and a significant decrease in NE turnover in the SCH-PO and ME were observed. LHRH levels in other regions were not significantly altered, although levels were already low before estrogen. Surprisingly, DA and NE turnover in the MBH, which had not changed after OVX, were significantly increased after E2 treatment. DA turnover in the SCH-PO was decreased after E2. These results show a temporal relationship between gonadotropin release and LHRH and CA content and turnover in different brain areas, and indicate that the acute negative feedback effect of E2 is mediated, at least in part, through changes in LHRH and NE metabolism in the SCH-PO and ME regions. These results also stress the necessity of studying smaller hypothalamic regions, since changes in the opposite direction in neighboring areas were observed after either OVX or E2 benzoate treatment...

Evidence that endopeptidase-catalyzed luteinizing hormone releasing hormone cleavage contributes to the regulation of median eminence LHRH levels during positive steroid feedback.

In previous studies we provided evidence that the degradation of LHRH is regulated so as to contribute to the establishment of appropriate levels of the decapeptide during the events leading to gonadotropin secretion in the first estrous cycle at puberty in the rat. In the present report, we present the first evidence that this apparent regulation of LHRH degradation can be studied in an experimental positive feedback model. We show that LHRH degradation in the median eminence was decreased 3 h after progesterone administration, at a time when LHRH content in this region is increasing, and when serum levels of LH remained at basal levels. Six h after progesterone administration, at the time of the LH surge, median eminence LHRH degradation was still low and LHRH content had fallen to basal levels. Additionally, we exploited this model to examine the mechanism of peptidase activity change by showing that blockade of noradrenergic neurotransmission by diethyldithiocarbamate abolishes the inhibition of LHRH degradation observed prior to the secretion of LH. We conclude that the degradation of LHRH by an endopeptidase may contribute to the regulation of LHRH levels appropriate for gonadotropin release, and that this can be studied in the ovariectomized, estrogen-progesterone-treated rat.

Hyperprolactinemia-induced precocious puberty: studies on the mechanism(s) by which prolactin enhances ovarian progesterone responsiveness to gonadotropins in prepubertal rats.

Hyperprolactinemia induced in immature female rats by treatment with sulpiride, a dopaminergic receptor blocker, increased the in vitro release of ovarian progesterone (P) in response to different doses of both highly purified hCG and human FSH. The increased P response to gonadotropins was also observed in ovaries of animals injected with ovine PRL or in rats in which the hyperprolactinemic condition was induced by pimozide, a more typical dopaminergic receptor blocker. In addition, the pimozide treatment advanced the time of puberty in a manner similar to that previously observed with sulpiride. In other experiments in which only the ovarian response to hCG, but not that to FSH, was evaluated, it was found that the in vivo treatment of hypophysectomized immature rats with sulpiride did not modify the almost undetectable serum PRL levels of these animals and failed to increase the in vitro ovarian P response to hCG. By contrast, sc injection of PRL to hypophysectomized rats clearly enhanced the in vitro release of P in response to the gonadotropin. Adrenalectomy of otherwise intact rats significantly decreased the in vitro ovarian P response to hCG and blunted the increase in the P response induced by hyperprolactinemia. These effects, however, were almost completely reversed by concomitant corticosterone replacement therapy. The ovarian content of hCG receptor in normal rats was found to increase during juvenile development (days 22-31). Hyperprolactinemic animals showed a greater ovarian hCG receptor content than age-matched 31-day-old controls. In contrast, the FSH receptor contents were similar in both groups. The increase in hCG receptor content induced by hyperprolactinemia was even more clearly manifested in isolated granulosa cells, but, as in the case of the whole ovaries, the FSH receptor content in these cells remained essentially the same in hyperprolactinemic and control rats. The results indicate that the enhanced ovarian P response to hCG induced by PRL in prepubertal rats is, at least in part, mediated by an increase in the LH receptor content of the granulosa cells of the developing follicle. It also appears that the sensitizing effect of PRL on the prepubertal ovarian P response to gonadotropins is modulated by the adrenal gland through an effect exerted by corticosterone. The mechanisms by which PRL enhances the ovarian P response to FSH do not appear to involve changes in FSH receptors. However, the occurrence of enlarged uteri in hyperprolactinemic rats suggest that the PRL-induced increase in the P response to FSH may be related to the presence of more mature, estrogen-secreting follicles resulting from the hyperprolactinemic condition.

In vivo biosynthesis of hypothalamic luteinizing hormone releasing hormone in individual free-running female rats.

The present studies examined the feasibility of performing in vivo biosynthetic studies of hypothalamic LHRH in preoptic area cannulated individual ovariectomized, estradiol benzoate (EB)-progesterone (P) treated rats. One day after EB treatment, rats were bilaterally cannulated in the anterior ventromedial preoptic area (POA). The following day, rats were treated with P, and a mix of 3H-proline and 3H-leucine (500 muCi) was continuously infused (for 8 hr) into the POA using an Alzet osmotic minipump delivery system at a rate of 1 microliter/h/cannula. 3H-LHRH was acid-extracted from discrete hypothalamic regions (POA, medial basal hypothalamus [MBH]and median eminence [ME]) and was purified to constant specific activity by sequential High Performance Liquid Chromatography. 3H-LHRH accounted for 0.010%, 0.018% and 1.6% of the total tissue 3H recovered in the POA, MBH and ME, respectively. These results demonstrate the feasibility of performing physiological studies on the biosynthesis and transport of hypothalamic LHRH using POA cannulated free-running female rats.

The maturation of estradiol-negative feedback in female rats: evidence that the resetting of the hypothalamic "gonadostat" does not precede the first preovulatory surge of gonadotropins.

Several experiments were performed to study the changes in the negative feedback of estradiol on gonadotropin secretion around the time of puberty in the female rat. Ovariectomy of juvenile, first diestrus, or adult animals elevated FSH and LH levels 2 and/or 4 days later. Estradiol administered via Silastic capsules, at several dose levels, was much more effective in preventing the postcastration rise of gonadotropins in juvenile than in the older animals. A dose of estradiol that inhibited gonadotropin levels in juvenile rats, but not in adult animals, maintained preovariectomy serum estradiol levels more efficiently in the adult rats. Therefore, a more rapid removal of estradiol from the blood stream cannot explain its lower effectiveness in suppressing gonadotropin release in adult rats. Estradiol-negative feedback effectiveness remained maximal until the day of first proestrus and decreased markedly on the next day (first estrus), remaining low thereafter. "Resetting" of the gonadostat to estradiol negative feedback was advanced by inducing precocious puberty by means of hyperprolactinemia, but not by mimicking the periovulatory changes in serum estradiol and progesterone in the absence of an LH surge. Serum progesterone levels were much higher in postpubertal rats than in juvenile animals. Ovariectomy of juvenile rats slightly decreased the already low levels of serum progesterone, but it produced a striking progesterone decrease in postpubertal animals. Quantitative replacement of preovariectomy serum progesterone levels in adult rats, treated with an ineffective dose of estradiol, almost completely restored the prepubertal effectiveness of estradiol in inhibiting LH release and, to a lesser extent, release of FSH...

Delayed puberty induced by chronic suppression of prolactin release in the female rat.

To study the effect of PRL deficiency on the onset of puberty, PRL release was chronically inhibited by treating immature female rats with the dopaminergic receptor agonist, bromoergocriptine (CB-154). The resulting alterations in the time of puberty onset, and in other associated parameters, such as serum levels of pituitary hormones, ovarian responsiveness to gonadotropins and ovarian hCG receptor content were then evaluated. CB-154 was provided in the drinking water from day 22 onward at the concentration of 20 and 100 micrograms/ml. The treatment resulted in almost complete suppression of serum PRL levels throughout the entire period studied (day 22 to first diestrus). In contrast, serum GH, FSH, and LH levels were not depressed. Likewise, pulsatile release of FSH was not affected and only a subtle alteration in pulsatile LH release was apparent. The onset of puberty, as determined by the age at vaginal opening, and at first diestrus after the first estrus and by the presence of corpora lutea at sacrifice (first diestrus), was markedly delayed in the hypoprolactinemic (HPO) rats. This inhibitory effect of CB-154 was completely prevented by concomitant administration of PRL. Ovarian weight was significantly decreased in HPO rats at the three ages studied (27, 32, and 36 days of age). By day 36, 50% of the control animals had already ovulated, as compared with only 9% of the HPO rats. Microscopic examination of ovaries from HPO rats revealed a retarded follicular development. In vitro ovarian progesterone response to hCG studied at day 32 and 36 of age was reduced in the HPO rats. Uterine growth was also depressed in HPO rats, the ovaries of which, when incubated in vitro, failed to show the prepubertal increase in estrogen response to hCG seen in control rats between day 32 ad 36. Aromatase activity, as measured by the in vitro release of estradiol from ovaries incubated in the presence of an excess of androgen substrate, was depressed in HPO rats. hCG receptor content in the ovaries from HPO rats (counts per min [125]hCG bound per micrograms DNA) was also lower than that of control animals at day 32 and 34 but not at day 36. However, at this later time the hCG receptor content per milligram of ovary was still significantly reduced in HPO rats. The results support the view that PRL plays an important role in the process of ovarian development that leads to the onset of puberty in the female rat and that this effect is, at least in part, exerted through a positive influence of PRL on ovarian LH receptor content.

Relation of biogenic amines to onset of puberty in the female rat.

Changes in hypothalamic concentration and turnover index (TI) of norepinephrine (NE), dopamine (DA), and serotonin (5HT) were studied during the first estrous cycle at the onset of puberty. Rats were killed when 40% of the population showed open vaginas. They were classified according to the state of their reproductive tract and serum LH values, as in anestrus, early proestrus, late proestrus, estrus, or diestrus. Serum LH, PRL, and hypothalamic LHRH were measured by RIA. The TIs for DA and NE were measured by estimating their rate of decrease after administration of alpha-methyl-para-tyrosine, and for 5HT-TI by its increase after pargyline administration. An increase in NE-TI in the hypothalamus was observed between anestrus and early proestrus, a decrease in DA-TI during early and late proestrus, and a small increase in 5HT-TI between early and late proestrus. LH and PRL peaks were observed at late proestrus when DA-TI and NE-TI reached their lowest values and hypothalamic LHRH concentration was failing. After late proestrus, the DA-TI rose sharply, NE-TI remained essentially unchanged. 5HT-TI fell, and serum LH and PRL declined to basal levels. These results suggest that the LH and PRL surges at late proestrus during the first estrous cycle at puberty are associated with an increase in NE-TI at early proestrus, a progressive decline in DA-TI at early and late proestrus, and an increase in 5HT-TI at late proestrus.

Activation of growth hormone short loop negative feedback delays puberty in the female rat.

Implantation of GH in the median eminence of the hypothalamus of 23-day-old female rats tonically inhibited serum GH levels throughout prepubertal development (days 23-36) and depressed GH diurnal pulsatile release. Puberty was significantly delayed in GH-deficient animals, a delay associated with a blunted in vitro ovarian steroidal responsiveness to gonadotropins (particularly estradiol) and a marked decrease in prepubertal uterine weight, as evaluated 4, 7, and 13 days after GH implantation. The prepubertal body weight increase was also depressed. Neither ovarian weight nor serum levels of LH, FSH, PRL, or TSH were consistently altered by the GH implant. In addition, evaluation of pulsatile PRL release in 33-day-old rats revealed no difference between control and GH-deficient animals. Ovarian LH receptor content was lower in GH-implanted rats than in controls, suggesting that a decrease in LH receptors may be one of the mechanisms by which a chronic decrease in serum gH depressed the prepubertal ovarian estradiol and, to a lesser extent, the progesterone response to gonadotropins. A direct ovarian site of action for GH was indicated by the results of experiments in which GH was administered to immature hypophysectomized estrogen-treated rats. The in vitro ovarian progesterone response of the GH-treated animals to both hCG and human FSH was distinctly increased by prior in vivo GH treatment. This effect of GH was not reproduced either by the in vivo administration of LH at a dose calculated by RIA to be contaminating the GH preparation or by FSH at a dose that induced a marked increase in aromatase activity in the same ovaries. GH treatment of hypophysectomized rats failed to affect either aromatase activity or hCG-induced estradiol release, indicating that GH does not directly facilitate the production of estradiol by the ovary. The fact that intact rats with GH implants did not actually show a decrease in the ovarian estradiol response to hCG but, rather, failed to show an increased response at the same time as control animals strongly suggests that ovarian maturation was delayed in GH-deficient rats. It is suggested that, in addition to PRL, GH may also play a role in the process of prepubertal reproductive development by enhancing the steroidal response of the ovary to gonadotropins.

Pulsatile luteinizing hormone (LH) patterns in ovariectomized rats: involvement of norepinephrine and dopamine in the release of LH-releasing hormone and LH.

Cannulated ovariectomized (OVX) rats were bled every 10 min for 2 h to characterize their individual patterns of LH release. After the last sample, all rats were decapitated, and their brains were removed for analysis of LHRH, norepinephrine (NE), and dopamine (DA) levels in median eminence (ME), arcuate-ventromedial, and suprachiasmatic-medial preoptic region (Sch-PO) to determine if changes in NE, DA, and LHRH levels in any of those areas could be observed at different points during the pulsatile release of LH. The results showed that when LH levels started to increase, NE levels peaked in the Sch-PO, whereas a sharp drop in DA and LHRH levels in the ME was detected, which may reflect an acute release in the ME of both DA and LHRH. When LH levels reached peak values, NE levels in the Sch-PO returned to lower values, and DA and LHRH in the ME rose to higher levels. Inhibition of NE synthesis with diethyldithiocarbamate resulted in the suppression of LH pulses. L-Dopa administered after diethyldithiocarbamate induced an increased release of LH in both OVX and OVX, estrogen-primed rats with a simultaneous inhibition of PRL release. Peak levels of LH after L-dopa treatment coincided with increased DA levels in the ME, no change in NE, and a clear drop in LHRH. The results suggest that both NE and DA are involved in the pulsatile release of LH in OVX rats.

Direct hypothalamic control of vasoactive intestinal peptide (VIP) levels in the developing rat ovary.

We have previously identified the neurotransmitter vasoactive intestinal peptide (VIP) in nerve fibers of the immature rat ovary and showed that it stimulates steroid release by a mechanism involving increased synthesis of the components of the cholesterol side-chain cleavage enzyme complex. The present experiments were undertaken to study the ontogeny of ovarian VIP levels and to determine if they change in relation to the initiation of puberty. VIP was already detected in 2-day-old ovaries; levels remained constant at approximately 4.5 pg/mg ovary until the end of juvenile development (day 30). Thereafter, and preceding the peripubertal activation of the ovary, VIP levels increased two-fold, decreased gradually towards the first proestrus, and returned to juvenile values after the first ovulation. Transection of the ovarian nerves eliminated radioimmunoassayable VIP levels in both intact and hypophysectomized rats, indicating that ovarian VIP derives mostly from the extrinsic innervation of the gland. Treatment of hypophysectomized immature female rats with human chorionic gonadotropin (hCG), follicle stimulating hormone (FSH), growth hormone (GH), prolactin (Prl), estradiol, or their combination, failed to reproduce the peripubertal increase in VIP. In contrast, unilateral direct anodal current lesions of the left preoptic-anterior hypothalamic area (POA-AHA) of hypophysectomized juvenile rats led to a significant increase in VIP in the ipsilateral ovary. Surprisingly, both bilateral lesions and lesions of the right POA-AHA also increased VIP levels in the left ovary suggesting the existence of a marked asymmetry in the hypothalamic control of ovarian VIP. Lesions of the ventromedial nucleus, dorsal AHA or small lesions of the AHA were ineffective.(ABSTRACT TRUNCATED AT 250 WORDS)

Studies on the central effect of prolactin in inducing precocious puberty in the female rat.

Implantation of prolactin (PRL) into the median eminence (PRL-ME implants) of 23 day old female rats markedly advanced the onset of puberty, as measured by the age at vaginal opening and at first ovulation. Precocious puberty was preceded by steroidogenic activation of the ovary, as reflected by increases in uterine weight and an enhanced in vitro steroidal responsiveness of the ovary to hCG. The stimulatory effect of PRL-ME implants could not be attributed to alterations in the release of LH, FSH, GH or TSH from the anterior pituitary. Likewise, the PRL effect was neither exerted through the adrenal gland nor involved activation of a direct neural, vagal-mediated influence on the ovary. Furthermore, the effect of PRL-ME implants was not due to a decrease in pituitary secretion of opioid substances, which appear to restrain chronically gonadotropin release during female prepubertal development. These latter experiments also showed that administration of the opioid agonist, morphine, can delay the onset of puberty in the female rat. Although local exposure of the medial basal hypothalamus to high PRL levels is extremely effective in accelerating puberty, the mechanisms by which this effect is exerted remains to be elucidated.

Comparative ability of chicken and mammalian LHRH to release LH from rooster pituitary cells in vitro.

Biological properties of homogeneous solutions of chicken (c) and mammalian (m) LHRH were compared by their ability to release LH, in vitro, from a rooster pituitary cell incubation system. Homogeneity of the two LHRH species was confirmed by High Performance Liquid Chromatography (HPLC) using linear gradients of acetonitrile and phosphate buffer. A clear HPLC separation of [Gln8]-LHRH ( cLHRH ) and [Arg8]-LHRH ( mLHRH ) was obtained, with the former having a consistently longer retention time than the latter. cLHRH cause a greater (p less than .025) in vitro release of LH at low doses (less than 1 ng/2 X 10(5) live pituitary cells), but not at high doses (greater than 10 ng/2 X 10(5) live pituitary cells), than that caused by mLHRH . Our results indicate that rooster pituitary cells are significantly more sensitive to low doses of cLHRH than to similar doses of mLHRH , when assessed by their ability to release LH in vitro.

Median eminence and anterior pituitary degradation of luteinizing hormone releasing hormone in hens undergoing changes in luteinizing hormone secretion.

Studies described in this report provide physiological evidence for a possible involvement of median eminence (ME) and anterior pituitary (AP) luteinizing hormone releasing hormone degrading activity (LHRH-DA) in the genesis of the hen's preovulatory surge of luteinizing hormone (LH). Serum LH and progesterone (P4), ME LHRH content and LHRH-DA, and AP LHRH-DA were determined in laying hens in the following reproductive conditions: 1) a "spontaneous" preovulatory LH surge; 2) a "premature" preovulatory LH surge; and 3) an ovulatory failure induced by feed withdrawal. The premature preovulatory surge of LH occurred 3 h after P4 administration and was preceded by an increase in both ME and AP LHRH-DA and by a decrease in ME LHRH content 1 h after P4 administration. However, the premature preovulatory LH surge was associated with a decrease in LHRH-DA back to control levels as ME LHRH content increased, even though LHRH was presumably being released from the ME at this time to maintain the preovulatory surge of LH. Although similar changes in LHRH-DA (an increase in both ME and AP LHRH-DA, followed by a decrease) also preceded the spontaneous preovulatory surge of LH, its profile was significantly blunted and no changes in ME LHRH content were associated with this LH surge. In contrast, ovulatory failure was correlated with a decrease in ME LHRH content but no changes in LHRH-DA. Therefore, in the hen, ME and AP enzymatic degradation of LHRH I might be involved in the genesis of a premature preovulatory surge of LH.

The median eminence as a site for neuroendocrine control of reproduction in hens.

Based on events occurring during the genesis of a preovulatory surge of luteinizing hormone (LH) and an ovulatory failure of central origin, the hypothalamic median eminence (ME) is probably a major neuroendocrine control site for reproductive activity in the hen. The ratio of facilitatory to inhibitory (F:I) inputs on LH-releasing hormone (LHRH) neuronal terminals is an important determinant of the ME control site. The word "facilitatory" is defined as "stimulatory or increasing responsiveness to inputs". A relative increase in the F:I ratio of inputs on LHRH neuronal terminals at the ME is apparently involved in the genesis of the preovulatory surge of LH. Both an increase in neuropeptide Y (NPY) facilitatory inputs and a decrease in beta-endorphin (beta END) inhibitory inputs to ME-LHRH neuronal terminals are involved in the increase in the F:I ratio preceding the preovulatory surge of LH. Although the NPY component of this ratio (F) is apparently driven by the preovulatory surge of progesterone (P4) itself, its beta END component (I) might be related to the preovulatory surge of estradiol accompanying or preceding that of P4. As the egg-producing life of the hen progresses, a relative decrease in the F:I ratio on ME-LHRH neuronal terminals occurs. As a consequence of this decrease, stress-related inputs (e.g., feed restriction) can induce an hypothalamic failure of central origin in laying hens ending their 1st yr of production, but not in birds at the start of their egg-laying life. An increase in ME dopaminergic inhibitory inputs (I) on LHRH neuronal terminals is an apparent cause of the ovulatory failure induced by feed restriction.(ABSTRACT TRUNCATED AT 250 WORDS)