The role of the pineal gland in the regulation of LH-release in rats with different types of the anovulatory syndrome.

The effect of different doses of testosterone propionate was investigated in provoking the development of the constant estrous anovulatory (CEA) syndrome in the rat. A direct relationship was observed between the dose of neonatally administered androgen (NA) and the percentage occurrence of this syndrome. Pinealectomy and superior cervical sympathetic ganglionectomy elicited the development of marked thecal luteinization in the NA-CEA rat, but the formation of corpora lutea was limited after these operations. The efficacy of pinealectomy and ganglionectomy in provoking luteinization was inversely related to the dose of testosterone used for neonatal androgenization. The LH-RH sensitivity of the adenohypophysis to release LH was decreased in the NA animals, as well as in the light-induced CEA syndrome (LCE), whereas it was increased in those CEA rats in which this syndrome was provoked by frontal hypothalamic deafferentation (FHD). Pinealectomy and ganglionectomy were able to elicit ovulation and luteinization in the FHD animals, but were ineffective in the LCE and high dose NA rats. The results are discussed in relation to those reported by others, in an attempt to explain the multitude of dissociated effects.     

Effect of pinealectomy and of bilateral cervical ganglionectomy on serum LH levels in constant estrous-anovulatory rats.

The effects of pinealectomy or bilateral superior cervical ganglionectomy on serum luteinizing hormone (LH) levels and ovulation were studied in constant estrous-anovulatory rats bearing a frontal hypothalamic deafferentation (FHD). Ovulation returned in 65% of the animals undergoing pinealectomy or bilateral superior cervical ganglionectomy. Almost 50% of these ovulating animals showed elevated serum levels of LH on the day of vaginal proestrus, while the remainder ovulated without appreciable changes in serum LH concentrations. It is suggested that the absence of an LH peak in the latter cases was due to an insufficient sampling size. The results suggest that pinealectomy and cervical sympathectomy may result in LH release by interference with the hypothalamic and brain stem serotonergic system involved in the inhibition of LH release.     

Regulation of anterior pituitary and brain beta-adrenergic receptors by ovarian steroids

Ovariectomy of adult female rats (200-230g) resulted in an increase in beta-adrenergic receptors in the cerebral cortex, hypothalamus and anterior pituitary. The anterior pituitary had the largest overall increase as well as the most rapid increase in beta-adrenergic receptor density of the tissues examined. The increase in hypothalamic or cerebral cortical beta-adrenergic receptors became apparent only long after ovariectomy (7-14 days). Fourteen days after ovariectomy, the density of beta-adrenergic receptors was 79%, 40%, and 24% in excess of control values in crude membranes prepared from anterior pituitary, hypothalamus and cerebral cortex, respectively. Over the same interval, the plasma concentration of luteinizing hormone (LH) increased 28-fold, while the concentration of follicle-stimulating hormone (FSH) rose 5-fold compared to control levels. Estradiol replacement (20 micrograms/kg/day) in these animals for four days before sacrifice concomitantly reduced plasma levels of the gonadotropins as well as the density of beta-adrenergic receptors in both the anterior pituitary and the hypothalamus. Long-term steroid replacement during the fifth and sixth week after ovariectomy, with implants of estradiol and progesterone which released the steroids in approximately physiological concentrations, significantly reduced beta-adrenergic density in anterior pituitary, but not in the hypothalamic membranes. This treatment significantly reduced plasma LH, but not FSH. Beta-adrenergic receptor density was also found to fluctuate significantly during the 4-day estrous cycle. The highest values were found on proestrus, and the lowest on diestrus 1. These studies indicate that changes in plasma concentrations of gonadal steroids (e.g. during the estrous cycle) influence the density of beta-adrenergic receptors in tissues involved in the control and release of anterior pituitary gonadotropins.     

Role of the serotoninergic neuron system of the brain stem on the release of thyrotrophic and luteinizing hormone

1. Decrease of brain serotonin concentration, elicited by either parachlorophenylalanine treatment, surgical interruption of the ascending serotoninergic fibres, or by pinealectomy provokes an enhanced release both of TSH and LH. 2. Increased serotonin content of the brain, produced by intraventricular, intrahypothalamic or systemic administration of serotonin, results in a significant inhibition of the release of these two trophic hormones. 3. It is concluded that the serotoninergic neuron system of the brain stem represents an inhibitory mechanism in the neuroendocrine circuit regulating pituitary trophic hormone release.     

Relation of endogenous opioid peptides and morphine to neuroendocrine functions.

Morphine and the endogenous opioid peptides (EOP) exert similar effects on the neuroendocrine system. When adminstered acutely, they stimulate growth hormone (GH), prolactin (PRL), and adrenocorticotropin (ACTH) release, and inhibit release of luteinizing hormone (LH), follicle stimulating hormone (FSH),and thyrotropin (TSH). Recent studies indicate that the EOP probably have a physiological role in regulating pituitary hormone secretion. Thus injection of naloxone (opiate antagonist) alone in rats resulted in a rapid fall in serum concentrations of GH and PRL, and a rise in serum LH and FSH, suggesting that the EOP help maintain basal secretion of these hormones. Prior administration of naloxone or naltrexon inhibited stress-induced PRL release, and elevated serum LH in castrated male rats to greater than normal castrate levels. Studies on the mechanisms of action of the EOP and morphine on hormone secretion indicate that they have no direct effect on the pituitary, but act via the hypothalamus. There is no evidence that the EOP or morphine alter the action of the hypothalamic hypophysiotropic hormones on pituitary hormone secretion; they probably act via hypothalamic neurotransmitters to influence release of the hypothalamic hormones into the pituitary portal vessels. Preliminary observations indicate that they may increase serotonin and decrease dopamine metabolism in the hypothalamus, which could account for practically all of their effects on pituitary hormone secretion.     

Effects of 6-methoxy-1,2,3,4-tetrahydro-beta-carboline and yohimbine on hypothalamic monoamine status and pituitary hormone release in the rat

Shortly after administration of 6-methoxy-1,2,3,4-tetrahydro-beta-carboline (6-MeOTHBC) and yohimbine to normal or hypothyroid rats [the latter exhibiting chronically elevated levels of serotonin (5-HT) neuronal activity in the hypothalamus] there was a highly significant increase in hypothalamic noradrenaline (NA) activity and in ACTH release concomittant with a reduction in hypothalamic 5-HT activity (P less than 0.01) and in growth hormone (GH) (P less than 0.01) and in thyroid stimulating hormone (TSH) (P less than 0.01) release from the pituitary. Both compounds caused an increase in hypothalamic dopamine (DA) metabolism and in pituitary prolactin (PRL) release in normal rats (P less than 0.01) but only yohimbine exerted this action in hypothyroid rats. Lower doses of 6-MeOTHBC exerted a relatively specific effect in hypothyroid rats, reducing (P less than 0.01) 5-HT neuronal activity in parallel with pituitary TSH secretion (P less than 0.05). While gross effects of 6-MeOTHBC and yohimbine were similar with respect to their effects on NA and 5-HT status in the hypothalamus, there were quantitative differences. 6-MeOTHBC always caused a greater decrease in 5-HT turnover and a lesser increase in NA turnover than did yohimbine. On the basis of these studies we suggest that the effect of tetrahydro-beta-carboline-related alkaloids on pituitary hormone release may be due to their influence on hypothalamic monoamine status and the subsequent alteration of the hypothalamic-pituitary control system.     

Detrimental effects of short-term ethanol exposure on reproductive function in the female rat

To more completely assess the means by which alcohol impairs the female reproductive cycle in rats, we have measured hypothalamic luteinizing hormone-releasing hormone (LHRH), pituitary LHRH receptor content, and the serum levels of luteinizing hormone (LH), follicle-stimulating hormone (FSH), prolactin (Prl), and progesterone (P). After two successive cycles, the animals began receiving either an alcohol or a isocaloric control liquid diet regimen beginning on the first day of diestrus, with continued monitoring of the estrous cycle throughout the experiment. An additional set of controls consisted of animals maintained on lab chow and water provided ad libitum. Our results indicate that those animals receiving the control diets showed uninterrupted estrous patterns, whereas those animals receiving the alcohol diet remained in diestrus. Additionally, the alcohol-treated animals showed an increase (p less than 0.05) in LHRH content, with a concomitant decrease (p less than 0.01) in serum LH, and an increase (p less than 0.01) in serum Prl. No significant differences were detected in serum FSH levels or pituitary LHRH receptor content. No differences were detected in serum P levels. These results indicate that short-term alcohol administration disrupts the female reproductive cycle, causing persistent diestrus, and support our hypothesis that the alcohol-induced depression in serum LH levels is due to a diminished release rate of hypothalamic LHRH.     

Serum luteinizing hormone and ovulatory response to luteinizing hormone-releasing hormone in the estrous and anestrous domestic cat.

A heterologous double antibody radioimmunoassay was developed to measure changes in serum luteinizing hormone (LH) concentrations in estrous and anestrous queens (female domestic cats), following a single injection of varying doses (0--25 microgram) of luteinizing hormone-releasing hormone (LH-RH). No increase in serum LH was detected in any of the estrous or anestrous queens following a single saline injection. Treatment with LH-RH resulted in a sharp increase in serum LH concentration in both estrous and anestrous queens. Ovulations as observed by the presence of corpora lutea at laparoscopy occurred in none of four, one of four, two of four and four of four estrous queens receiving 0, 5, 10 or 25 microgram of LH-RH, respectively. Mean serum LH concentration of the ovulating queens was maintained at a higher level and did not return to basal level at the same time as that of nonovulating queens. The data show that: LH-RH can cause release of LH in both estrous and anestrous queens and induce ovulation in the estrous cat; the magnitude of LH response is influenced by the stage of the reproductive cycle; and the duration during which LH is maintained above basal level may play a significant role in ovulation induction in this coitus-induced ovulatory species.     

Seasonal variation in hypothalamic content of gonadotropin-releasing hormone (GnRH), pituitary receptors for GnRH, and pituitary content of luteinizing hormone and follicle-stimulating hormone in the mare

Seasonal changes in the hypothalamic-hypophyseal axis were investigated using tissue from 49 light-horse mares, of mixed breeding. Hypothalamic and pituitary tissues were collected at 5 intervals throughout the years 1981 and 1982, representing midbreeding season (July, n = 10), transition out of the breeding season (October, n = 11), midanestrus (December, n = 8), transition into the breeding season (March, n = 10), and again in the following midbreeding season (July, n = 10). The hypothalamic region was dissected into preoptic area, body and median eminence. Gonadotropin-releasing hormone (GnRH) was extracted from hypothalamic samples with methanol-formic acid and quantified by radioimmunoassay. The anterior pituitary was homogenized and receptors for GnRH were quantified in a crude membrane fraction. Concentrations of luteinizing hormone (LH) and follicle-stimulating hormone (FSH) were measured in the resulting supernatant. Content of GnRH in each of the 3 hypothalamic areas varied with season (P less than 0.01) and was lowest during midanestrus (P less than 0.05). There was no effect of season (P greater than 0.01) on either concentration or total number of receptors for GnRH, or concentration of FSH in the anterior pituitary. Concentrations of LH in the anterior pituitary varied with season (P less than 0.001). Means (+/- SEM) for the 5 collection times were 15.5 +/- 2.7, 9.7 +/- 2.4, 2.3 +/- 0.5, 2.7 +/- 0.4 and 11.7 +/- 1.5 microgram LH/mg anterior pituitary, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)     

Influence of endogenous opiates on anterior pituitary function

In general, the endogenous opioid peptides (EOP), morphine (MOR), and related drugs exert similar effects on acute release of pituitary hormones. Thus administration of opiates produces a rapid increase in release of prolactin (PRL), growth hormone (GH), adrenocorticotropin (ACTH), and antidiuretic hormone (ADH), and a decrease in release of gonadotropins and thyrotropin (TSH). Although not yet fully established, there is growing evidence that the EOP participate in the physiological regulation of pituitary hormone secretion. Thus naloxone (NAL), a specific opiate antagonist, has been shown to reduce basal serum levels of PRL and GH, and to elevate serum levels of LH and follicle stimulating hormone in male rats. Other reports have shown that NAL can inhibit the stress-induced rise in serum PRL, raise the castration-induced increase in serum LH to greater than normal castrate values, and counteract the inhibitory effects of estrogen and testosterone on LH secretion. Opiates appear to have no direct action on the pituitary, but there is evidence that they can alter activity of hypothalamic dopamine and serotonin in modulating secretion of pituitary hormones.     

A Mathematical Model for the Actions of Activin,Inhibin, and Follistatin on Pituitary Gonadotrophs

The timed secretion of the luteinizing hormone (LH) and follicle stimulating hormone (FSH) from pituitary gonadotrophs during the estrous cycle is crucial for normal reproductive functioning. The release of LH and FSH is stimulated by gonadotropin releasing hormone (GnRH) secreted by hypothalamic GnRH neurons. It is controlled by the frequency of the GnRH signal that varies during the estrous cycle. Curiously, the secretion of LH and FSH is differentially regulated by the frequency of GnRH pulses. LH secretion increases as the frequency increases within a physiological range, and FSH secretion shows a biphasic response, with a peak at a lower frequency. There is considerable experimental evidence that one key factor in these differential responses is the autocrine/paracrine actions of the pituitary polypeptides activin and follistatin. Based on these data, we develop a mathematical model that incorporates the dynamics of these polypeptides. We show that a model that incorporates the actions of activin and follistatin is sufficient to generate the differential responses of LH and FSH secretion to changes in the frequency of GnRH pulses. In addition, it shows that the actions of these polypeptides, along with the ovarian polypeptide inhibin and the estrogen-mediated variations in the frequency of GnRH pulses, are sufficient to account for the time courses of LH and FSH plasma levels during the rat estrous cycle. That is, a single peak of LH on the afternoon of proestrus and a double peak of FSH on proestrus and early estrus. We also use the model to identify which regulation pathways are indispensable for the differential regulation of LH and FSH and their time courses during the estrous cycle. We conclude that the actions of activin, inhibin, and follistatin are consistent with LH/FSH secretion patterns, and likely complement other factors in the production of the characteristic secretion patterns in female rats.     

Vasopressin regulation of the proestrous luteinizing hormone surge in wild-type and Clock mutant mice

In the female mouse, ovulation and estrous cyclicity are under both hormonal and circadian control. We have shown that mice with a mutation in the core circadian gene Clock have abnormal estrous cycles and do not have a luteinizing hormone (LH) surge on the afternoon of proestrus due to a defect at the hypothalamic level. In the present study, we tested the hypotheses that vasopressin (AVP) can act as a circadian signal to regulate the proestrous release of LH, and that this signal is deficient in the Clock mutant. We found that Avp expression in the suprachiasmatic nucleus (SCN) and AVP 1a receptor (Avpr1a) expression in the hypothalamus is reduced in Clock mutant mice compared to wild-type mice. Intracerebroventricular (i.c.v.) injection of AVP on the afternoon of proestrus is sufficient to induce LH secretion, which reaches surge levels in 50% of Clock mutant mice. The effect of AVP on the Clock mutant LH surge is mediated by AVPR1A, as co-infusion of AVP and an AVPR1A-specific antagonist prevents AVP induction of LH release, although infusion of an AVPR1A antagonist into wild-type mice failed to prevent a proestrous LH surge. These results suggest that reduced hypothalamic AVP signaling plays a role in the absence of the proestrous LH surge in Clock mutant mice. The results also support the hypothesis that AVP produced by the SCN may be a circadian signal that regulates LH release.     

Binding studies of substance P anterior pituitary binding sites: changes in substance P binding sites during the rat estrous cycle

Previous studies have shown that substance P (SP), an undecapeptide widely distributed in the gastrointestinal tract and in the peripheral and central nervous system, is a putative regulatory peptide involved in the control of reproductive function. Specifically, SP inhibited, at the anterior pituitary (AP) level, the stimulatory action of a physiological concentration (10(-8) M) of Gonadotropin Releasing Hormone (GnRH) on the release of the luteinizing hormone (LH). In the present work, we have demonstrated the presence of specific SP binding sites in the AP and related changes in the number of these sites to GnRH receptor number, hypothalamic SP and GnRH content and LH secretion during the rat estrous cycle. High affinity saturable SP binding sites (Kd, 1.5 approximately equal to 10 nM) were demonstrated in AP membranes using [3H]-SP or a novel analog, [125I]-(D-Tyr0, NorLeu11)SP. The binding affinity of SP fragments decreased with progressive removal of amino acid residues from N or C termini of the molecule. Other neuropeptides had low affinity for the SP binding sites. During the rat estrous cycle, SP and GnRH binding capacity of the anterior pituitary were inversely related. At the time of the proestrous LH surge, the AP binding capacity was low for GnRH but high for SP. The highest content of SP in the hypothalamus were recorded during the afternoon of proestrus when hypothalamic GnRH levels were lowest and the preovulatory surge occurred. These studies have established the presence of high affinity specific binding sites for SP in the AP which alter during the estrous cycle in a manner appropriate for mediating the direct inhibitory effects of SP on LH release in vitro.     

Circadian clock mutation disrupts estrous cyclicity and maintenance of pregnancy

Classic experiments have shown that ovulation and estrous cyclicity are under circadian control and that surgical ablation of the suprachiasmatic nuclei (SCN) results in estrous acyclicity in rats. Here, we characterized reproductive function in the circadian Clock mutant mouse and found that the circadian Clock mutation both disrupts estrous cyclicity and interferes with the maintenance of pregnancy. Clock mutant females have extended, irregular estrous cycles, lack a coordinated luteinizing hormone (LH) surge on the day of proestrus, exhibit increased fetal reabsorption during pregnancy, and have a high rate of full-term pregnancy failure. Clock mutants also show an unexpected decline in progesterone levels at midpregnancy and a shortened duration of pseudopregnancy, suggesting that maternal prolactin release may be abnormal. In a second set of experiments, we interrogated the function of each level of the hypothalamic-pituitary-gonadal (HPG) axis in order to determine how the Clock mutation disrupts estrous cyclicity. We report that Clock mutants fail to show an LH surge following estradiol priming in spite of the fact that hypothalamic levels of gonadotropin-releasing hormone (GnRH), pituitary release of LH, and serum levels of estradiol and progesterone are all normal in Clock/Clock females. These data suggest that Clock mutants lack an appropriate circadian daily-timing signal required to coordinate hypothalamic hormone secretion. Defining the mechanisms by which the Clock mutation disrupts reproductive function offers a model for understanding how circadian genes affect complex physiological systems.     

Stimulation of the hypothalamic-pituitary-adrenal axis and inhibition of growth hormone release via increased central noradrenaline neuronal activity by urethane anaesthesia in the rat: blockade by clonidine

Computerized gas chromatography-mass spectrometry was used to measure precisely the hypothalamic levels of noradrenaline (NA), dopamine and serotonin together with those of their major neuronal metabolites 3,4-dihydroxyphenylethyleneglycol (DHPG), 3,4-dihydroxyphenylacetic acid and 5-hydroxyindoleacetic acid in normal male rats 45 min after stimulation of hypothalamic-pituitary-adrenal function by urethane (1.3 g/kg) administration. Urethane treatment resulted in a significant elevation of central noradrenergic neuronal activity (NNA) as assessed from marked rises in hypothalamic DHPG concentrations and the ratio (DHPG/NA). At the same time there was significant stimulation of ACTH and corticosterone release and inhibition of growth hormone release. These hormonal and central effects of urethane (but not anesthesia) were inhibited when the alpha 2-agonist clonidine (150 micrograms/kg) was co-administered. Urethane had no major effect on hypothalamic dopamine or serotonin status. We propose that the release of ACTH and the suppression of growth hormone release following urethane anaesthesia is a result of activation of central NNA and suggest that the hormonal responses are mediated via hypothalamic noradrenergic facilitation of corticotrophin releasing factor and somatostatin release to the anterior pituitary.     

Naloxone stimulation of luteinizing hormone release in prepubertal female rats; role of serotonergic system

The purpose of this study was to determine whether naloxone stimulated LH release via a serotonergic mechanism. Injection of naloxone hydrochloride (2 mg/kg B.W.) into 25-day old female prepubertal rats resulted in a significant elevation in serum LH 30 min later. Injection of this dose of naloxone together with morphine sulfate (2 or 5 mg/kg B.E.) resulted in inhibition of naloxone-induced LH release. When rats were first injected with 5-hydroxytryptophan (5-HTP) to increase hypothalamic serotonin content, naloxone failed to increase serum LH levels. On the other hand, when parachlorophenylalanine (PCPA) was given first to reduce hypothalamic serotonin content, naloxone-induced LH release was potentiated. Morphine failed to inhibit the naloxone-induced rise in serum LH when PCPA was first administered. Neither 5-HTP nor PCPA, when injected alone, altered serum LH values. These results suggest that naloxone promotes LH release by reducing hypothalamic serotonergic activity, and morphine inhibits LH release by increasing hypothalamic serotonergic activity. This does not exclude possible involvement of other neurotransmitters.     

Depressed prolactin cell activity in long-term blind-pinealectomized female hamsters is due to loss of estrous cyclicity

Pinealectomy in the female golden Syrian hamster is not always completely effective in preventing the suppressive effects of long-term light deprivation due to blinding on pituitary prolactin (PRL) cell activity. We examined this curious phenomenon by measuring pituitary PRL mRNA levels, PRL synthesis, and radioimmunoassayable PRL, and correlating these changes with the status of estrous cyclicity. As expected, 12 weeks of light deprivation resulted in loss of estrous cyclicity and a greater than 90% decline in all indices of pituitary PRL cell activity, compared with intact cycling controls. Pinealectomy prevented only 40-50% of this decline. However, if noncycling light-deprived pinealectomized animals were excluded, pinealectomy was completely effective, i.e., cycling intact control animals were no different than cycling blind-pinealectomized. We conclude that the inability of pinealectomy to completely prevent the decline in prolactin cell activity seen after blinding is due to the loss of estrous cyclicity in some blind-pinealectomized females, with the attendant loss of the prolactin-stimulating hormone estrogen.     

Peptidase activity in the hypothalamus and pituitary of the rat: fluctuations and possible regulatory role of luteinizing hormone releasing hormone-degrading activity during the estrous cycle

Peptidase activity capable of inactivating luteinizing hormone (LHRH) may have a physiological role in partially determining hypothalamic LHRH levels as well as LHRH levels at the gonadotrope. In our previous work ( Lapp and O' Conner , 1984, companion paper), use of the synthetic substrate leucine-p-nitroanilide (Leu-p-NA) to assay LHRH-degradative activity was validated by several methods. The current studies were conducted in order to monitor peptidase activity in the hypothalamus and pituitary throughout the rat 4-day estrous cycle. Activity in both tissues was significantly decreased during proestrus and diestrus I. It seems possible that the proestrous reduction in peptidase activity represents a permissive period necessary for the induction of the LHRH and LH surges. The decreased degradative activity in the pituitary on diestrus I may be involved in inducing the pituitary LHRH receptors which are reportedly synthesized prior to proestrus. The peptidase exhibits positive cooperativity with Leu-p-NA, and the degree of this cooperativity also fluctuates during the estrous cycle. Estradiol and progesterone given alone or in combination to prepubertal castrate animals increased the activity of the hypothalamic peptidase in vitro. The degree of positive cooperativity with which the enzyme functioned was also apparently altered by these gonadal steroids.     

Effect of naloxone and beta-casomorphin on the hypothalamic-pituitary-luteinizing hormone axis in vitro.

The effect of naloxone and beta-casomorphin on luteinizing hormone (LH) release from pituitary cell aggregates, obtained by three-dimensional culture, with or without mediobasal hypothalamic fragments was studied in vitro. Short-term naloxone perifusion at a concentration of 10(-5)M did not modify either basal or LHRH-stimulated LH release from the pituitary cell aggregates. In contrast, a 12-min naloxone perifusion at the same concentration caused an increase in LH release in the mediobasal hypothalamic-pituitary cell aggregate axis. This increase was rapid (12-16 min after time pulse), marked [up to 10 times (p less than 0.004) the initial base line], short (return to the base line secretion 32-40 min after the beginning of the time pulse) and dose-dependent, with a rise greater than 1000% at a concentration of 10(-4) (p less than 0.006). The same effect was observed when a second pulse was applied 48 min after the first one. LH release induced by naloxone was antagonized 56 +/- 2% (p less than 0.03) by beta-casomorphin (an exogenous opiate) at a concentration of 10(-5) M. beta-casomorphin alone did not modify LH basal secretion, but inhibited 25.1 +/- 2.4% (p less than 0.008) LH release enhanced by LHRH. These results indicate that naloxone, an opiate antagonist, markedly increases LH release via a mu-type opioid receptor mechanism at the hypothalamic level only, during short-term exposure.     

Change in concentrations of luteinizing hormone subunit messenger ribonucleic acids in the estrous cycle of beef cattle.

Changes in the concentrations of LH subunit messenger ribonucleic acids (mRNAs) and in the LH content of the anterior pituitary of beef cattle were studied during the estrous cycle. Japanese beef cows were classified according to the expected day of the estrous cycle: stage I (early-luteal phase, days 1-4; day 1=day of ovulation), stage II (early-mid-luteal phase, days 5-10), stage III (late-mid-luteal phase, days 11-17) and stage IV (follicular phase, days 18-20), according to the morphology of the ovaries. The anterior pituitaries of the cows were collected and the levels of alpha and LHbeta subunit mRNAs were determined by slot-blot analyses. The LH content of the anterior pituitary was measured by radioimmunoassay. The level of alpha subunit mRNA in the pituitary of cows was highest in stage I and decreased significantly by stage II (P<0.05); thereafter it tended to increase. The level of LHbeta subunit mRNA did not change significantly during the estrous cycle. The LH content of the pituitary of cows was low in stage I and tended to increase by stage II, then to decrease from stage II to III, and to increase significantly from stage III to IV (P<0.05). These results suggest that the highest levels of gene expressions of alpha subunit in the anterior pituitary occur in the early-luteal phase of beef cows, while the LH content is increased most in the follicular phase. The enhanced gene expressions of common alpha subunit in the early-luteal phase could be important in replenishing the bovine anterior pituitary with LH, which is depleted of hormone by the LH surge or the enhanced pulsatile release.