Diestrous Female Rats Research Articles

Intranasal application of PACAP and \u03b2-cyclodextrin before the \u201ccritical period of proestrous stage\u201d can block ovulation

IntroductionIt was previously shown that intracerebroventricular administration of pituitary adenylate cyclase-activating polypeptide (PACAP) prior to GnRH mobilization in proestrus prevents ovulation in rats. In this study, we examined whether PACAP given intranasally could influence luteinizing hormone (LH) and prolactin (PRL) surges and ovulation.MethodsOn the day of proestrus PACAP, p-cyclodextrin (modifier of blood-brain barrier) or PACAP + p-cyclodextrin was applied intranasally between 12:30 and 13:00. Blood samples were taken at 16:00, 18:00, and 20:00 for measuring plasma hormone levels. In the next morning, the expelled ova were counted. p-Cyclodextrin was also administered to male and diestrous female rats between 12:30 and 13:00 and blood was taken at 18:00.ResultsPACAP prevented LH and PRL surges and ovulation in about half of the rats, p-cyclodextrin alone more effectively prevented ovulation. When PACAP and p-cyclodextrin were administered together, more rats ovulated like when PACAP given alone. p-Cyclodextrin did not influence LH and PRL levels in diestrous females; however, in males, it significantly enhanced PRL level.DiscussionNot only the intracerebroventricular, but the intranasal application of PACAP prevented ovulation. p-Cyclodextrin alone is more effective than PACAP and enhances PRL levels in male rats. PACAP and p-cyclodextrin given together weaken each other’s effect. p-Cyclodextrin, as excipient of various drugs, has to be used carefully in human medications.

Tropisetron, a 5-HT3 receptor antagonist, enhances object exploration in intact female rats

The effects of a 5-HT(3) receptor antagonist, tropisetron, on cognitive functions were evaluated using the object-recognition test in estrous (postovulatory) and in diestrous female rats. Recognition was measured by the ability of rats to discriminate between a familiar and a new object in a T-maze after a 3-h delay. Rats from both stages spent equivalent amounts of time exploring the objects on trial 1. Three hours after trial 1, trial 2 (test trial) was initiated. Before the test trial, rats were injected with either saline or 1.5 or 2.5 mg/kg tropisetron. During the test trial, one arm of a T-maze contained an object from trial 1 (familiar) and a new object (novel) was introduced into the other arm. Rats from both stages responded to tropisetron by showing a greater percentage of time exploring the novel object. These findings indicate that tropisetron facilitates cognition in female rats by improving the recognition of familiar information.

Low doses of cocaine decrease, and high doses increase, anxiety-like behavior and brain progestogen levels among intact rats

There are sex and hormonal differences in response to cocaine that have been demonstrated in people and animal models. Cocaine can alter secretion of progestogens, such as progesterone (P), and its neuroactive metabolite, 5α-pregnan-3α-ol-20-one (3α,5α-THP). However, little research has been done on the neuroendocrine effects in the initiation phase of cocaine use. We hypothesize that some sex/hormonal differences in initiation phase responses to cocaine may be related to formation of progestogens. To investigate the role of progestogens in sex differences in response to acute cocaine, male and female rats in the high (proestrous) or low (diestrous) progestogen phase of the estrous cycle were administered cocaine (0, 5, 10, or 20 mg/kg, IP). We examined cocaine's acute neuroendocrine effects on P and 3α,5α-THP levels, as well as its effects on acute psychomotor stimulation, anxiety, and sexual behaviors. Among rats that had P and/or 3α,5α-THP levels increased in response to cocaine, enhanced acute psychomotor stimulation was observed. Results suggest that cocaine produces U-shaped curves for progestogens, and anxiety-like behaviors. Male rats were less susceptible to these effects of cocaine than were proestrous or diestrous female rats. However, cocaine's disruption of sexual behaviors was similar among males and proestrous females. These data suggest a complex interaction between hormonal milieu and the neuroendocrine and behavioral effects of cocaine.

Sex-specific 24-h profile of extracellular serotonin levels in the medial prefrontal cortex

The medial prefrontal cortex (mPFC) controls emotional responses in many species, receiving serotonergic innervation from the dorsal and median raphe nucleus (DRN and MRN). To examine the sex difference in 24-h profiles of extracellular serotonin (5HT) levels in the mPFC, an in vivo microdialysis study was performed using intact male, diestrous female, and proestrous female rats. Dialysates were automatically collected by a microdialysis probe from the mPFC every 30 min for more than 24 h under freely moving conditions. The levels of 5HT in dialysates were quantified by high performance liquid chromatography. Extracellular 5HT levels exhibited episodic changes in the mPFC of both sexes of rats, with both diestrous and proestrous females exhibiting a clear diurnal change; the 5HT levels were high during the dark phase, but low during the light phase. In contrast, male rats exhibited relatively high 5HT levels throughout the day without significant diurnal changes. At mathematically analyzed trough, males showed higher 5HT levels than diestrous or proestrous females. The overall 24-h 5HT levels in males were significantly greater than proestrous females, but were not different from diestrous females. Further, stereological methods were used to examine the number of tryptophan hydroxylase (TrpH), but no sex differences in the number of TrpH immunoreactive cells in the DRN and MRN were observed. These results suggest that sex and/or the gonadal steroid environment may affect the 24-h profile of extracellular 5HT in the mPFC of rats without changes in the number of 5HT neurons in the DRN and MRN.

Overlapping litters in rats: Effects on maternal behavior and offspring emotionality

Female rats have a fertile postpartum estrus, which can result in a simultaneous gestation and lactation and later in the overlapping of two different-age litters. These different physiological and contextual situations may affect the maternal behavior of lactating rats and provoke long-lasting changes in the affective behavior of the litter. Therefore, we aimed to assess the effect of pregnancy and of litter overlapping on the maternal behavior of lactating rats and to describe the maternal- and anxiety-like behaviors of the juveniles that remained in contact with their younger siblings. Results showed that pregnant lactating rats spent more time outside the nest and less time nursing than non-pregnant mothers. On the other side, mothers with overlapping litters licked less the newborn pups than mothers with single litters. These deficits in maternal licking received by neonates were overridden by the juveniles' licking behavior to their younger siblings. Adult male and diestrous female rats reared with younger siblings showed a reduced anxiety-like behavior as compared to age-weaning matched animals without this experience. Thus, natural changes in the reproductive conditions and in the early experience, affect the maternal behavior and long-term modulate affective behavior of the individuals.

Antinociceptive Activity of Pleurotus ostreatus., an Edible Mushroom, in Rats

The antinociceptive potential of Pleurotus ostreatus. (Jacquin: Fries) P. Kummer (Tricholomataceae) was investigated in rats (doses used: 125, 500, and 1000 mg/kg). Male rats and female rats in proestrous, estrous, and diestrous stages were orally administered 1000 mg/kg of freeze-dried P. ostreatus. and the reaction times on hot-plate and tail-flick tests were recorded. In the hot-plate test, the reaction time was significantly prolonged in male rats and diestrous female rats. Marked and significant prolongation in the reaction time at 1 h in males (28% mid and 32% high dose) and up to 2 h in diestrous females (57% mid and 79% high dose after 1 h) were observed on the hot-plate test. This effect was dose-dependent. In contrast, none of the rats showed increases in reaction time in the tail-flick test. In the formalin test, in rats administered a 1000 mg/kg dose of P. ostreatus., pain was significantly suppressed in both phases (females: licking time, 23%, 48%; licking frequency, 28%, 28%; males: licking time, 32%, 43%; licking frequency, no significant change). P. ostreatus. possessed antihistamine activity (histamine wheal test). Naloxone blocked the antinociceptive activity in the hot-plate test; however, metochlopramide did not abolish the activity. P. ostreatus. also showed mild antioxidant activity. Further, a 1000 mg/kgdose of P. ostreatus. did not induce sedation (hole-board test). This dose did not cause mortality or show signs of acute toxicity or stress. It is concluded that P. ostreatus. shows antinociception against neurogenic and continuous inflammatory pain possibly by opioid mechanisms, antioxidative and antihistamine activities.

Orphanin FQ/Nociceptin Is a Physiological Regulator of Prolactin Secretion in Female Rats

Orphanin FQ/nociceptin (OFQ/N), the most recently identified endogenous opioid peptide, stimulates prolactin secretion in both male and female rats. OFQ/N, however, did not elicit this stimulatory effect through the mu-, delta-, or kappa-opiate receptor subtype. The role OFQ/N plays in prolactin regulation under physiological conditions and its mechanism of action are not known. The purpose of these studies was to determine the physiological significance and pharmacological specificity of the prolactin secretory response to OFQ/N. In addition, the role of the tuberoinfundibular dopaminergic (TIDA) neurons in mediating this response was examined. Opioid receptor-like-1 (ORL-1) receptors were blocked by pretreatment with compound B (Comp B), a purported OFQ/N antagonist, or receptor synthesis was disrupted by pretreatment with ORL-1 receptor antisense oligonucleotides. The prolactin secretory response to OFQ/N administration in diestrous females was measured. Furthermore, the suckling-induced prolactin response was also determined after Comp B pretreatment. TIDA neuronal activity was quantified in diestrous female rats to determine whether OFQ/N stimulates prolactin release by inhibiting TIDA neurons. OFQ/N significantly inhibited the TIDA neurons by 1 min, preceding the prolactin secretory response. Both Comp B and antisense pretreatment blocked the stimulatory effects of OFQ/N on prolactin release, and Comp B abolished the suckling-induced prolactin response. These studies indicate that OFQ/N is a potent stimulus for prolactin secretion in female rats and that it mediates this effect by rapid and transient inhibition of TIDA neuronal activity. Furthermore, OFQ/N plays a physiologically significant role in the regulation of prolactin secretion during lactation, and it mediates its effects via actions at the ORL-1 receptor subtype.

Sedative and GABAergic Effects of Ethanol on Male and Female Rats

Women consume less alcohol than men, and yet they are more susceptible than men to the negative medical consequences of alcohol use, such as cirrhosis of the liver, cardiac disease, and cognitive impairments. This sex difference is also reflected in animal studies, in which male and female rats differ on both behavioral and electrophysiological measures. Given that one significant difference between males and females is the cycling fluctuations of the sex hormones, this study aimed to compare the relative sensitivity of adolescent and adult rats of both sexes and varying estrous stages to the behavioral and electrophysiological effects of ethanol. Adult female rats were lavaged daily for estrous cycle assessment. Following administration of 5 g/kg ethanol, adolescent and adult male and female animals were observed for loss of the righting reflex. Then, using whole-cell recording, we tested the response of spontaneous, gamma-aminobutyric acid (GABA(A)) receptor-mediated inhibitory postsynaptic currents (sIPSCs) in hippocampal slices from drug-naïve adult male and female rats. Consistent with previous findings, adolescent animals were less sensitive than adults to the effect of ethanol on the righting reflex. In addition, adult proestrous and diestrous female rats were less sensitive than male rats to the sedative effects of ethanol. Finally, ethanol increased the frequency of sIPSCs in hippocampal pyramidal neurons and did so more potently in cells from male rats than in those from female rats. Female animals are less sensitive to the behavioral sedative effects of ethanol than adult male rats, and the effect is pronounced in the proestrous and diestrous states. This sex difference may be related to differential sensitivity of GABA receptor-mediated central nervous system function to ethanol in females.

The anorectic effect of fenfluramine is influenced by sex and stage of the estrous cycle in rats

The controls of food intake differ in male and female rats. Daily food intake is typically greater in male rats, relative to female rats, and a decrease in food intake, coincident with the estrous stage of the ovarian reproductive cycle, is well documented in female rats. This estrous-related decrease in food intake has been attributed to a transient increase in the female rat's sensitivity to satiety signals generated during feeding bouts. Here, we investigated whether sex or stage of the estrous cycle modulate the satiety signal generated by fenfluramine, a potent serotonin (5-HT) releasing agent. To examine this hypothesis, food intake was monitored in male, diestrous female, and estrous female rats after intraperitoneal injections of 0, 0.25, and 1.0 mg/kg D-fenfluramine. The lower dose of fenfluramine decreased food intake only in diestrous and estrous females, suggesting that the minimally effective anorectic dose of fenfluramine is lower in female rats, relative to male rats. Although the larger dose of fenfluramine decreased food intake in both sexes, the duration of anorexia was greater in diestrous and estrous female rats, relative to male rats. Moreover, the magnitude of the anorectic effect of the larger dose of fenfluramine was greatest in estrous rats, intermediate in diestrous rats, and least in male rats. Thus our findings indicate that the anorectic effect of fenfluramine is modulated by gonadal hormone status.

Red blood cell damage after trauma-hemorrhage is modulated by gender.

Previous studies have shown that trauma-hemorrhagic shock (T/HS) causes significant alterations in red blood cell (RBC) deformability and shape. Gender is becoming well recognized as a modulating factor in the pathophysiologic response to trauma. We hypothesize that female subjects are more resistant to adverse effects of T/HS on RBC deformability and shape than male subjects because of estrogen protection. Elongation index, a measure of RBC deformability, was examined in six male rats and four groups of female rats at different stages of the estrous cycle (six animals per group) before and 6 hours after T/HS by laser ektacytometry. RBC shape was determined by scanning electron microscopy. Lipid peroxidation was evaluated by measurement of malonyldialdehyde in plasma 6 hours after T/HS. Male rats had a significant decrease in RBC elongation index after T/HS as compared with preshock values. RBC deformability of the proestrous (PES) and estrous female rats did not decrease after T/HS. Postshock RBC deformability values in metestrous and diestrous female rats did not differ from male rats. Male rats and metestrous and diestrous females had a significant increase in the percentage of abnormally shaped RBCs immediately after shock and during the postresuscitation period. In PES and ES female rats, RBC shape distribution did not significantly change after T/HS. Plasma malonyldialdehyde levels at 6 hours post-T/HS were higher in the male rats than in PES female rats. These data show that high levels of estrogen in female rats are protective against shock-induced RBC damage and suggest that this effect is associated with a decrease in lipid peroxidation.

Regulation of leptin mRNA and protein expression in pituitary somatotropes.

Leptin, the ob protein, regulates food intake and satiety and can be found in the anterior pituitary. Leptin antigens and mRNA were studied in the anterior pituitary (AP) cells of male and female rats to learn more about its regulation. Leptin antigens were found in over 40% of cells in diestrous or proestrous female rats and in male rats. Lower percentages of AP cells were seen in the estrous population (21 +/- 7%). During peak expression of antigens, co-expression of leptin and growth hormone (GH) was found in 27 +/- 4% of AP cells. Affinity cytochemistry studies detected 24 +/- 3% of AP cells with leptin proteins and growth hormone releasing hormone (GHRH) receptors. These data suggested that somatotropes were a significant source of leptin. To test regulatory factors, estrous and diestrous AP populations were treated with estrogen (100 pM) and/or GHRH (2 nM) to learn if either would increase leptin expression in GH cells. To rule out the possibility that the immunoreactive leptin was bound to receptors in somatotropes, leptin mRNA was also detected by non-radioactive in situ hybridization in this group of cells. In estrous female rats, 39 +/- 0.9% of AP cells expressed leptin mRNA, indicating that the potential for leptin production was greater than predicted from the immunolabeling. Estrogen and GHRH together (but not alone) increased percentages of cells with leptin protein (41 +/- 9%) or mRNA (57 +/- 5%). Estrogen and GHRH also increased the percentages of AP cells that co-express leptin mRNA and GH antigens from 20 +/- 2% of AP cells to 37 +/- 5%. Although the significance of leptin in GH cells is not understood, it is clearly increased after stimulation with GHRH and estrogen. Because GH cells also have leptin receptors, this AP leptin may be an autocrine or paracrine regulator of pituitary cell function.

Effect of prenatal stress and gonadal hormone condition on depressive behaviors of female and male rats

Whether prenatal stress (PNS) and gonadal hormones may influence depressive behavior of rats in the forced swim test was investigated. In Experiment I, adult diestrous female rats had increased immobility, which is indicative of depression, but did not show any significant difference in the duration of struggling compared to intact adult males. In Experiment 2, the behavior of adult intact, castrated, or castrated dihydrotestosterone (DHT)- or estrogen (E 2)-replaced offspring of dams that were restrained under lights for 45 min on gestational day 18 (PNS) or were not subjected to gestational stress (non-PNS, control condition) were compared. There were no effects of PNS, but DHT and E 2 produced anti-depressant effects on behavior of male rats. Castration decreased struggling and increased immobility compared to intact rats. DHT or E 2 replacement was able to partially reinstate struggling and immobility behavior but not to levels of intact males. In Experiment 3, behavior of PNS or control rats that were in proestrus or were ovariectomized and DHT, E 2, or vehicle-replaced were compared. Ovariectomy decreased struggling and increased immobility compared to that of proestrous rats. E 2 or DHT to control females increased anti-depressant struggling behavior compared to ovariectomized control or PNS rats administered vehicle, which demonstrated greater duration of struggling than did E 2-primed, PNS rats. E 2 or DHT administration decreased immobility of PNS and control females. These findings suggest that E 2 and DHT have some anti-depressant effects but that modest PNS may alter E 2's ability to alleviate some depressive behavior in female, but not male rats.

Epidermal Growth Factor and Gonadotropin-Releasing Hormone Stimulate Proliferation of Enriched Population of Gonadotropes

Recent studies of epidermal growth factor (EGF) receptors on gonadotropes show that they appear early in the estrous cycle on immature gonadotropes, most of which could be identified by LH messenger RNA only. As diestrous gonadotropes translate the messenger RNAs, the percentages of LH and FSH cells with EGF receptors increase to reach a peak during proestrus. To learn more about the function of EGF in gonadotrope regulation, parallel studies of its mitogenic potential were conducted. To test this in a cell growth assay, we initially developed a protocol for enrichment of gonadotropes by counterflow centrifugation (elutriation). Analysis of immunolabeled cells in the enriched fraction showed that the population contained 90–95% cells with LH and/or FSH antigens. Less than 4% have TSH or PRL antigens, and less than 7% have ACTH antigens. About 15% of the enriched population expressed GH antigens in male rats and nearly 30% of the population express GH in females. This agrees with the known hormone storage overlap between these cells, especially in proestrous female rats. The MTT cell growth/cell death assay was then used to test the mitogenic potential of EGF, GnRH, and activin. This assay showed a linear relationship between plated cell numbers and optical density of the media after the MTT reaction was run. The enriched gonadotropes were plated in 96-microwell trays and grown for 3–4 days in the presence of defined media alone (no serum), or defined media containing 0.5–10 ng/ml EGF, 0.5–1 nm GnRH, 60 ng/ml activin or two of these factors. In all of the 12 experiments, each of the factors stimulated a 3- to 10-fold increase in optical density values, depending on the dose of the stimulating factor. The effects of any two factors were not additive. Because the MTT assays do not discriminate between mitogenic effects and enhanced cell survival, a second group of tests was run with mixed cultures of pituitary cells from diestrous female rats. These cells were cultured in the same combinations of EGF with and without GnRH for 3 h. During the last hour of culture, they were exposed to bromodeoxyuridine (BrDU) to identify cells that were synthesizing DNA. Cells in the S phase were thereby detected with dual immunocytochemical labeling for nuclear BrDU and gonadotropins. The analysis of dual labeled cells showed a 3-fold increase in percentage of LH or FSH cells with BrDU labeled nuclei following EGF or GnRH stimulation. The effects of the two growth factors were not additive. Collectively, these data confirm previous studies showing mitogenic functions for activin and now add EGF and GnRH as mitogens for gonadotropes.

Epidermal growth factor and gonadotropin-releasing hormone stimulate proliferation of enriched population of gonadotropes.

Recent studies of epidermal growth factor (EGF) receptors on gonadotropes show that they appear early in the estrous cycle on immature gonadotropes, most of which could be identified by LH messenger RNA only. As diestrous gonadotropes translate the messenger RNAs, the percentages of LH and FSH cells with EGF receptors increase to reach a peak during proestrus. To learn more about the function of EGF in gonadotrope regulation, parallel studies of its mitogenic potential were conducted. To test this in a cell growth assay, we initially developed a protocol for enrichment of gonadotropes by counterflow centrifugation (elutriation). Analysis of immunolabeled cells in the enriched fraction showed that the population contained 90-95% cells with LH and/or FSH antigens. Less than 4% have TSH or PRL antigens, and less than 7% have ACTH antigens. About 15% of the enriched population expressed GH antigens in male rats and nearly 30% of the population express GH in females. This agrees with the known hormone storage overlap between these cells, especially in proestrous female rats. The MTT cell growth/cell death assay was then used to test the mitogenic potential of EGF, GnRH, and activin. This assay showed a linear relationship between plated cell numbers and optical density of the media after the MTT reaction was run. The enriched gonadotropes were plated in 96-microwell trays and grown for 3-4 days in the presence of defined media alone (no serum), or defined media containing 0.5-10 ng/ml EGF, 0.5-1 nM GnRH, 60 ng/ml activin or two of these factors. In all of the 12 experiments, each of the factors stimulated a 3- to 10-fold increase in optical density values, depending on the dose of the stimulating factor. The effects of any two factors were not additive. Because the MTT assays do not discriminate between mitogenic effects and enhanced cell survival, a second group of tests was run with mixed cultures of pituitary cells from diestrous female rats. These cells were cultured in the same combinations of EGF with and without GnRH for 3 h. During the last hour of culture, they were exposed to bromodeoxyuridine (BrDU) to identify cells that were synthesizing DNA. Cells in the S phase were thereby detected with dual immunocytochemical labeling for nuclear BrDU and gonadotropins. The analysis of dual labeled cells showed a 3-fold increase in percentage of LH or FSH cells with BrDU labeled nuclei following EGF or GnRH stimulation. The effects of the two growth factors were not additive. Collectively, these data confirm previous studies showing mitogenic functions for activin and now add EGF and GnRH as mitogens for gonadotropes.

Effects of indomethacin on the selective release of follicle-stimulating hormone during the period of ovulation in the rat.

To determine whether indomethacin, a potent inhibitor of prostaglandins endoperoxide synthetase, affects the selective follicle-stimulating hormone (FSH) surge during the period of ovulation, the compound was administered intravenously (i.v.), concurrent with 10 IU human chorionic gonadotropin (hCG), to diestrous female rats at 16:00 hr. Indomethacin inhibited the number of ovulations in a dose-dependent manner, and treatment with 500 micrograms indomethacin reduced number of oocytes in the ampullae most effectively without enteric lesions. In the histological observation, oocytes that had began to mature were found not only in unruptured luteinized follicles but also in ovarian interstitium beneath ruptured luteinized follicle. Despite the inhibitory effects of indomethacin on ovulation, peri-ovulatory FSH and progesterone surges occurred in comparable levels and duration to vehicle-treated animals. These results indicate that indomethacin-induced inhibition of prostaglandin synthesis does not affect the selective release of FSH during the peri-ovulatory period.

Ketanserin and Anxiety Levels: Influence of Gender, Estrous Cycle, Ovariectomy and Ovarian Hormones in Female Rats

DÍAZ-VÉLIZ, G., N. T. ALARCÓN, C. ESPINOZA, N. DUSSAUBAT AND S. MORA. Ketanserin and anxiety levels: Influence of gender, estrous cycle, ovariectomy and ovarian hormones in female rats. PHARMACOL BIOCHEM BEHAV 58(3) 637–642, 1997.—The influence of gender, estrous cycle, ovariectomy and ovarian hormones on the behavioral effects of the 5-HT 2 receptor antagonist, ketanserin (KET), was studied. Intact males, female rats in the four stages of the estrous cycle and ovariectomized (OVX) female rats 14 days after surgery were used. The OVX rats received progesterone [PROG, 25 mg/kg, subcutaneously (SC)] and/or estradiol benzoate (EB, 10 μg/kg, SC). KET (3 mg/kg, SC) was injected 30 min before testing. All the animals were subjected to the following behavioral tests: exploration of an elevated plus-maze and retention of a passive-avoidance response. KET enhanced the exploration of the open arms in diestrous female rats but inhibited this behavior during the other stages of the cycle and in OVX rats injected either with oil or EB. This dose of KET was ineffective in males and in OVX rats injected with PROG. Furthermore, KET inhibited the retention of the passive avoidance response in males, in diestrous and metestrous female rats and in OVX rats injected with oil. In estrous females and in OVX rats injected with EB, KET enhanced the passive-avoidance response. These results demonstrate that the sensitivity to KET differs with the gender, estrous cycle and hormonal treatment and suggest that central serotonergic activity is influenced by the hormonal status of the animal.

Multiple opiate receptor subtypes are involved in the stimulation of growth hormone release by beta-endorphin in female rats.

The growth hormone (GH) secretory response to beta-endorphin and the involvement of opiate receptor subtypes in this response were determined in diestrous female rats. The involvement of the mu (mu), delta (delta) and/or kappa (kappa) site was determined by administering specific antagonists for each of these sites prior to beta-endorphin. beta-Funaltrexamine (1 or 5 micrograms) was administered to block mu sites, ICI 154,129 (5 or 25 micrograms) blocked delta sites and nor-binaltorphimine (8 micrograms) blocked kappa sites. The ability of these antagonists to block GH secretion following intravenous morphine administration was also determined. The opiate antagonists and beta-endorphin were administered into the lateral ventricle. A dose-response study for beta-endorphin indicated that 0.5 micrograms beta-endorphin was the minimum stimulatory dose for GH release, producing an approximately 4-fold increase in circulating levels of GH; lower doses of beta-endorphin did not stimulate secretion. All three antagonists were capable of blocking the stimulatory effects of beta-endorphin. These results provide evidence that all three opiate receptor subtypes are involved in the stimulatory effect of beta-endorphin on GH release.

Opiate receptor subtype involvement in the stimulation of prolactin release by beta-endorphin in female rats.

The prolactin secretory response to beta-endorphin and the involvement of opiate receptor subtypes in this response was determined in both diestrous and postpartum, lactating female rats. The involvement of the mu-, delta- and/or kappa-site was determined by administering specific antagonists for each of these sites prior to beta-endorphin. beta-Funaltrexamine (beta-FNA, 1 or 5 micrograms) was administered to block mu-sites, ICI 154,129 (5, 10 or 25 micrograms) blocked delta-sites and nor-binaltorphimine (norBNI, 8 micrograms) blocked kappa-sites. The ability of beta-FNA and ICI 154,129 to block prolactin secretion following morphine administration was also determined. A dose response study for beta-endorphin indicated that beta-endorphin, at doses as low as 25 ng, was a potent stimulus for prolactin release producing an increase in prolactin that mimicked the suckling-induced prolactin increase. In addition, all three antagonists were capable of antagonizing the stimulatory effect of beta-endorphin in both diestrous and postpartum female rats. These results indicate that beta-endorphin is a potent stimulus for prolactin secretion and that these three opiate receptor subtypes interact to produce its stimulatory effect on prolactin release.

The Role of the Mu1 Opioid Receptor Subtype in the Regulation of Prolactin and Growth Hormone Secretion by Beta‐Endorphin in Female Rats: Studies with Naloxonazine

The μ opioid receptor subtype has been reported to mediate the prolactin secretory response to opioids. This receptor subtype has been implicated in the morphine-induced prolactin increase, as well as the prolactin response to μ-specific opioid peptides. Subtypes of the μ receptor have been proposed and the μ(1) , site has been postulated as the receptor subtype involved in the morphine-induced prolactin secretory response. However, the role of this receptor subtype in mediating the endocrine effects of the endogenous opioid peptides has not been characterized. In order to determine the physiological significance of this receptor subtype, animals were pretreated with saline, WIN 44,441-3 (a μ, δ and κ antagonist) or naloxonazine (a μ(1) antagonist) followed by a stimulatory dose of morphine or β-endorphin. A dose response study for β-endorphin was conducted to determine the minimal stimulatory dose of β-endorphin on the prolactin and growth hormone (GH) secretory response. The dose response study indicated that β-endorphin is a more potent stimulus for prolactin release than for GH. A dose as low as 25 ng increased prolactin levels as much as 100-fold in both lactating and diestrous female rats. In contrast, 2.5 μg β-endorphin was required in order to consistently and significantly increase circulating levels of GH by 2- to 3-fold. WIN 44,441-3 antagonized the stimulatory effects of β-endorphin on both prolactin and GH secretion. Naloxonazine pretreatment abolished the morphine-induced prolactin secretory response, without affecting the GH increase in diestrous females. Naloxonazine also antagonized the prolactin response to β-endorphin in both lactating and diestrous females. In addition, it attenuated the GH secretory response but did not totally abolish it. These data indicate that β-endorphin elicits an increase in prolactin release through an opioid specific receptor which appears to be the μ(1) opioid receptor subtype. They further suggest that β-endorphin may increase GH levels, at least partially, via its action at this μ(1) site.

Growth hormone secretion in old female rats analyzed by the reverse hemolytic plaque assay.

Growth hormone (GH) release was studied in young (3-4-month-old) and old persistent diestrous (20-21-month-old) female rats using the reverse hemolytic plaque assay. A bimodal distribution of reverse hemolytic plaque area was observed in both young and old female rats. The mean and median of the plaque area of GH cells from old females were smaller than those from young female rats. The percentage of plaque-forming GH cells in old female rats was lower than in young female rats. The percentage of large plaque-forming GH cells (plaque area, more than 8 x 10(3) microns2) was lower in old female rats than in young female rats. GH-releasing hormone (GHRH) increased the mean and median of plaque areas in both young and old female rats. However, responsiveness to GHRH was reduced in old female rats. These results indicate that the amount of GH released from individual GH cells decreases with age in female rats, resulting in diminished GH secretion.