Β-endorphin Neurons Research Articles

Inhibitory pathways and the inhibition of luteinizing hormone-releasing hormone release by alcohol.

In this research we examined the mechanisms by which ethanol (EtOH) inhibits luteinizing hormone-releasing hormone (LHRH) release from incubated medial basal hypothalamic explants. EtOH (100 mM) stimulated the release of two inhibitory neurotransmitters: gamma-aminobutyric acid (GABA) and beta-endorphin. EtOH also inhibited NO production, indicative of a suppression of nitric oxide synthase (NOS) activity. This inhibition was reversed by naltroxone (10(-8) M), a micro-opioid receptor blocker, indicating that the inhibition of NOS by EtOH is mediated by beta-endorphin. EtOH also blocked N-methyl-d-aspartic acid-induced LHRH release, but the blockade could not be reversed by either the GABA receptor blocker, bicuculline (10(-5) M), naltroxone (10(-8) M), or both inhibitors added together. However, increasing the concentration of naltrexone (10(-6) M) but not bicuculline (10(-4) M) reversed the inhibition. When we lowered the concentration of EtOH (50 mM), the EtOH-induced blockade of LHRH release could be reversed by either bicuculline (10(-5) M), naltroxone (10(-8) M), or the combination of the two blockers. Therefore, GABA is partially responsible for the blockade of N-methyl-d-aspartic acid-induced LHRH release. The block by GABA was exerted by inhibiting the activation of cyclooxygenase by NO, because it was reversed by prostaglandin E(2), the product of activation of cyclooxygenase. Because the inhibition caused by the higher concentration of EtOH could not be reduced by bicuculline (10(-4) M) but was blocked by naltroxone (10(-6) M), the action of alcohol can be accounted for by stimulation of beta-endorphin neurons that inhibit LHRH release by inhibition of activation of NOS and stimulation of GABA release.

Control of rat hypothalamic pro-opiomelanocortin neurons by a circadian clock that is entrained by the daily light-off signal

Previous studies have clearly demonstrated that the immediate-early gene, c- fos can regulate, through its protein product Fos, the expression of the pro-opiomelanocortin gene. In the present study, immunohistochemistry for Fos and β-endorphin was used to assess the basal activity of hypothalamic pro-opiomelanocortin-producing neurons throughout a 12 h light/12 h dark cycle. Here, we showed that Fos is undetectable in most β-endorphin neurons from late morning until 30 min after light offset in the evening, whereas Fos is spontaneously expressed in these neurons after 1 h following dark onset. The number of β-endorphin neurons expressing Fos increases continuously during the first half of the dark phase, is maximal at the middle of this phase and decreases through late night and early morning, reaching a nadir 2–3 h after light onset. Acute shifts of lighting parameters allowed us to demonstrate that the light-off signal per se is neither sufficient nor necessary for Fos expression in β-endorphin neurons. However, when recurrent, this signal is able to entrain Fos expression after a period of adaptation to the new light/dark schedule. Moreover, an expression of Fos in β-endorphin neurons persists during subjective night in rat exposed to constant light or constant dark for two to three days. Thus, the occurrence of the daily rhythmic increase in the expression of Fos protein in hypothalamic pro-opiomelanocortin neurons exclusively at (subjective) night suggests that these neurons are, most likely, controlled by a (circadian) nocturnal oscillator. Our data also reveal an interesting property of this oscillator: its entrainment by the daily light-to-dark transition signal.

Proopiomelanocortin gene expression is decreased in the infundibular nucleus of postmenopausal women

Previous studies have shown that estrogen withdrawal decreases the secretion of β-endorphin from the monkey hypothalamus. In addition, there are consistent age-associated changes in β-endorphin neurons in the rodent. Based on these findings, we hypothesized that the activity of hypothalamic β-endorphin neurons would be decreased in the hypothalamus of postmenopausal women. In the present study, we examined the expression of proopiomelanocortin (POMC) mRNA, the precursor mRNA for β-endorphin, in the medial basal hypothalamus of premenopausal and postmenopausal women. Every 20th sagittal section through the hypothalamus was hybridized with a synthetic [ 35 S ]labeled, 48-base oligonucleotide probe complementary to POMC mRNA. Labeled neurons were counted and their somatic profile areas were measured with an image-combining computer microscope system. The number of POMC mRNA-containing neurons/section in the infundibular nucleus was reduced by 65% in postmenopausal women. In contrast, there was no significant difference in the number of neurons expressing POMC gene transcripts in the retrochiasmatic region. The POMC neurons in the retrochiasmatic area were also distinct morphologically from those in the infundibular nucleus. The differences between the infundibular and retrochiasmatic regions suggest that functional subgroups of POMC neurons exist in the human hypothalamus. Our findings provide evidence that the activity of hypothalamic POMC neurons is decreased in the infundibular nucleus of postmenopausal women. Both aging and gonadal steroid withdrawal may contribute to the decline in POMC gene expression in postmenopausal women.

Proopiomelanocortin (POMC) mRNA expression: distribution and region-specific down-regulation by chronic morphine in female guinea pig hypothalamus

There is compelling evidence that endogenous opioid peptides are regulated by exogenous opiates. Our previous studies have shown that the μ-opioid receptor protein and mRNA are down-regulated in the mediobasal hypothalamus of the female guinea pig following chronic morphine treatment. In addition, electrophysiological studies have shown that hypothalamic β-endorphin ( β-EP) neurons express μ-opioid receptors that are uncoupled and down-regulated following chronic morphine treatment. Currently, we tested the hypothesis that chronic morphine, which produces down-regulation of μ-opioid receptors, causes a down-regulation of pro-opiomelanocortin (POMC, the precursor of β-EP) mRNA expression in female guinea pig hypothalamus. Female guinea pigs were ovariectomized and implanted subcutaneously (s.c.) with 4×75 mg pellets for 2 days plus six more pellets of either morphine ( n=6) or placebo ( n=6) for another 5 days. Animals were sacrificed between 1000 and 1100 h on day 7. The expression of POMC mRNA were investigated using in situ hybridization histochemistry with a guinea pig specific 35S-labeled cRNA probe in hypothalamic tissue sections. POMC mRNA was localized to the arcuate nucleus (Arc) and median eminence (ME) of the medial basal hypothalamus. The distribution pattern was the same in both morphine and placebo control animals. However, the density of silver grains was less in morphine treated animals versus placebo control animals. Overall, the level of POMC mRNA was decreased by 22% in the Arc of morphine-treated guinea pigs as compared with the placebo controls ( p<0.05). This decrease in POMC mRNA expression was even greater in the caudal Arc (28%, p<0.01) in morphine-treated animals. These results suggested that the biosynthetic activity of POMC neurons is down-regulated with chronic exposure to morphine.

The Peptide Orphanin FQ Inhibits β-Endorphin Neurons and Neurosecretory Cells in the Hypothalamic Arcuate Nucleus by Activating an Inwardly-Rectifying K+ Conductance

Orphanin FQ (OFQ) is a novel heptadecapeptide whose structure resembles that of dynorphin A_1–17. Its receptor shares appreciable homology with µ-, δ- and ĸ-opioid receptors, and is highly expressed in the hypothalamus. The present study examined the effects of OFQ on neurons within the arcuate nucleus (ARC) of the mediobasal hypothalamus, using intracellular recordings from coronal slices. In current clamp, OFQ produced a hyperpolarization of ARC neurons, including those immunopositive for β-endorphin, tyrosine hydroxylase and gonadotropin-releasing hormone. This hyperpolarization was dose-dependent, insensitive to antagonism by naloxone and was associated with a decrease in input resistance. In voltage clamp, OFQ produced an outward current associated with an increase in conductance. Varying the extracellular K⁺ concentration shifted the reversal potential for the OFQ response to the degree predicted by the Nernst equation. Furthermore, barium chloride markedly attenuated both the OFQ-induced hyperpolarization and decrease in input resistance. Administration of maximally effective concentrations of OFQ, followed by coadministration of maximal concentrations of either OFQ and the µ-opioid receptor agonist DAMGO or OFQ and the GABA_B receptor agonist baclofen produced additive hyperpolarizations and outward currents. If DAMGO was applied first, followed by the coadministration of DAMGO and OFQ, then the responses were occluded. Taken together, these results indicate that OFQ inhibits β-endorphin neurons, as well as A₁₂ dopamine and GnRH neurosecretory cells, within the ARC by activating a subset of inwardly-rectifying K⁺ channels. This suggests that OFQ is not only an antiopioid peptide, but that it also modulates the hypothalamo-pituitary axis and, ultimately, reproductive behavior.

The secretory response of hypothalamic β-endorphin neurons to acute and chronic nicotine treatments and following nicotine withdrawal

In the present study, we determined the effect of acute and chronic nicotine treatments on the secretion of immunoreactive β-endorphin (IR-β-EP) and cell viability of cultured hypothalamic neurons. Also, we determined the secretory response of IR-β-EP following withdrawal from a long-term nicotine treatment. Fetal hypothalamic cells were dissociated and maintained in cultures for 9 days and were treated with various doses of nicotine (1, 6,12 and 18 μM) for 6 h (acute treatment) or treated with nicotine at 12 h intervals for 96 h (chronic treatment). Determination of IR-β-EP concentrations in the media revealed that 6–18 μM doses of nicotine increased IR-β-EP secretion from these cultures for a period of 24 h; after this period, the cultured cells did not respond to these doses of nicotine. The desensitization of β-EP neurons 24 h after treatment with nicotine did not appear to be related to the loss of viable cells. Determination of withdrawal response after 72 h of constant nicotine (6 μM) treatments revealed that the hypothalamic neurons secrete elevated amounts of IR-β-EP for a period of 72 h after nicotine withdrawal. These results suggest that: 1) acute treatment with nicotine stimulates hypothalamic IR-β-EP release; 2) chronic nicotine treatment desensitizes β-EP-secreting neurons and, 3) β-EP neurons in primary culture show withdrawal response to nicotine.

NPY Y1 receptor like immunoreactivity exists in a subpopulation of β-endorphin immunoreactive nerve cells in the arcuate nucleus: a double immunolabelling analysis in the rat

Double immunolabelling immunohistochemistry in the arcuate nucleus of the rat demonstrates that neuropeptide Y (NPY) Y1 receptor like immunoreactivity of β-endorphin is strongly present in a subpopulation β-endorphin immunoreactive nerve cell bodies, while the small NPY immunoreactive nerve cell bodies lie in an arcuate area rich in NPY immunoreactive nerve terminals forming an uniform plexus. It is postulated that NPY Y1 receptors in β-endorphin neurons may mediate some actions of NPY on motivational processes and pain control as well as on hypophyseal hormone secretion, involving at least in part a regulation of the tubero-infundibular dopamine (DA) neurons.

The effect of estradiol-induced hypothalamic pathology on sulfated glycoprotein-2 (clusterin) expression in the hypothalamus

Sulfated glycoprotein-2 (SGP-2 or clusterin) is a complex multifunctional molecule that has recently been implicated in neuronal degeneration and remodeling. We have shown that estradiol treatment results in a selective destruction of β-endorphin neurons in the hypothalamic arcuate nucleus. We have used immunocytochemistry to determine the distribution of SGP-2 immunoreactivity in the rat hypothalamus and to assess the effects of the estradiol-induced destruction of β-endorphin neurons on SGP-2 expression. We have found that SGP-2-immunopositive neurons normally occur in the medial preoptic area (MPOA), supraoptic nucleus (SON), paraventricular nucleus (PVN), dorsomedial nucleus (DM), and the lateral hypothalamic area (LHA) in both males and females. The neuropil appears free of label. Treatment with estradiol valerate results in the appearance of immunopositive punctate deposits in the neuropil in the MPOA, PVN and DM. The number and distribution of SGP-2-positive neurons are unaffected by estradiol treatment except in the MPOA, where there are twice as many SGP-2-positive neurons as in controls. These effects are precluded by treatment with vitamin E, which blocks the cytotoxic action of estradiol on β-endorphin neurons. Thus, we interpret these changes as responses to the loss of β-endorphin afferents.

Estrogen modulation of two subpopulations of β-endorphin neurons in ovariectomized guinea pigs distinguished by peripherally injected fluorogold

β-endorphin released by neurons in the arcuate nucleus affects the output of several neuroendocrine systems and estrogen levels modulate the production and secretion of β-endorphin. We used intraperitoneal injection of fluorogold to retrogradely label the cell bodies of neurons that project outside the blood-brain-barrier in conjunction with immunohistochemistry for β-endorphin to dual label the subpopulation of β-endorphin neurons that project to the median eminence or other sites of access to the peripheral circulation. We found that some identified β-endorphin neurons in the arcuate nucleus of ovariectomized guinea pigs sequestered fluorogold. Approximately 7% of β-endorphin-containing cells co-localized with fluorogold. The effect of estrogen on the number of identified β-endorphin cells was examined. A single estradiol benzoate injection to ovariectomized guinea pigs 24 h prior to sacrifice dramatically decreased the total number of β-endorphin cells identified in the rostral, medial and the caudal portions of the arcuate nucleus. Also, a significantly smaller percentage of fluorogold-filled cells was found to contain β-endorphin immunoreactivity in the estrogen-treated group. These data suggest that a subpopulation of β-endorphin neurons has access to the peripheral circulation and may alter the output of neurosecretory terminals at the level of the median eminence. Furthermore, estrogen affects this subpopulation and the general population of β-endorphin neurons in the arcuate nucleus in a similar manner.

Effects of age and long-term ovariectomy on the estrogen-receptor containing subpopulations of beta-endorphin-immunoreactive neurons in the arcuate nucleus of female C57BL/6J mice.

We have reported a decrease in the number of arcuate nucleus (ARC)-immunoreactive beta-endorphin neurons in old (24 months) female C57BL/6J mice versus young (5 months) mice. Here, we have tested by immunocytochemistry whether age-related changes in beta-endorphin neuron numbers are selective for beta-endorphin neurons which do or do not contain estrogen receptors (E2R). We also compared beta-endorphin neuron number in mice with short- (S) and long-duration (L) ovariectomy (OVX), since the latter may protect against neuroendocrine aging. Mice were studied at 5 (young), 12 (middle-aged), or 23-24 months (old). When the mean number of neurons per tissue section (15 sections per animal) was examined, there were no significant differences between young and middle-aged S-OVX females for either beta-endorphin, E2R, or beta-endorphin/E2R neuron number. However, there were significant decreases in beta-endorphin-containing neurons in the oldest age group versus young females (young S-OVX: 74.4 +/- 11 (+/- SD) immunopositive neurons per tissue section, n = 10 mice; young L-OVX: 61.6 +/- 6.9, n = 6; old S-OVX: 45.7 +/- 9.9, n = 7; and old L-OVX: 37.5 +/- 7.3, n = 7). There were also decreases in beta-endorphin neurons which contained E2R in the oldest animals (young S-OVX: 16.6 +/- 6.4; young L-OVX: 13.7 +/- 1.3; old S-OVX: 9.2 +/- 1.8; L-OVX: 6.0 +/- 1.5) (p < 0.05 ANOVA). Both age (p < or = 0.001, two-way ANOVA) and ovarian status (p < or = 0.05) independently affected neuron number for both the beta-endorphin and beta-endorphin/E2R populations versus young mice. We tested whether the observed age and/or ovarian-related decreases were proportionally greater in the subpopulation of beta-endorphin neurons which contained E2R compared to the total beta-endorphin neuron population. In the oldest age group, there was no significant difference in the decrease with age in the population of beta-endorphin neurons which contained E2R and the total beta-endorphin population (p = 0.208). When we examined the E2R neuron population as compared to the beta-endorphin neuron populations, age-related decreases in the beta-endorphin neuronal population tended to be greater than the decreases seen in the E2R neuron population (p = 0.054 repeated measures ANOVA). The tyrosine hydroxylase (TH) neuron population was studied to test whether there were changes in another ARC neuron population.(ABSTRACT TRUNCATED AT 400 WORDS)

Tolerance in hypothalamic β-endorphin neurons following chronic morphine

Tolerance in hypothalamic β-endorphin neurons following chronic morphine

The 21-aminosteroid antioxidant, U74389F, prevents estradiol-induced depletion of hypothalamic β-endorphin in adult female rats

A single intramuscular injection of 2 mg estradiol valerate (EV) results in neuronal degeneration and β-endorphin depletion in the hypothalamic arcuate nucleus of adult female rats. We have hypothesized that peroxidase-positive astrocytes in this brain region oxidize estrogens and catecholestrogens to semiquinone radicals which mediate oxidative neuronal injury. In the present study, dietary administration of the potent antioxidant 21-aminosteroid, U-74389F, completely blocked EV-induced β-endorphin depletion in the hypothalami of adult female rats. Neither EV nor 21-aminosteroid treatment had any effect on hypothalamic concentrations of neuropeptide Y and Met-enkephalin, confirming that the estradiol lesion is fairly selective for the β-endorphin cell population. The present findings support the hypothesis that the toxic effect of estradiol on hypothalamic β-endorphin neurons is mediated by free radicals.

Immunohistochemical colocalization of tyrosine hydroxylase and estradiol receptors in the sheep arcuate nucleus

In sheep, the arcuate nucleus contains numerous tyrosine hydroxylase (TH) and estradiol receptor (rE2) immunoreactive (IR) perikarya and it has been shown previously in this species that catecholaminergic neurons can mediate the gonadal steroid action on the reproductive function. In the present study, double immunohistochemical labelling with antibodies against TH and rE2 have been used to demonstrate the presence of rE2 in TH-IR neurons in the arcuate nucleus where the distribution of TH-IR and rE2-IR neurons overlap each other. Only less than 10% of all the rE2-IR perikarya presented TH immunoreactivity. It was therefore hypothesized that either such a low number of double labelled neurons can support the effects of estradiol in this area or that the effect of this steroid was indirect. In the latter case it might be first mediated by β-endorphin neurons which have been previously described in this nucleus.

GABAergic and catecholaminergic innervation of mediobasal hypothalamic β-endorphin cells projecting to the medial preoptic area

In the absence of cellular estrogen receptors or proven direct estrogen action in the rat, it is assumed that estrogen indirectly regulates the secretory activity of the preoptic area luteinizing hormone-releasing hormone-producing cells. We have previously shown that pro-opiomelanocortin neurons in the arcuate nucleus of the rat send axons rostrally to connect with luteinizing hormonereleasing hormone neurons of the preoptic area. An experiment combining retrograde tracing and double-immunostaining was used to test the hypothesis that rat GABAergic and/or catecholaminergic neurons can influence luteinizing hormone-releasing hormone-producing cells via mediobasal hypothalamic β-endorphin neurons. The retrograde tracer horseradish peroxidase was injected into the medial preoptic area; two days later, arcuate nucleus Vibratome sections were double-immunostained for β-endorphin and glutamate decar☐ylase or tyrosine hydroxylase. Light and electron microscopic analysis of these triple-labeled sections demonstrated that a population of β-endorphin-immunoreactive neurons concentrated in the ventromedial arcuate nucleus contain retrogradely transported horseradish peroxidase granules and form synaptic contacts with glutamate decar☐ylase- and tyrosine hydroxylase-immuno-reactive axon terminals. The present data suggest that arcuate nucleus GABA and catecholamine fibers may influence luteinizing hormone-releasing hormone-containing neurons via projective pro-opiomelanocortin cells.

β-Endorphin-immunoreactive neurons in the hypothalamus and medulla of the rat brain: Effect of prenatal ethanol

The number and distribution of β-endorphin neurons in the hypothalamus and medulla of rats is compared among three experimental treatment groups: prenatally ethanol-exposed rats, pair-fed control rats, and normally fed control rats. Quantitative analysis of β-endorphin-immunoreactive neurons in 12 sections through the rostral-caudal extent of the mediobasal hypothalamus revealed no significant difference in the total number or distribution of β-endorphin neurons among any of the three treatment groups. The mean number of β-endorphin neurons was consistently less in each of the 12 sections of pair-fed rats compared to normally fed rats, but there was no consistent difference between the mean number of neurons/section in prenatally ethanol-exposed rats compared to the normally fed controls. The distribution and time of appearance of a second population of β-en-dorphin neurons was analyzed in the caudal medulla. β-endorphin-immunoreactive neurons were easily visualized in the caudal medulla of rats from each treatment group on Postnatal Days 9–11. However, this population could not be identified with antisera against β-endorphin on Postnatal Days 19–21 in any treatment group, even though neurons in the hypothalamus of the same rats were easily visualized histochemically at this age. These data suggest that the well-characterized prenatally ethanol-induced alterations in central opioid systems cannot be attributed to changes in the number, distribution, or time of appearance of β-endorphin neurons in the brain.

Central peptidergic neurons as targets for glucocorticoid action. Evidence for the presence of glucocorticoid receptor immunoreactivity in various types of classes of peptidergic neurons

By means of double immunolabeling procedures it has been possible to demonstrate glucocorticoid receptor (GR) immunoreactivity (IR) in large numbers of various peptidergic neurons of the brain including neurons containing gastrointestinal peptides, opioid peptides, and peptides with a hypothalamic hormone function. For each peptide system, however, marked heterogeneities exist among brain regions. Thus, in the neocortex and the hippocampal formation most of the brain peptide neurons lack GR IR, while the same types of peptide neurons in the arcuate and paraventricular nucleus [e.g. neuropeptide Y (NPY), somatostatin (SRIF) and the cholecystokinin (CCK) neurons] possess strong GR IR. Furthermore, in the arcuate, parvocellular part of the paraventricular nuclei aand the central amygdaloid nucleus practically all the peptidergic neurons are strongly GR IR, while in the lateral hypothalamus, mainly the neurotensin (NT) and galanin (GAL) IR neurons are GR IR. These marked differences among areas probably reflect functional differences dependent upon their participation in stress regulated circuits. All the paraventricular NT, corticotropin-releasing factor (CRF), growth hormone-releasing factor (GRF), thyrotropin-releasing hormone (TRH) and SRIF IR neurons appear to contain GR IR, while the luteinizing hormone-releasing hormone (LHRH) IR neurons lack GR IR, underlying the importance of glucocorticoids (GC) in controlling endocrine function. Finally, the GC may influence pain and mood control mainly via effects on enkephalin (ENK) neurons especially in the basal ganglia (mood) and on all β-endorphin (β-END) neurons of the arcuate nucleus, while most of the dynorphin neurons are not directly controlled by GC.

Opioids Hyperpolarize β-Endorphin Neurons via μ-Receptor Activation of a Potassium Conductance

Intracellular recordings were made from hypothalamic arcuate (ARC) neurons with biocytin-filled electrodes under current- and voltage-clamp in slices prepared from ovariectomized guinea pigs which were pretreated with estradiol. Forty-three neurons were identified after linking the intracellular biocytin with streptavidin-FITC and subsequently were examined for beta-endorphin immunoreactivity. Ten of these neurons were immunoreactive for beta-endorphin. beta-Endorphin neurons displayed the following passive membrane properties: RMP:-56 +/- 2 mV; Rin: 439 +/- 66 M omega; tau: 17.5 +/- 2.4 ms; and often fired spontaneously (5.9 +/- 2.2 Hz). These membrane characteristics were not different from identified neurons in the ARC that were not immunoreactive for beta-endorphin. beta-Endorphin neurons exhibited instantaneous inward rectification and time-dependent rectification. The mu-opioid agonist Tyr-D-Ala-Gly-MePhe-Gly-ol (DAGO) decreased spontaneous firing, induced membrane hyperpolarization (12 +/- 2 mV; range 6-22 mV) and decreased the Rin (38 +/- 4%) of the beta-endorphin neurons. These effects of DAGO were blocked by the opioid antagonist naloxone (1 microM) and were not blocked by 1 microM TTX. DAGO-responsive cells were unaffected by either kappa- or delta-receptor opioid agonists. These results indicate that mu-receptors may be autoreceptors on ARC beta-endorphin neurons and that activation of opioid mu-receptors hyperpolarizes beta-endorphin neurons via an increase in K+ conductance. Therefore, opioid peptides may modulate opioid tone through an 'ultra-short loop' feedback control mechanism.

The effects of castration, testosterone replacement and photoperiod upon hypothalamic β-endorphin levels in the male syrian hamster

Syrian hamsters kept in long day-lengths have active gonads and high circulating levels of gonadal steroids. Under the influence of the pineal gland, animals exposed to short photoperiods undergo testicular regression, have low circulating levels of testosterone and gonadotrophins and elevated levels of β-endorphin within the hypothalamus. This paper describes the interaction between testosterone and photoperiod in the regulation of β-endorphin levels in three regions of the hypothalamus. Hypothalamic β-endorphin levels were measured by a combination of high-performance liquid chromatography and radioimmunoassay techniques that allows separation of the β-endorphin (1–31) peptide from its metabolites and precursors. All of the β-endorphin-like immunoreactivity in the hypothalamus of the male hamster, in both photoinhibited and photostimulated conditions, was found to represent the 31-aminoacid peptide. In photostimulated hamsters, chronic castration was associated with a significant increase of β-endorphin levels in the anterior hypothalamus and mediobasal hypothalamus, which was reversed by treatment with exogenous testosterone. Castration prevented the ability of naloxone, an opiate receptor antagonist, to release luteinizing hormone, and this effect was also reversed by exogenous steroid. In photoinhibited hamsters, however, castration had no effect upon β-endorphin levels in the preoptic area or mediobasal hypothalamus, and there was only a small increment in the anterior hypothalamus. Significantly, β-endorphin levels in all areas of the hypothalamus of photoinhibited castrates were not decreased by testosterone treatment. In addition, administration of exogenous testosterone did not restore sensitivity to naloxone in these animals. These results show that inhibitory photoperiods uncouple the functional interaction between testosterone and the hypothalamic β-endorphin system, and reinforce the proposition that a change in the activity of β-endorphin neurons is a component of the central mechanism which mediates the environmental control of reproduction.

Estradiol target sites immunoreactive for β-endorphin in the arcuate nucleus of rat and mouse hypothalamus

Combined autoradiography and immunocytochemistry was performed to examine estradiol-concentrating neurons of the arcuate nucleus in the hypothalamus of ovariectomized rats and mice for β-endorphin-like immunoreactivity. A high number of [ 3H]estradiol-labeled cells occurred in the arcuate nucleus of both species. Neurons immunoreactive for β-endorphin could be visualized throughout the arcuate nucleus as well as ventrolaterally outside of this nucleus. Colocalization of [ 3H]estradiol nuclear labeling and β-endorphin-like cytoplasmic immunoreactivity could be found scattered throughout the arcuate nucleus, most frequently in the ventromedial part, although most of the [ 3H]estradiol target neurons did not react with the antibodies. The results suggest that a subpopulation of β-endorphin neurons is directly addressed by estradiol and that the arcuate nucleus contains different groups of estradiol receptive neurosecretory cells.

Acetylated and nonacetylated forms of β-endorphin in rat brain and pituitary

Extracts of rat posterior intermediate pituitary and extracts of brains from normal and hypophysectomized rats were separated by gel filtration chromatography and fractions were analyzed by both a classical β-endorphin radioimmunoassay and by a radioimmunoassay specific for α-N-acetyl β-endorphin. In posterior intermediate pituitary extracts, more than 90 percent of the β-endorphin-sized immunoreactive material was α-N-acetylated. In extracts of brains from normal rats, less than 2 percent of the β-endorphin-sized immunoreactive material corresponded to α-N-acetylβ-endorphin, whereas in brains from hypophysectomized animals, no α-N-acetylβ-endorphin-like material could be detected. Immunofluorescence on normal brain sections, using either affinity purified antibodies to α-N-acetylβ-endorphin or conventional β-endorphin antibodies, showed no α-N-acetylβ-endorphin immunoreactivity in β-endorphin neurons. Only in brain sections which had been acetylated in vitro prior to immunostaining could α-N-acetylβ-endorphin-like material be detected in the β-endorphin neurons. These results suggest that—in contrast to the cells in the intermediate lobe of the pituitary—the β-endorphin in brain neurons is not α-N-acetylated and that the small amount of α-N-acetyl β-endorphin which can be found in extracts of brains from normal animals is probably of pituitary origin.