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Aims: The objective of this study was to examine the involvement of endogenous opioid peptides and opioid receptor (OR) subtypes in the cardioprotective effect of adaptation to chronic hypoxia in rats., Main Methods: Rats were exposed to continuous normobaric hypoxia (CNH; 12% oxygen) for 3 weeks. Myocardial ischemia was induced by 20-min coronary artery occlusion followed by 3-h reperfusion in anesthetized open-chest animals. Various OR antagonists were administered to rats prior to ischemia. The size of myocardial infarction and the incidence of ischemic ventricular arrhythmias were assessed. Myocardial and plasma concentrations of opioid peptides (met-enkephalin, β-endorphin, and endomorphins) were determined., Key Findings: Adaptation to CNH significantly increased myocardial and plasma concentrations of opioids, potentiated their further elevation by ischemia/reperfusion, and reduced myocardial infarct size, but it did not affect the incidence of ischemic arrhythmias. The infarct size-limiting effect of CNH was abolished by OR antagonists naltrexone (non-selective), naloxone methiodide (non-selective peripherally acting), TIPP[ψ] (δ-OR), naltriben (δ2-OR), or CTAP (μ-OR), while BNTX (δ1-OR) and nor-binaltorphimine (κ-OR) had no effect., Significance: The results suggest that the infarct size-limiting effect afforded by adaptation to CNH is mediated by activation of peripheral δ2- and μ-ORs by elevated levels of endogenous opioid peptides., (© 2013.)

Although serotonin, serotonin uptake inhibitors and serotonin precursors (including tryptophan or 5-hydroxytryptophan) are known to have hypoglycemic action in rodents or human, it is not clear whether serotonin has hypoglycemic effect in streptozotocin-induced diabetic rats (STZ-diabetic rats). The aim of this study was to investigate the action of serotonin in regulating the plasma glucose STZ-diabetic rats. Plasma glucose, insulin, beta-endorphin and adrenaline were assessed after intraperitoneal administration of serotonin. Serotonin produced hypoglycemic effects without altering plasma insulin and adrenaline levels but increasing beta-endorphin level in STZ-diabetic rats. The glycogen content in soleus muscle was increased at 90 min after application of serotonin (0.3 mg/kg) in STZ-diabetic rats. Dihydroergotamine (non-selective 5-HT receptor blocker) and pimozide (5-HT(7) receptor blocker) abolished the hypoglycemic effect of serotonin in STZ-diabetic rats. Serotonin-induced hypoglycemic effect in association with the increase of beta-endorphin release was abolished in bilaterally adrenalectomized STZ-diabetic rats. In isolated adrenal gland of STZ-diabetic rats, the increase of beta-endorphin secretion in response to serotonin was reduced by either dihydroergotamine or pimozide. Pretreatment with naloxone (1.0 mg/kg, i.p.) prevented serotonin-induced plasma glucose lowering effect in STZ-diabetic rats. The results demonstrated that serotonin may activate 5-HT(7) receptor on rat adrenal gland to enhance of beta-endorphin secretion, which then stimulates the opioid receptor to increase peripheral glucose utilization, resulting in decreased plasma glucose levels in STZ-diabetic rats.

The immunomodulatory properties of melatonin (Mel) are generally recognized but the mechanisms of its action are not fully understood. In mammals, some of the immunomodulatory effects of Mel are mediated by opioids synthesized by immune cells under its influence. The present study was performed to examine whether Mel-induced opioids are involved in the immunomodulatory activity of Mel in chickens. Experimental peritonitis was evoked by a single ip injection of thioglycollate (TG), and half of the birds were pre-treated with Mel. Some of the Mel-treated birds were additionally pre-treated with naltrexone, an antagonist of opioid receptors. Control birds received an injection of saline, Mel or were untreated. At specific post-injection intervals chickens were sacrificed, the peritoneal cavity was flushed out and peritoneal leukocytes (PTLs) were counted. The activity of PTLs was measured in vitro by the level of reactive oxygen species (ROS). Splenocytes were isolated aseptically and mitogen-stimulated in vitro proliferation was assessed. In PTLs and splenocytes the expression of opioid (proopiomelanocortin and proenkephalin) genes was also examined. Mel exerted a bi-phasic effect on TG-induced peritonitis in chickens: initially it blocked the development of peritonitis, decreasing the number of PTLs and intracellular ROS level (anti-inflammatory action), and thereafter an increase in both PTL number and ROS level was observed (pro-inflammatory action). The pro-inflammatory effect occurred a few hours after the induction of expression of the proenkephalin gene in PTLs and both the proenkephalin and proopiomelanocortin genes in splenocytes. These effects were prevented by naltrexone, suggesting involvement of the opiatergic mechanism.

The main biological role of angiotensin II type 2 receptor (AT2) has not been established. We made use of targeted disruption of the mouse AT2 gene to examine the functional role of the AT2 receptor in the central nervous system (CNS). We have previously shown that AT2-deficient mice displayed anxiety-like behavior in comparisons with wild-type mice. In the present study, we analyzed the pain threshold, learning behavior and brain edema formation using the tail-flick test, the tail-pinch test, the passive avoidance task and cold injury, respectively. In the passive avoidance task and cold injury, no differences were found between wild-type mice and AT2-deficient mice. In contrast, the pain threshold was significantly lower in AT2-deficient mice, compared with findings in wild-type mice. The immunohistochemical distribution of beta-endorphin in the brain was analyzed quantitatively in AT2-deficient mice and wild-type mice, using microphotometry. The fluorescence intensity of beta-endorphin in the arcuate nucleus of the medial basal hypothalamus (ARC) was significantly lower in AT2-deficient mice, compared with findings in wild-type mice. We found that the AT2 receptor does not influence learning behavior and brain edema formation. As AT2-deficient mice have increased sensitivity to pain and decreased levels of brain beta-endorphin, AT2 receptors may perhaps mediate regulation of the pain threshold.

The aim of the present study was to investigate the effect of somatostatin administration in arthritic rats. Inflammation was induced by daily interplantar injection of 100 microl of Freund's complete adjuvant into the left hind paw of the rat. Arthritis developed 20 days following the first injection and was stable in the inoculate paw. Arthritic rats were treated interplantarly with somatostatin (5 or 10 microg) or with indomethacin (100 microg) daily for 14 days. Inflammatory response was studied at 12 h, 7 and 14 days following drug administration. The effect of somatostatin was determined by local (into popliteal lymph nodes) and systemic production of beta-endorphin. Our results showed that somatostatin treatment significantly increased beta-endorphin levels in the blood and lymphocytes from popliteal lymph nodes. Greater efficiency was seen when 5 microg instead of 10 microg of somatostatin was used. A significant decrease of absolute leukocytosis was observed at the 14th day following somatostatin administration. Moreover, a significant reduction of plasmatic beta-globulins at 12 h and the 7th day and of plasmatic alpha2-globulins at the 14th day was observed after the beginning of somatostatin treatment.

Previous studies from our laboratory have characterized a naloxone-insensitive beta-endorphin (beta-End) receptor on the human pro-monocytic cell line U937. Since monocytes are macrophage precursors, we sought to identify and characterize this site on fully differentiated effector macrophages. Mice (ICR females, 6-8 wk old) were injected (i.p.) with 1 mL of thioglycollate to induce an inflammatory response. Elicited cells were harvested 3 d later by lavage. Macrophages were enriched by adherence and analyzed via radioreceptor assay (with [125I] beta-End, 2,000 Ci mmol-1) as either intact cells or membrane preparations. Scatchard analysis revealed a single saturable binding site for beta-End (Kd = 9.75 +/- 2.6 x 10(-9) M; 8218 +/- 2360 sites/cell). Competition studies showed that other opiate receptor ligands including naloxone, DAMGO, U69593, or 2,5 DPDP-enkephalin were ineffective at displacing [125I] beta-End when compared to unlabeled beta-End. Analysis of competition studies utilizing fragments and analogs of beta-End revealed that beta-End (6-31) and beta-End (1-5, 16-31) were equipotent, and N-acetylated beta-End was less potent, than beta-end (1-31) in displacing [125I] beta-End binding. In contrast, beta-End (1-27) and beta-End (28-31) were ineffective. In summary, we have identified a naloxone-resistant beta-End binding site on murine peritoneal macrophages that is similar to one we have previously characterized on U937 cells and cultured murine splenocytes.

The present study was conducted to evaluate the effects of caffeine and the benzodiazepine agonist diazepam, and a combination of both on beta-endorphin (beta-EN) levels in specific rat brain regions. Male Sprague-Dawley rats (150-200 g) adapted to a 12-hour light: 12-hour dark illumination cycle were used in this study. Caffeine (10 mg/kg), diazepam (2 mg/kg) or a combination of caffeine (10 mg/kg) and diazepam (2 mg/kg) were administered intraperitoneally to rats at 11:00 hr. Control animals were injected with saline. Animals were sacrificed by decapitation 1 h after injection, the brains were immediately removed; the cortex, hippocampus, hypothalamus and midbrain were dissected and their B-EN levels measured by radioimmunoassay. Caffeine administration significantly increased B-EN levels in the cortex. Similarly, administration of diazepam alone resulted in a significant increase of B-EN levels in cortex. However, concurrent administration of diazepam and caffeine resulted in higher increase of B-EN levels in cortex. No significant changes in B-EN levels were detected in hippocampus and midbrain after administration of either caffeine or diazepam alone. On the other hand, when diazepam and caffeine were concurrently administered a significant increase of B-EN levels were observed in the midbrain. Moreover, administration of diazepam alone resulted in a significant increase of B-EN levels in hypothalamus. This increase was still observed following concurrent administration of diazepam and caffeine. These results clearly indicate that diazepam alters caffeine-induced effects on B-EN in specific rat brain regions.

In the present study, we determined the effect of acute and chronic nicotine treatments on the secretion of immunoreactive beta-endorphin (IR-beta-EP) and cell viability of cultured hypothalamic neurons. Also, we determined the secretory response of IR-beta-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 microM) for 6 h (acute treatment) or treated with nicotine at 12 h intervals for 96 h (chronic treatment). Determination of IR-beta-EP concentrations in the media revealed that 6-18 microM doses of nicotine increased IR-beta-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 beta-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 microM) treatments revealed that the hypothalamic neurons secrete elevated amounts of IR-beta-EP for a period of 72 h after nicotine withdrawal. These results suggest that: 1) acute treatment with nicotine stimulates hypothalamic IR-beta-EP release; 2) chronic nicotine treatment desensitizes beta-EP-secreting neurons and, 3) beta-EP neurons in primary culture show withdrawal response to nicotine.

The intermediate lobe of the mammalian pituitary is highly responsive to dopamine inhibition of beta-endorphin secretion. In this study, the ability of aged (12 months) intermediate lobes to respond to dopamine was compared to that of young (6 weeks) tissue, using a short-term in vitro incubation of isolated rat neurointermediate lobes, with measurement of peptide release by radioimmunoassay. Tissue from the aged rats released greater amounts of beta-endorphin peptide than amounts measured from young tissue at all time periods studied. The aged lobes were also found to be significantly more sensitive to dopamine than young glands, as measured by percent change of each group compared to respective baseline release. In comparison, incubation of tissue from young animals in which the intermediate lobe had been acutely denervated by treating rats with injections of the catecholamine neurotoxin, 6-hydroxydopamine, did not differ in responsiveness to dopamine as compared to tissue from control rats. The observations suggest that aging intermediate lobe, while being hypersecretory, is supersensitive to dopamine, perhaps as the result of gradually reduced innervation.

Previous reports have demonstrated that beta-endorphin stimulates the clonal growth of human small cell lung carcinoma (SCLC) cell lines. In this study, the human SCLC lines, NCI-H69, NCI-H345, and NCI-N417, were observed to be highly-enriched in saturable, high-affinity binding sites which are labeled by [125I]beta-endorphin. In contrast to conventional opioid receptors, [125I]beta-endorphin SCLC binding was insensitive to naloxone and other mu, delta, or kappa opioid ligands. Further analysis of the NCI-H69 cells demonstrated that specific (naloxone-insensitive) binding was dependent on receptor concentration, reversible, sensitive to sodium ion, but insensitive to the GTP analogue, Gpp(NH)p. These results suggest a role for naloxone-insensitive beta-endorphin in modulating SCLC metabolism.

Testicular function is sensitive to chemical and thermal stresses. To investigate the effects of small temperature changes on CRH-stimulated beta EP release, we employed TM3 cells, a mouse prepubertal Leydig cell line that secretes ir beta EP. To monitor beta EP secretion from these cells we used the reverse hemolytic plaque assay. After 3.5 hr incubation of cells with hormone, the EC50 of the CRH dose-response curve at 34 degrees C and 37 degrees C were 0.1 nM and 1 nM, respectively. For comparison, we also investigated the effect of temperature on CRH-stimulated beta EP release from a non-testicular cell line, AtT-20, a mouse anterior pituitary cell line. Using radioimmunoassay to measure ir beta EP levels in the media of AtT-20 cells, the EC50s for the CRH dose-response curve at 34 degrees C and 37 degrees C were 0.2 nM and 2 nM, respectively, at 1 h. After 3.5 h this temperature dependent difference in EC50 was still observed. These results suggest that CRH receptors or post-receptor actions in Leydig cells and anterior pituitary corticotropes are sensitive to small temperature changes.

Administration of Hypnorm, an anaesthetic containing the known mu-opiate receptor agonist fentanyl, elicited dose- and time-related elevation of plasma ACTH, beta-endorphin and corticosterone levels in 10-day old rat pups. Pretreatment with specific antibodies (raised against CRH, AVP and ACTH resp.) revealed that Hypnorm administration activated the ACTH-corticosterone system in the 10-day old rat and its effect is mediated by CRH and/or AVP. Hypnorm anaesthesia was associated with significant decrease in the ACTH and beta-endorphin levels in the pituitary lobes as well as in beta-endorphin content of the hypothalamus and medulla oblongata. Latter results may indicate that the beta-endorphinergic system in the brain of the 10-day old rat is activated by Hypnorm, an effect most probably elicited by the opiate agonist fentanyl.

The purpose of this study was to immortalize porcine endometrial cells and to characterize the transformed cells. Primary porcine endometrial cells were transfected with the plasmid vector (pmk16) containing SV40 DNA using a liposome-mediated method. The viral DNA was from a replication-defective, origin-minus, temperature-sensitive mutant strain (A58). One clone, designated PE-1, has been propagated for over 120 passages. PE-1 cells grown at 33C (33C cells) exhibit spindle-shaped morphology; when cultured at 40C (40C cells), they took on a polygonal or spherical shape. Morphology of 40C cells returned to the spindle shape after culture flasks were shifted back to 33C. During a 2-week period, 33C cells propagated approximately 30-fold faster than 40C cells, whereas protein concentration was higher in 40C cells. Southern blot analysis of PE-1 cells demonstrated successful integration of the ts-SV40 DNA sequence into the porcine endometrial cells, possibly at multiple sites. The presence of cytokeratin on PE-1 cell membranes was shown by immunocytochemical studies, suggesting that the PE-1 cell clone was of epithelial origin. Reverse phase (RP)-HPLC analysis of PE-1 cell extract indicated that the majority of immunoreactive beta-endorphin (ir-BEND) eluted with a hydrophobicity similar to that of synthetic BEND and alpha-N-acetylated BEND (Nac-BEND). These results demonstrate that a porcine endometrial cell line has been established, and that this cell line possesses characteristics of temperature sensitivity in cell morphology, growth rate, and protein synthesis.

Dopamine (DA) neurons participate in tonic inhibition of prolactin (PRL), whereas beta-endorphin (beta-End) and serotonin (5-HT) neurons appear to be important stimulatory links for nocturnal PRL surges that occur throughout the first half of pregnancy in the rat. The purpose of this study was to determine how these neuronal components might be organized within the pathway controlling PRL release during gestation. Maximal stimulation of DA receptors with the agonist bromocriptine mesylate (Bromo) completely blocked the PRL response to beta-End (100 ng/microliters/min for 15 min) given intracerebroventricularly (i.c.v.) on day 8 of pregnancy. DA receptor blockade, produced by implanting a 25 mg pellet of haloperidol (Hal) on day 7 of pregnancy, resulted in PRL levels of 500-600 ng/ml by the following morning. beta-End i.c.v. or 250 mg/ml/kg BW of the DA synthesis inhibitor, alpha-methyl-p-tyrosine (alpha-MPT), given during the intersurge period, were equally effective in significantly increasing PRL (p less than 0.01) above pretreatment levels. beta-End and alpha-MPT evoked similar increases in rats pretreated with Hal, suggesting the stimulatory effect of beta-End on nocturnal PRL surges may primarily be due to DA inhibition. The next objective was to determine how beta-End and 5-HT might interact to stimulate the nocturnal surge. Day 8 pregnant rats were infused continuously with the opioid receptor blocker, naloxone hydrochloride (Nal), at a rate of 2.0 mg/10 min from 1000-1300 h. The PRL response to an injection of 20 mg/kg BW 5-hydroxytryptophan (5-HTP) at 1200 h was greatly attenuated, compared to controls infused with saline instead of Nal. This suggests that 5-HT stimulates PRL, at least in part, by an action at opioid receptors. Distilled H2O or 10 mg/kg BW of the selective S2 receptor blocker, ketanserin tartrate (Ket), was given intraperitoneally (i.p.) during the intersurge period on day 8 of pregnancy. All animals demonstrated an identical response to beta-End given 2 hours later, regardless of the type of pretreatment. It appears that beta-End does not stimulate PRL by way of an S2 receptor. Although beta-End induced a significant increase in PRL on day 16 of pregnancy, the response was attenuated by more than 60% compared to the response on day 8 of pregnancy. This attenuation may involve placental lactogens, shown to be secreted during this time and to inhibit PRL secretion.(ABSTRACT TRUNCATED AT 400 WORDS)

The mouse neuroblastoma x rat glioma hybrid NG108-15 was previously shown to express delta opioid receptors. Because neuroblastoma cells display different phenotypes and cloned cell lines are heterogenous, we studied the characteristics and distribution of human 125I-beta-endorphin (125I-beta E) binding sites in cultures of NG108-15 cells with the use of micro-autoradiography and light microscopy. 125I-beta E labeled delta sites in NG108-15 in the presence of the non-opioid blocking peptide, beta-endorphin (6-31) (beta E (6-31)). Silver grains resulting from 125I-beta E binding to the opioid sites occurred in diffuse patches over several cells, with preferential location in dense cell patches. Pretreatment of NG108-15 with the delta agonist DADLE, previously shown to decrease beta E binding to delta sites on intact cells, also reduced silver grain density; however, some cells located in dense cell clusters were resistant to substantial agonist induced loss of labeling. These results suggest that delta opioid binding has a heterogenous cellular distribution in NG108.

We have measured mu and delta opioid receptor sites on intact SK-N-SH and NG108-15 neuroblastoma cells, respectively, in culture. Use of 125I-beta-endorphin (beta E) as a tracer, together with beta E(6-31) to block high-affinity non-opioid binding in both cell lines, permitted the measurement of cell surface mu and delta opioid receptor sites. Labeling was at delta sites in NG108-15 cells and predominantly at mu sites in SK-N-SH cells. Pretreatment with the mu and delta agonist, DADLE, caused a rapid loss of cell surface delta receptor sites in NG108-15 cells, but failed to reduce significantly mu receptor density in SK-N-SH cells.

After 125I-beta-endorphin was intravenously injected to rats, an autoradiographic study of distal femur articular cartilage was performed. Results show a specific binding of 125I-beta-endorphin to chondrocytes, suggesting the possible existence of an opiate modulation of articular cartilage.

To characterize the effect of ethanol on the hypothalamic beta-endorphin-containing neurons, rat fetal hypothalamic neurons were maintained in primary culture, and the secretion of beta-endorphin (beta-EP) was determined after ethanol challenges. Constant exposure to ethanol at doses of 6-50 mM produced a dose-dependent increase in basal secretion of beta-EP from these cultured cells. These doses of ethanol did not produce any significant effect on cell viability, DNA or protein content. The stimulated secretion of beta-EP following constant ethanol exposure is short-lasting. However, intermittent ethanol exposures maintained the ethanol stimulatory action on beta-EP secretion for a longer time. The magnitude of the beta-EP response to 50 mM ethanol is similar to that of the beta-EP response to 56 mM of potassium. Ethanol-stimulated beta-EP secretion required extracellular calcium and was blocked by a calcium channel blocker; a sodium channel blocker did not affect ethanol-stimulated secretion. These results suggest that the neuron culture system is a useful model for studying the cellular mechanisms involved in the ethanol-regulated hypothalamic opioid secretion.

Corticotropin-releasing factor (CRF), a 41-aminoacid neuropeptide, can induce lymphocytes to production of beta-endorphin (beta E). Furthermore, the neuropeptide Arginine-Vasopressin (AVP) can enhance CRF-induced production of beta E. We have demonstrated that CRF acts by stimulating monocytes to production of the cytokine interleukin-1 (IL-1). IL-1 can in its turn activate the lymphocytes to secretion of beta E. Here we demonstrate that the glucocorticoid analogue dexamethasone is capable of modulating CRF-induced beta E secretion by lymphocytes. It appeared that dexamethasone can inhibit secretion of lymphocyte-derived beta E. The mechanism by which dexamethasone exerts its inhibitory activity is by blocking CRF-induced production of IL-1, thereby preventing induction of beta E secretion by B cells. These results support the concept that peptide hormones and glucocorticoids are mediating a reciprocal modulation of neuroendocrine and immunological activities.

The significance of beta-endorphin for drug dependence was explored by measuring the levels of beta-endorphin-immunoreactivity (beta E-IR) in plasma and parts of pituitary and brain of rats self-administering heroin or cocaine as compared to animals offered saline. Rats that had intravenously self-administrated heroin for 5 consecutive daily sessions of 6 h, and were decapitated immediately after the last session, showed a decreased concentration of beta E-IR in the anterior lobe (AL) of the pituitary while rats that had taken cocaine showed a decreased concentration of beta E-IR in the septum. Rats that had self-administrated heroin or cocaine and were decapitated 18 h after the last session, showed an increased concentration of beta E-IR in plasma and decreased concentrations in the AL of the pituitary and in specific areas of the brain limbic system, i.e. nucleus accumbens, septum, hippocampus and rostral striatum. The finding that self-administration of both the opiate heroin, inducing psychic and physical dependence, and the non-opiate cocaine, inducing psychic but not physical dependence, is accompanied by similar changes in beta E-IR concentrations particularly in limbic brain structures, and that these effects are present 18 h but not immediately after the last session, suggests that beta E and related peptides in limbic brain regions may represent a neurochemical correlate for psychic dependence on drugs.

The present study examines whether two treatments known to induce refractoriness to exogenous morphine produce this desensitization through a change in the posttranslational processing of brain beta-endorphin (beta-End). The first experiment examined whether an ovarian steroid regimen which produces a transient desensitization of brain opiate receptor mechanisms alters beta-End processing in the preoptic area (POA), medial basal hypothalamus (MBH), and brainstem (BS). The second experiment monitored the effects of morphine pellet treatment, known to produce morphine dependency, on immunoreactive beta-End forms in the hypothalamus and periaquaductal gray area of the midbrain (PAG). The individual molecular forms of beta-End were separated using ion exchange chromatography and collection fractions were quantitated for beta-End immunoreactivity by RIA. The results show that regional differences occur in the posttranslational processing of beta-End. In the hypothalamus, MBH and POA, beta-End-(1-31) and its non-acetylated C-terminal cleavage products, beta-End-(1-27) and beta-End-(1-16) were the predominant forms of beta-End. The PAG pools produced a beta-End peptide elution profile similar to the hypothalamus, although small amounts of N-acetyl-beta-End-(1-31) were also identified. The BS exhibited the least posttranslational processing of beta-End; beta-End-(1-31) was the primary product with smaller amounts of beta-End-(1-27) and beta-End-(1-26) observed. However, neither ovarian steroid treatment nor chronic morphine produced any changes in posttranslational processing of beta-End or in total beta-End concentration in any of the brain regions examined in these experiments. These data indicate that the refractoriness or tolerance to exogenous morphine associated with steroid or chronic morphine treatment cannot be explained by alterations in the biological activity of beta-End resulting from the differential regulation of its posttranslational processing products.

The concentrations of alpha-, beta- and gamma-endorphin were determined by radioimmunoassay in HPLC fractionated extracts of post mortem hypothalamic tissue obtained from schizophrenic patients and controls. The hypothalamic concentration of alpha- and gamma-endorphin was significantly higher in patients than in controls (+72.9% and +50.5% respectively). No difference was found in the concentration of beta-endorphin, the putative precursor of alpha- and gamma-endorphins. These results suggest a deviant metabolism of beta-endorphin in the brain of schizophrenic patients. Whether this phenomenon is related to the psychopathology, or is a consequence of ante mortem farmacotherapy, remains to be established.

The effect of chronic administration of morphine and abrupt and naloxone-precipitated withdrawal on the levels of beta-endorphin and methionine-enkephalin in spleen, adrenals and thymus glands of Sprague-Dawley rats was determined. Rats were made tolerant to and dependent on morphine by subcutaneous implantation of 6 morphine pellets (75 mg morphine in each) during a 7-day period. The tolerant-dependent (with pellets intact) and abstinent (pellets removed 18 hours earlier) rats were sacrificed. In another group, rats with pellets intact were injected with naloxone and sacrificed 10 min later (precipitated abstinence). The weights of the tissues under any of the above treatments did not change nor did the levels of methionine-enkephalin and beta-endorphin in adrenals. The level of beta-endorphin was elevated in the spleen and thymus of morphine tolerant-dependent rats, while the levels of methionine-enkephalin in rats undergoing abrupt or naloxone-precipitated abstinence were significantly higher than in their respective placebo controls. The levels of methionine-enkephalin in the thymus gland of rats with placebo and morphine pellets left intact did not differ. It is concluded that in morphine tolerant-dependent rats the levels of beta-endorphin in spleen and thymus are elevated. During abrupt and naloxone-precipitated abstinence, the levels of methionine-enkephalin in the thymus gland are significantly elevated possibly due to an inhibition of their release. Since these opioid peptides have been implicated in immunomodulation, and alterations were seen in organs controlling immune function, the present results may be helpful in explaining altered immune function in morphine dependent and abstinent states.

We have investigated the ontogeny of immunoreactive beta-endorphin (i-beta E) in the testes of rats from 5 to 150 days of age. i-beta E was measured by RIA in acid extracts of decapsulated testes and characterized by gel filtration chromatography. Significant age-related differences in both the levels and type of i-beta E were observed. Total levels of i-beta E in the testes were very low and barely detectable from 5-20 days of age, but rose sharply in parallel with testes weights from 20-60 days of age; thereafter, no significant changes in i-beta E were found through 150 days of age. Concentrations of i-beta E, expressed in pmol/g testis, fell precipitously between days 5 and 10 and remained relatively constant from 10-150 days. Most of the i-beta E at 5 and 15 days chromatographed like authentic beta-endorphin. However, with the onset of puberty (30-35 days) and during sexual maturation, much of the total i-beta E chromatographed like its' precursor beta-lipotropin (beta LPH). Hypophysectomy decreased the weight and total i-beta E levels of the testes to the same extent without altering the concentrations of i-beta E or the chromatographic pattern of i-beta E. These results indicate that beta E-like and beta LPH-like peptides are present in the rat testis and that age-related changes in both the levels and type of i-beta E correlate with various structural and functional aspects of testicular development.

Immunoreactive beta-endorphin (IR-BE) was measured in the plasma, anterior pituitary (AP), neurointermediate lobe of the pituitary (NIL) and hypothalamus of male rats treated chronically (once daily for ten days) with cocaine. Cocaine produced a consistent elevation in the concentration of IR-BE in the plasma, the AP and the NIL at doses of 2.5 - 20 mg/kg/ip. The release of IR-BE from the AP and the NIL was determined in vitro and was found to be increased by treatment with cocaine. Chronic administration of cocaine did not affect the concentration of IR-BE in the hypothalamus. Chromatographic analysis revealed that cocaine produced a slight decrease in the amount of beta-endorphin relative to beta-lipotropin in the AP. Beta-endorphin was the major form of IR-BE released by the AP and the sole constituent and secretory product of the NIL. These data indicate that chronic administration of cocaine stimulates the endogenous opiate system, elevating the levels of IR-BE in the pituitary and promoting beta-endorphin release.

The equilibrium dissociation constants and maximal binding capacities of 3H-dihydromorphine (DHM), 3H-D-Ala2-D-leu3-enkephalin (DADL), and 3H-dynorphin A(1-8) for their respective mu, delta, and kappa opiate binding sites were studied in brain membrane preparations from lean and genetically obese-hyperglycaemic (Aston ob/ob) mice. The concentration of kappa binding sites was 2.7 fold higher in obese compared with lean mouse brain (231 +/- 44.6 versus 83.8 +/- 10.3 fmoles 3H-dynorphin/mg protein respectively, mean +/- SEM). The concentration of delta binding sites in obese was 1.6 fold that in lean mouse brain (94.5 +/- 8.6 versus 59.5 +/- 6.5 fmoles 3H-DADL/mg protein). In contrast, the concentration of brain mu receptors was 40% lower in obese compared with lean mice (20.8 +/- 2.19 and 34.8 +/- 3.1 fmoles 3H-DHM/mg protein respectively). Binding affinities of delta and kappa sites for their respective ligands were not significantly different in lean v. obese mice. However, for mu sites, lean mouse binding data showed two affinities, one was not significantly different from obese (0.35 nM) the second was lower (1.18 nM) for DHM. Increases of approximately 5 fold and 3 fold in the brain content of beta-endorphin and met-enkephalin respectively, and no differences in brain dynorphin levels, were demonstrated in obese mice compared with lean controls. In obese mice, pituitary beta-endorphin content was 9 fold higher, met-enkephalin 4 fold higher and dynorphin 12 fold higher than in lean mice. The striking differences in opioid binding-site characteristics and in endogenous opioid peptide levels in obese compared with lean mice may contribute to the hyperphagia and, directly or indirectly, to the development of hyperglycaemia and hyperinsulinaemia in obese mice.

An in vitro perifusion system was used to investigate the effects of GnRH stimulation on LH, ACTH, and immunoreactive beta-endorphin (i beta-END) release from ovariectomized (1 week) rat anterior hemipituitaries. Either 0, 8 or 80 nM GnRH was administered as a 15 min pulse followed 30 min later by a prolonged 45 min infusion. Both 8 and 80 nM GnRH induced comparable LH release in response to the 15 min as well as the 45 min GnRH stimulation. The initial 15 min exposure to either 8 or 80 nM GnRH did not induce significant changes in ACTH or i beta-END release. In contrast, the subsequent 45 min exposure to 8 nM GnRH induced a significant (p less than 0.01) increase in ACTH release, and the 45 min exposure to 80 nM GnRH induced a significant (p less than 0.01) increase in ACTH as well as i beta-END release. Equimolar (i.e. 8 or 80 nM) GnRH receptor antagonist (ANT) blocked the stimulatory effects of GnRH in all cases. These results demonstrate that GnRH can stimulate not only LH but also ACTH and i beta-END release from ovariectomized rat anterior hemipituitaries in vitro, apparently by a GnRH receptor mediated mechanism independent of actual LH release. Although the time course of these responses appears to be consistent with the hypothesis that GnRH-stimulated gonadotropes release paracrine factor(s) which stimulate corticotrope activity, the mechanism of these responses remains to be determined.