Opiate receptor subtypes in the regulation of thyrotropin and prolactin secretion in the rat

In the present study both MR 2034 (kappa-agonist) and DAMME (mu-agonist) decreased thyrotropin (TSH) secretion stimulated by cold in the rat when infused into the 3rd ventricle. After infusion into the posterior hypothalamus (PH), a small dose of MR 2034 increased the TSH response to cold whereas other doses did not. The stimulatory (at PH) but not the inhibitory (at 3rd ventricle) effect of MR 2034 was antagonized by naloxone. DAMME had no statistically significant effect at this location. Both the mu- and kappa-agonist stimulated prolactin secretion when infused into the 3rd ventricle, but DAMME was more effective than MR 2034. Furthermore, the stimulatory effect of DAMME, but not that of MR 2034, on prolactin secretion was antagonized by naloxone.     

Endocrine actions of opioids

The widespread occurrence of opioid peptides and their receptors in brain and periphery correlates with a variety of actions elicited by opioid agonists and antagonists on hormone secretion. Opioid actions on pituitary and pancreatic peptides are summarized in Table 1. In rats opioids stimulate ACTH and corticosterone secretion while an inhibition of ACTH and cortisol levels was observed in man. In both species, naloxone, an opiate antagonist, stimulates the release of ACTH suggesting a tonic suppression by endogenous opioids. In rats, a different stimulatory pathway must be assumed through which opiates can stimulate secretion of ACTH. Both types of action are probably mediated within the hypothalamus. LH is decreased by opioid agonists in many adult species while opiate antagonists elicit stimulatory effects, both apparently by modulating LHRH release. A tonic, and in females, a cyclic opioid control appears to participate in the regulation of gonadotropin secretion. Exogenous opiates potently stimulate PRL and GH secretion in many species. Opiate antagonists did not affect PRL or GH levels indicating absence of opioid control under basal conditions, while a decrease of both hormones by antagonists was seen after stimulation in particular situations. In rats, opiate antagonists decreased basal and stress-induced secretion of PRL. Data regarding TSH are quite contradictory. Both inhibitory and stimulatory effects have been described. Oxytocin and vasopressin release were inhibited by opioids at the posterior pituitary level. There is good evidence for an opioid inhibition of suckling-induced oxytocin release. Opioids also seem to play a role in the regulation of vasopressin under some conditions of water balance. The pancreatic hormones insulin and glucagon are elevated by opioids apparently by an action at the islet cells. Somatostatin, on the contrary, was inhibited. An effect of naloxone on pancreatic hormone release was observed after meals which contain opiate active substance. Whether opioids play a physiologic role in glucose homeostasis remains to be elucidated.     

The met-enkephalin analog FK 33-824 directly inhibits ACTH release by the rat pituitary gland in vitro

Systematic administration of the enkephalin analog FK 33-824 was previously shown to stimulate PRL secretion and to inhibit ACTH secretion in man. Naloxone prevented the effect on PRL release, but not on ACTH release. In this study, the direct action of this analog on hormone release by rat anterior pituitary lobes $\text{in}$$\text{vitro}$ were investigated. 1 uM FK 33-824 inhibited basal ACTH secretion by anterior pituitary glands in vitro, while 0.1 uM and 1 uM attenuated the lysine vasopressin stimulated ACTH release. Naloxone did not reverse the inhibitory action of the analog on ACTH release. β-Endorphin (0.01 - 1 uM) did not directly affect ACTH release. Basal and dopamine-induced inhibition of PRL release by anterior pituitary glands was neither influenced by FK 33-824 (0.1 and 1 uM), nor by β-endorphin (0.1 and 1 uM) with or without bacitracin. This study shows that the long-acting met-enkephalin analog FK 33-824 differentially affects PRL and ACTH secretion by the pituitary gland. It seems to stimulate PRL release at a suprapituitary site and this action probably involves u opiate receptors, because naloxone prevents these stimulatory effects. The inhibitory effect of FK 33-824 on ACTH release, however, is mediated via a direct effect at the pituitary level, which does not involve u receptors, as naloxone did not prevent this effect. In this respect, its action differs from that of β-endorphin, which does not directly affect ACTH release by the anterior pituitary gland.     

Effect of central naloxone on hormone and blood pressure responses to hemorrhage in conscious sheep

The role of the brain opioid system in the control of hypothalamic-pituitary-adrenal activity was studied in 10 conscious sheep with an indwelling cannula in a cerebral lateral ventricle. On separate days, sheep received infusions of artificial CSF (control) and the opiate antagonist, naloxone (100 micrograms/hr) before and during acute moderate hemorrhage (15 ml/kg over 10 min). Infusion of naloxone before hemorrhage raised plasma ACTH and resulted in a significant increase in cortisol compared to the control infusion. In contrast, ACTH and cortisol responses to hemorrhage tended to be blunted by central naloxone infusion. The responses of vasopressin, aldosterone and the catecholamines remained unaffected by naloxone. The fall in blood pressure and the rise in heart rate accompanying hemorrhage were likewise unaltered. These results suggest that brain opioid peptides have an inhibitory effect on basal ACTH secretion but do not play a major role in modulating the hemodynamic or pituitary-adrenal responses to acute moderate hemorrhage in conscious sheep.     

Stereospecific increase by narcotic antagonists of evoked acetylcholine output in guinea-pig ileum.

In the myenteric plexus-longitudinal muscle preparation of the guinea-pig ileum, naloxone (30–100 nM) increases the output of acetylcholine evoked by electrical field stimulation at 0.017 Hz and to a lesser extent also at 10 Hz. The stereospecific requirements for this effect were studied with three pairs of optical isomers of antagonists of the benzomorphan series. The (−)-isomer of β-9-methyl-5-phenyl-2-allyl-2′-hydroxy-6,7-benzomorphan (GPA 1843) which had no agonist activity, had an effect similar to naloxone whereas the (+)-isomer was inactive in this respect. The (−)-isomer of antagonists with even weak agonist activity gave variable results. It is assumed that naloxone antagonises the action of enkephalin which has been shown to be present in the guinea-pig ileum. It is recommended to establish the stereospecificity of an antagonist action in order to exclude pharmacological effects not due to interaction with opiate receptors.     

Failure of vasopressin to prevent the natruiretic and diuretic response to unilateral stimulation of carotid baroreceptors.

Unanesthetized rats treated with deoxycorticosterone acetate were continuously infused with a maximally effective dose of vasopressin (50 muU/min per 100 g). After a control period of 2 h the animals were subjected to a 2 h period of unilateral stimulation of the carotid baroreceptors. There was a large natriuretic response accompanied by diuresis and a fall in urine osmolality. It was concluded that neither the natriuretic nor the diuretic response could be explained by inhibition of vasopressin secretion. Analysis of kidney tissue indicated that the diuresis was associated with partial dissipation of the inner medullary concentration gradient.     

Naloxone does not antagonize PCP-induced stimulation of the pituitary-adrenal axis in the rat

Phencyclidine (PCP) has been shown to stimulate the pituitary-adrenal axis in the rat. The purpose of the present study was to determine whether opiate receptors are involved in this effect by testing whether pretreatment with the opiate antagonist naloxone can antagonize PCP-induced ACTH and corticosterone release. PCP (10.0 mg/kg) produced increases in plasma ACTH and corticosterone 60 min after s.c. administration. Pretreatment with naloxone (2.0 mg/kg s.c.) did not reduce the rise in plasma levels of ACTH or corticosterone produced by PCP. These results indicate that naloxone-sensitive opiate receptors are not involved in the PCP-induced stimulation of the pituitary-adrenal axis in rats.     

Stereospecific reversal of nitrous oxide analgesia by naloxone

The opiate antagonist naloxone was found to block nitrous oxide analgesia in a stereospecific fashion. Using a modified hotplate test in mice, the (-)-enantiomer of naloxone (which has a KD of approximately 1 nM for opiate receptors) antagonized the analgesic actions of nitrous oxide in a dose-dependent (2.5-20 mg/kg) fashion. In contrast, the (+)-enantiomer (KD approximately 10,000 nM) had no effect on nitrous oxide analgesia at the highest dose tested (40 mg/kg). These data strongly suggest that nitrous oxide analgesia is mediated via opiate receptors and is consistent with the hypotheses that this effect occurs either through the release of endogenous opioids or by physical perturbation of the opiate receptors.     

On the site of action of naloxone-stimulated cortisol secretion in gilts

The increase in serum cortisol concentrations following naloxone administration to female pigs was abolished by hypophysial stalk-transection, even though CRH and ACTH stimulated cortisol release in these animals. We suggest that the opioid antagonist enhances cortisol secretion primarily by a central action in pigs.     

Effects of intracerebroventricular administration of opiate receptor antagonists on the suppressed pulsatile LH release during acute fasting in ovariectomized estradiol-treated rats.

The involvement of specific opiate receptors in the suppression of LH release during acute fasting in ovariectomized estradiol-treated rats was examined by intracerebroventricular (i.c.v.) administration of opiate receptor antagonists that exert a specificity directed mainly, although not absolutely, towards the delta-, kappa- or mu-opiate receptors. Fasting for 48 h significantly decreased mean plasma LH levels in estradiol-treated animals by increasing sensitivity to the negative feedback effect of estradiol. Injecting i.c.v. the mu-opiate receptor antagonist naloxone (10 or 100 nmol in 2 microliters of saline) blocked the inhibitory effect of fasting on pulsatile LH release and reinstated LH pulses. On the other hand, i.c.v. administration of the same dosages of a delta-opiate receptor antagonist ICI 174,864 or a kappa-opiate receptor antagonist WIN 44441-3 did not have any effect. These results suggest that the increased sensitivity of the LH-releasing mechanism to the negative feedback effect of estradiol during fasting involves the endogenous opioids mainly through the selective activation of the mu-opiate receptors.     

Interactions between CRF, epinephrine, vasopressin and glucocorticoids in the control of ACTH secretion

The secretion of ACTH by corticotrophs in the anterior lobe of the rat pituitary gland is under the stimulatory influence of at least three receptors, namely that for peptidic CRF (corticotropin-releasing factor), vasopressin and alpha 1-adrenergic agents. CRF is a potent stimulator of cyclic AMP accumulation as well as adenylate cyclase activity in the rat adenohypophysis, thus suggesting an important role of cyclic AMP as mediator of CRF action on ACTH secretion. Vasopressin causes a 2-fold increase of the stimulatory effect of CRF on ACTH release in rat anterior pituitary cells in culture. The potentiating effects of vasopressin on CRF-induced ACTH release are accompanied by parallel changes of intracellular cyclic AMP levels. Vasopressin, while having no effect on basal cyclic AMP levels, causes a 2-fold increase in CRF-induced cyclic AMP accumulation without affecting the ED50 value of CRF action. ACTH secretion is also stimulated by a typical alpha 1-adrenergic receptor. Epinephrine causes a marked stimulation of ACTH release which is additive to that of CRF. Epinephrine, in analogy with vasopressin, although having no effect alone on basal cyclic AMP levels, causes a marked potentiation of CRF-induced cyclic AMP accumulation. Glucocorticoids cause a near-complete inhibition of epinephrine-induced ACTH secretion within 4 h with the following order of ED50 values: triamcinolone acetonide (0.2 nM) greater than dexamethasone (1.0 nM) much greater than cortisol (11 nM) greater than corticosterone (22 nM). Similar effects are observed for CRF- and vasopressin-induced ACTH release. Although the activity of the pituitary-adrenocortical axis in the rat is highly dependent upon sex steroids, 17 beta-estradiol, 5 alpha-dihydrotestosterone and the pure progestin R5020 have no detectable effect on basal or epinephrine-induced ACTH release, thus illustrating the high degree of specificity of glucocorticoids in their feedback control of ACTH secretion. Moreover, glucocorticoids have no effect on CRF-induced cyclic AMP accumulation, thus indicating that their inhibitory effect is exerted at a step following cyclic AMP accumulation.     

Opiate receptor blockade and diurnal pituitary and adrenal hormone levels

Peripheral pituitary hormone levels exhibit circadian variations though the mechanism of these changes is unknown. In order to investigate the possible role of endogenous opiates in such changes we have studied the influence of opiate receptor blockade with naloxone (6.8 mg) on pituitary hormones in the morning and again in the evening in six normal male volunteers. Basal ACTH, cortisol, aldosterone and prolactin were higher in the morning than in the evening. Following naloxone at 0700h both ACTH and cortisol rose indicating a tonic inhibition of ACTH by endogenous opiates at that time. At 2230h cortisol rose following naloxone but ACTH did not, suggesting that endogenous opiates do not play an important role in the diurnal rhythm of this hormone and consistent with the suggestion that endogenous opiates can effect cortisol levels independently of their action on ACTH. Neither aldosterone nor prolactin were influenced by naloxone. In contrast TSH was unaffected by naloxone in the morning but fell in the evening (mean + SE decrement over 120 min -0.6 +/- 0.3 mU/l as compared with the control +0.6 +/- 0.4 mU/l; p less than 0.01). Thus, endogenous opiates probably tonically stimulates TSH levels in the evening when TSH may increase and possibly play a role in the circadian rhythm of TSH.     

ACTH and corticosterone response to naloxone and morphine in normal, hypophysectomized and dexamethasone-treated rats

The effect of opiate receptors blocker naloxone on ACTH and corticosterone secretion in normal, dexamethasone-treated and hypophysectomized rats was studied. A dose-related increase in plasma corticosterone level was found at 45 min after s.c. injection of naloxone in a dose range of 0.25-2.0 mg kg-1. The rise in plasma corticosterone was preceded by a slight increase in plasma ACTH. Acute morphine administration in a relatively low dose (6 mg kg-1 s.c.) induced a significant rise in both plasma ACTH and corticosterone levels. Dexamethasone treatment was followed by low basal corticosterone level, by total inhibition of the stress response and response to morphine injection, while the response to ACTH administration was normal. Under these circumstances as well as in rats 6 days after hypophysectomy, naloxone failed to increase plasma corticosterone levels. It is concluded that a direct stimulation of corticosteroid biosynthesis in adrenal cortex is not involved in the mechanism of naloxone-induced activation of pituitary-adrenocortical function.     

Studies on the Inhibitory Action of Opiate Compounds in Isolated Bovine Adrenal Chromaffin Cells: Noninvolvement of Stereospecific Opiate Binding Sites

In isolated bovine adrenal chromaffin cells, beta-endorphin, dynorphin, and levorphanol caused a dose-dependent inhibition of catecholamine (CA) secretion elicited by acetylcholine (ACh), with an ID50 of 50, 1.3, and 4.3 microM, respectively. The inhibition by the opiate compounds was specific for the release evoked by ACh and nicotinic drugs and was noncompetitive with ACh. Stereospecific binding sites for the opiate agonist [3H]etorphine were found in homogenates of bovine adrenal medulla (KD = 0.59 nM). beta-Endorphin, dynorphin, levorphanol, and naloxone were potent inhibitors of the binding of [3H]etorphine with an ID50 of 12, 0.4, 5.2, and 6.2 nM, respectively. However, [3,5-I2Tyr1]-beta-endorphin, [3,5-I2Tyr1]-dynorphin, and dextrorphan, three opiate compounds with no or little activity in the guinea pig ileum assay, were relatively ineffective in inhibiting the binding of [3H]etorphine (ID50 700, 600, and 10,000 nM, respectively). On the other hand, these three compounds were equipotent with beta-endorphin, dynorphin, and levorphanol, respectively, in inhibiting the ACh-evoked release of CA from the adrenal chromaffin cells (ID50 of 10, 1.5, and 6 microM, respectively). Inhibition of CA release was also obtained with naloxone (ID50 = 14) microM) and naltrexone (ID50 greater than 10(-4) M), two classical antagonists of opiate receptors, and this effect was additive to that of beta-endorphin. These data indicate that the opiate modulation of CA release from adrenal chromaffin cells is not related to the stimulation of the high affinity stereospecific opiate binding sites of the adrenal medulla. The physiological function of these sites remains to be determined.     

Opioid control of oxytocin secretion: evidence of distinct regulatory actions of two opiate receptor types

Stress induced oxytocin (OT) secretion was measured in female rats following treatment with various opiate antagonists selective for different types of opiate receptor. Naloxone (mu selective) and MR2266 BS (kappa selective) potentiated the OT response to an emotional stress (1 min. immobilization) whereas the delta selective antagonist ICI 154129 was without effect. Similarly, naloxone and MR2266 BS, but not ICI 154129, potentiated the response to a physical stress (i.p. hypertonic saline). A dose response comparison of the actions of naloxone and MR2266 BS revealed that naloxone was most effective in potentiating the immobilization response whereas MR2266 BS elicited greater responses than naloxone when administered prior to hypertonic saline. The results indicate that the opioid regulation of stress induced OT secretion is primarily mediated via mu and kappa opiate receptor types, the two types differentially regulating the OT response to two different stressors.     

Effects of lithium on the hypothalamo-pituitary-adrenal axis

The effect of lithium on the hypothalamo-pituitary-adrenal axis was studied in vivo and in vitro. The levels of plasma vasopressin, ACTH and corticosterone increased after the administration of lithium (LiCl 4 mmol/kg BW, 11 days) in rats, while the tissue vasopressin concentration in the median eminence, the rest of the hypothalamus and the posterior pituitary was decreased. The CRF concentration in the posterior pituitary increased markedly, but it did not change significantly in the median eminence or the rest of the hypothalamus. The elevated plasma ACTH level might be at least partly due to the increased vasopression secretion. Lithium stimulated ACTH secretion per se and also enhanced vasopressin-induced ACTH secretion in cultured pituitary cells and in half pituitary incubations, while it did not affect CRF-induced ACTH secretion. Lithium inhibited CRF-induced cAMP accumulation in half pituitary incubations, while lithium and vasopressin did not affect cAMP accumulation per se or even when administered together. The results suggest that lithium-induced ACTH release is via a cAMP-independent mechanism. Thus, it is possible that lithium stimulates ACTH release by acting directly on the corticotroph, stimulating vasopressin release and potentiating vasopressin-induced ACTH release.     

The met-enkephalin analog FK 33-824 and naloxone do not directly influence cortisol secretion by cultured human adrenocortical cells

Systemic administration of the enkephalin analog FK 33.824 was previously shown to inhibit ACTH secretion in man. In this study, the direct action of this analog on cortisol release was studied. The enkephalin analog (1 uM and 10 uM) did not influence basal or ACTH-stimulated cortisol production by cultured isolated adrenocortical cells prepared from the hyperplastic adrenal glands from three patients with Cushing's disease. Naloxone (10 uM) had also no direct effect on cortisol release. It is concluded that the met-enkephalin analog used in this study and naloxone do affect the hypothalamo-pituitary-adrenal axis via a central effect.     

Effect of enkephalins on vasopressin and aldosterone levels in acute experimental myocardial ischemia

It has been demonstrated in experiments on rats that acute myocardial ischemia gives rise to a decrease in diuresis, elevation of antidiuretic activity of blood plasma and the blood concentration of immunoreactive aldosterone. Intraperitoneal injection of a synthetic enkephalin analog D-ala2-leu5-arg6-enkephalin in a dose of 1.25 nmol/kg bw resulted in partial normalization of diuresis, reduction in antidiuretic activity of blood plasma and blood aldosterone level to the control values. Naloxone eliminated the effects described. It is concluded that enkephalins have an inhibitory action on aldosterone and vasopressin secretion, with this action being mediated via opiate receptors.     

Growth hormone-releasing hormone facilitates hypoglycemia-induced release of cortisol

Early sleep in humans is characterized by a distinct suppression of pituitary-adrenal activity coinciding with enhanced activity of the somatotropic axis. Here, we tested in awake humans the hypothesis of an inhibiting influence of hypothalamic growth hormone-releasing hormone (GHRH) on pituitary-adrenal activity. For this purpose, pituitary-adrenal activity was stimulated in 10 men through a standard insulin-hypoglycemia-test (IHT) and in another 10 men through combined administration of CRH/vasopressin. Stimulation was performed in each man on three conditions following pretreatment with Placebo and GHRH administered intravenously (50 microg) or intranasally (300 microg) 1 h before. GH, ACTH and cortisol as well as blood pressure and heart rate were measured repeatedly. Contrary to expectations, pretreatment with GHRH did not suppress but enhanced secretion of cortisol upon insulin-induced hypoglycemia regardless of the route of GHRH pretreatment (p<0.05). In contrast, GHRH did not facilitate cortisol release after stimulation with CRH/vasopressin. Changes in ACTH remained inconsistent. Plasma levels of GH increased significantly after i.v. GHRH application, but remained unchanged after the intranasal administration. Blood pressure and heart rate were not influenced by the treatments. Results indicate facilitating effects of GHRH mediated at a suprapituitary (i.e. hypothalamic) level as suggested by restriction of the effect to the hypoglycemia-induced cortisol release with no effects after pituitary stimulation with CRH/vasopressin.     

Cyclical edema in a patient with hypothalamic disorders and chronic glomerulonephritis: arginine vasopressin-dependent atrial natriuretic hormone release.

A 28-year-old woman had hypothalamic disorders (amenorrhea, obesity, psychiatric abnormalities, polydipsia and fever) and chronic glomerulonephritis. She also suffered from general edema associated with cyclical oliguria and polyuria. Her body weight and plasma osmolality increased during the oliguria phase lasting 2 to 8 days and decreased after paroxysmal polyuria accompanied by the natriuresis. These episodes occurred repeatedly, regardless of the treatment with or without diuretics. The release of arginine vasopressin in response to increased plasma osmolality was exaggerated, but changes in plasma volume did not affect arginine vasopressin release. Plasma atrial natriuretic hormone increased in response to a rise in plasma arginine vasopressin and plasma volume during the oliguria phase, thereby resulting in the diuresis and natriuresis. The renin-angiotensin-aldosterone system was secondarily activated by body fluid depletion and diuretics, and this might play an additive role in general swelling. Plasma gonadal hormones did not change to explain the edema. The mechanism of this cyclical edema remains unknown, but it is likely that hypothalamic dysfunction related to psychiatric abnormalities may exaggerate arginine vasopressin release, and enhanced renal sympathetic activity may cause retention of Na and water, and the increase in atrial natriuretic hormone release responding to the plasma volume expansion may bring about the diuresis and natriuresis.