Antioxidant properties of melatonin--an emerging mystery.

Over three centuries ago, the French philosopher René Descartes described the pineal gland as "the seat of the soul." However, it was not until the late 1950s that the chemical identity and biosynthesis of melatonin, the principal hormone secreted by the pineal body, were revealed. Melatonin, named from the Greek melanos, meaning black, and tonos, meaning color, is a biogenic amine with structural similarities to serotonin. The mechanisms mediating the synthesis of melatonin are transcriptionally regulated by the photoperiodic environment. Once synthesized, the neurohormone is a biologic modulator of mood, sleep, sexual behavior, reproductive alterations, immunologic function, and circadian rhythms. Moreover, melatonin exerts its regulatory roles through high-affinity, pertussis toxin-sensitive, G-protein (or guanine nucleotide binding protein) coupled receptors that reside primarily in the eye, kidney, gastrointestinal tract, blood vessels, and brain. Additional evidence also indicates a role for melatonin in aging and age-related diseases, probably related to its efficient free radical scavenger (or antioxidant) activity. The potential clinical benefit of melatonin as an antioxidant is remarkable, suggesting that it may be of use in the treatment of many pathophysiological disease states including various cancers, hypertension, pulmonary diseases, and a variety of neurodegenerative diseases such as Alzheimer's disease. This review summarizes the biosynthesis of melatonin and its many endocrine and physiological functions, including its therapeutic potential in human disease states. Emphasis is placed on the recent speculations indicating that this pineal hormone serves as an endogenous antioxidant agent with proficient free radical scavenging activity.

Corticosterone facilitates the acquisition of cocaine self-administration in rats: opposite effects of the type II glucocorticoid receptor agonist dexamethasone.

The effect of corticosterone on the acquisition of cocaine-seeking behavior was investigated in rats using ascending dose-response curves for intravenous cocaine self-administration. Rats pretreated daily with corticosterone (2.0 mg/kg i.p.) acquired cocaine self-administration at a lower dose compared with vehicle-treated controls. In contrast, daily corticosterone pretreatment did not alter food-maintained responding. Cocaine self-administration was not affected by the type I (mineralocorticoid) receptor agonist, aldosterone (100 microgram/kg). However, rats treated with the type II (glucocorticoid) receptor agonist, dexamethasone (10 or 100 microgram/kg) did not acquire self-administration at any dose tested. The 100 microgram/kg dose of dexamethasone attenuated food-reinforced behavior and decreased body weight, but these effects were not observed with the 10 microgram/kg dose. Dexamethasone dose-dependently attenuated the plasma corticosterone response to self-administered infusions or intraperitoneal injections of cocaine, indicating that the ability of dexamethasone to block cocaine-induced corticosterone secretion may have contributed to its effects on self-administration. Administration of aldosterone (100 microgram/kg) together with 10 microgram/kg dexamethasone restored self-administration to the level of vehicle-treated rats, suggesting that type I receptor occupation by corticosterone may be required for the acquisition of this behavior. These results indicate that stress-induced corticosterone secretion may provide a substrate through which stressors interact with cocaine reinforcement. Additionally, the finding that dexamethasone blocks the acquisition of cocaine self-administration may be relevant to the development of novel approaches to the treatment of cocaine addiction.

Neuroendocrine and immunocytochemical demonstrations of decreased hypothalamo-pituitary-adrenal axis responsiveness to restraint stress after long-term social isolation.

We have studied the effects of long-term social isolation of male Wistar rats, after early weaning (16 days), on the activity of the hypothalamo-pituitary-adrenal (HPA) axis. In addition to studying basal HPA activity, the response of the HPA axis to 15 min of immobilization stress was examined. Plasma corticosterone concentrations were measured, and the relative weights of adrenal glands, thymus, and testes were obtained, the latter to check whether gonadal function was affected by the isolation paradigm. Moreover, we carried out a quantitative immunohistochemical study of pituitary ACTH and its hypothalamic secretagogues: CRF, arginine vasopressin (AVP), and oxytocin (OT), both at the level of the synthesizing cell bodies in the hypothalamic paraventricular nucleus and of the releasing fibers in the median eminence (ME). Body weight and daily consumption of food and water were not altered, but social isolation caused a reduction in plasma corticosterone levels, both under basal and stress-stimulated conditions; this was correlated with an increased thymus weight, without affecting adrenal or testicular weights. The immunohistochemical study revealed that isolation caused a smaller increase in the number of ACTH-immunoreactive cells in the pars distalis of the anterior pituitary after exposure to restraint stress, as compared with control animals. This result indicates that fewer corticotrophs were activated by restraint stress in isolated animals, such cells being smaller and exhibiting a smaller ACTH-immunoreactive area than in control animals. Isolated animals also showed an increase in the content of CRF-ir fibers in the ME and a smaller decrease in the neuropeptide immunoreactivity after stress than that observed in control animals. This result could indicate a reduced release of CRF into the portal vasculature in response to acute stress and may partially explain the reduced activation of corticotrophs observed in the pituitary of isolated animals. However, no changes were found in the content of CRF, AVP, or OT within the paraventricular nucleus, nor of the AVP or OT content in the ME. The results of this study show that long-term social isolation after early weaning caused a hypofunction of the HPA axis in the adult rat. This hypofunction was particularly evident after exposure to an acute stressor, suggesting a desensitization of this axis to stressful stimuli.

Dexamethasone suppression of the effects of cocaine on adrenocortical secretion in Lewis and Fischer rats.

There is evidence to suggest that cocaine acts centrally to enhance adrenocortical secretory activity and this effect may be associated with the reinforcing properties of this drug. Lewis (LEW) and Fischer (F344) rats are inbred strains which differ in their responses to the reinforcing effects of cocaine. Previous findings from this laboratory have demonstrated differences in the hypothalamic-pituitary-adrenocortical (HPA) responses to cocaine between these strains. To determine whether strain differences in glucocorticoid responsiveness play a role in the differential effects of cocaine on plasma corticosterone (CS) secretion in these strains, experiments were designed to suppress the HPA response to cocaine in these two rat strains. HPA activity was attenuated by central administration of the glucocorticoid agonist dexamethasone (DEX) using osmotic minipumps. A constant infusion of artificial cerebrospinal fluid or DEX (50, 100 or 500 ng/h) was delivered into the lateral ventricle of LEW and F344 rats. Four days later, the rats were challenged with cocaine HCl (0, 20 and 40 mg/kg, i.p.), and the plasma CS response 15 min later was quantified. Cocaine-induced alterations in circulating plasma CS were reduced in a dose-related manner by centrally administered DEX in both strains. Significant strain differences in the effects of DEX on the plasma CS response to cocaine were observed, suggesting that LEW rats were more sensitive to DEX suppression of HPA activity than F344 rats. DEX also produced dose-related effects on body weight in both strains and decreased adrenal weight at the highest dose in F344 rats. Blood collected on the final day of the experiment demonstrated that central infusions of DEX decreased plasma ACTH concentrations in both strains compared to control rats. These studies indicate that central administration of DEX produces a feedback inhibition of cocaine-induced glucocorticoid release and that LEW rats are more sensitive to DEX suppression than F344 rats.

Role of 5-hydroxytryptamine3 (5-HT3) antagonists in the prevention of emesis caused by anticancer therapy.

Most anticancer drugs are cytotoxic and produce various side-effects, among which nausea and vomiting are almost ubiquitous and usually extremely distressing to the patient. Cancer chemotherapy elicits two main phases of vomiting: an intense, acute phase of vomiting that occurs almost immediately following anti-cancer therapy and a milder, delayed phase of nausea and vomiting of longer duration. The mechanisms underlying the induction of nausea and vomiting after cancer chemotherapy are poorly understood but may be mediated by serotonin (5-hydroxytryptamine or 5-HT), particularly in the acute phase. Serotonin activates 5-HT3 receptors, which function as ligand-gated ion channels located either in the periphery and/or in the central nervous system to produce emesis, among other effects. The peripheral 5-HT3 receptors may be pharmacologically distinct from the central 5-HT3 receptors and may exhibit some association with GTP-binding proteins. In addition, different populations may exist as distinct subtypes of the same receptor. The 5-HT3 receptor antagonist ondansetron (GR 38032F) is effective in preventing the emesis induced by cytotoxic agents currently used in the treatment of many forms of cancer. Ondansetron has, comparatively, a much higher efficacy in the treatment of acute emesis following cancer chemotherapy than it does in the delayed phase, suggesting that the late phase of emesis may be mediated by other distinct mechanisms.

Differential neuroendocrine and behavioral responses to cocaine in Lewis and Fischer rats.

Lewis (LEW) and Fischer 344 (F344) rats differ in responsiveness of the hypothalamo-pituitary-adrenocortical (HPA) axis as well as in their behavioral responses to drugs of abuse. The present experiments were conducted to compare hypothalamic corticotropin-releasing factor (CRF), plasma adrenocorticotropic hormone (ACTH) and plasma corticosterone (CS) responses to cocaine (0-60 mg/kg, i.p.) in LEW and F344 rats. Acute administration of cocaine resulted in decreases in CRF and dose-related increases in CS and ACTH with significant differences observed between the strains. Cocaine also increased plasma norepinephrine concentrations. Although the CS response was increased in the F344 compared to LEW rats, the percent change in the CS response was markedly enhanced in LEW rats. Plasma ACTH concentrations as well as the percentage of the control response were dramatically increased at the 40 mg/kg cocaine dose in the LEW compared to F344 rats. Since cocaine-induced changes in HPA axis activity may contribute to behavioral responses to cocaine, another experiment was performed to compare the locomotor responses to novelty and to acute cocaine between LEW and F344 rats. Strain differences were not observed in the locomotor response to novelty or to cocaine. These data indicate that strain differences exist in the neuroendocrine response to acute cocaine exposure.

Effects of the serotonin1A agonist, 8-hydroxy-2-(di-n-propylamino)tetralin on neurochemical responses to stress.

The effects of intracerebroventricular administration of the 5-hydroxytryptamine (5-HT)1A agonist, 8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT; 0.1 pmol) on adrenocortical and neurochemical responses to stress were examined in conscious male rats. The following stress paradigms were used: acoustic stimulation (105 dB for 2 min); footshock (0.2 mA, five shocks over 5 min); conditioned fear (animals placed in a footshock chamber for 5 min, 24 h after footshock); restraint (5 min); intraperitoneal (i.p.) injection of recombinant human interleukin-1 alpha (rHu-IL-1 alpha, 20 micrograms/kg); and injection of cocaine hydrochloride (20 mg/kg, i.p.). As previously shown, 8-OH-DPAT was able to attenuate the adrenocortical response to acoustic stress, conditioned fear, rHu-IL-1 alpha, and cocaine administration. Cocaine decreased 5-hydroxyindoleacetic acid (5-HIAA)/5-HT and dihydroxyphenylacetic acid/dopamine (DOPAC/DA) ratios and norepinephrine (NE) concentration in the prefrontal cortex, hypothalamus, and brainstem in all experiments, and 8-OH-DPAT reversed the changes in DOPAC/DA ratio without affecting 5-HIAA/5-HT ratios or NE content. 8-OH-DPAT alone had no effect on these parameters, although it decreased NE content in the prefrontal cortex in several experiments, and in the brainstem in one experiment. Significant decreases in NE content were observed in some brain regions following some of the stressors, but these changes were not generally affected by 8-OH-DPAT. Increases in the 5-HIAA/5-HT and DOPAC/DA ratios were also observed in some brain sites following some stressors, but these changes were not affected by 8-OH-DPAT except in the case of the increased 5-HIAA/5-HT ratio in the prefrontal cortex following the conditioned fear response. These results indicate that although 8-OH-DPAT is able to decrease plasma corticosterone responses following acoustic stress, conditioned fear, rHu-IL-1 alpha, and cocaine administration, these effects do not appear to be related to an action of the 5-HT1A agonist on biogenic amine metabolism. This observation indicates that the predominant effect of 8-OH-DPAT on adrenocortical responses is mediated at postsynaptic sites not involved in the regulation of cerebral biogenic amine metabolism.

Adrenalectomy alters the response of neurons in the bed nucleus of the stria terminalis to electrical stimulation of the medial amygdala.

This study was performed to characterize the effects of adrenalectomy (ADX) on electrical activity and synaptic responses of bed nucleus of the stria terminalis (BNST) and preoptic area (POA) neurons, which are involved in the control of limbic-hypothalamo-pituitary-adrenocortical (LHPA) activity. Adrenalectomy altered the response of BNST neurons to medial amygdala (AME) stimulation, increasing the proportion of excitatory responses and reducing the number of cells inhibited. No such effects were found for neurons within the POA. The basal activity of neurons recorded within the BNST and POA, as well as the latencies and duration of responses, was not affected. The specificity of the effects upon BNST, but not POA, neurons suggests that the response of BNST neurons to AME stimulation is corticosteroid dependent, whereas the response of preoptic neurons is not.

Effects of prolonged social isolation on responses of neurons in the bed nucleus of the stria terminalis, preoptic area, and hypothalamic paraventricular nucleus to stimulation of the medial amygdala.

The studies presented demonstrate changes in hypothalamo-pituitary-adrenocortical secretion, and in electrical activity and synaptic responses of neurons in the bed nucleus of the stria terminalis, preoptic area, and hypothalamic paraventricular nucleus of rats exposed to early, long-term social isolation. Rats isolated from all social contact from an early preweaning time showed reduced basal plasma corticosterone concentrations, compared with littermate controls raised under social conditions. Isolated animals also exhibited a selective decrease in the spontaneous electrical activity of neurons within the hypothalamic paraventricular nucleus and lateral preoptic area, but not in adjacent structures. Moreover, isolation also altered the response of neurons in certain nuclei to electrical stimulation of the medial amygdala. Thus, a reduction in excitatory responses, and an increase in inhibition and nonresponsiveness, of preoptic area and paraventricular nucleus neurons was recorded, compared with control rats. Neurons in the bed nucleus of the stria terminalis were less affected, but showed an increase in the duration of excitatory responses following medial amygdala stimulation. These results, obtained from urethane-anesthetized rats, together with the reduced basal plasma corticosterone concentrations, suggest a reduction in limbic-hypothalamo-pituitary-adrenocortical (LHPA) activity following maternal deprivation and prolonged social isolation. This may result from altered limbic activity, specifically in the amygdala and its pathways to the paraventricular nucleus (PVN). Such alterations may include the stria terminalis, in so much as increased efficacy of inhibitory components and reduced efficacy of excitatory components was observed. The neural mechanisms underlying these alterations could involve an altered synaptology of the regions examined and/or a disruption of glucocorticoid feedback events.

The brain-immune axis: role of opiates and other substances of abuse, the hypothalamic-pituitary-adrenal axis and behavior.

The brain-immune axis is a highly complex and dynamic homeostatic mechanism. The effect of morphine, or other drugs of abuse, on this interactive system can have multiple consequences, depending on the specific site(s) of action, duration of exposure and underlying neuroendocrine, neural and behavioral status. This emphasizes the critical role of interdisciplinary studies, incorporating behavioral, pharmacological, neuroanatomical, cellular and molecular approaches, that are necessary for elucidating opiate-BIA interactions. To characterize fundamental mechanisms of action, the sheer complexity of this system often requires that more rigorous experimental methodology be applied by multidisciplinary groups of investigators than ever before.

Neuroendocrine effects of interferon-alpha in the rat.

We have previously found that recombinant human interferon-alpha 2A (rHu-IFN-alpha 2A) inhibits hypothalamo-pituitary-adrenocortical (HPA) axis activity following both peripheral and central administration. This effect is antagonized by mu-opioid receptor antagonists, suggesting transduction by this subtype of opioid receptors. We have now demonstrated that this effect is also observed with hybrid rHu-IFN-alpha A/D, rat kidney fibroblast-derived IFN-alpha, and recombinant rat IFN-alpha preparations. The inhibitory effects on HPA activity were observed after intraperitoneal (i.p.) injections of rHu-IFN-alpha2A(10(03)U), rHu-IFN-alpha A/D (10(4)U), and of Rat-IFN-alpha (1-10U). Similar effects were observed with intracerebroventricular (i.c.v.) administration of all four IFN-alpha preparations. No increases in plasma corticosterone concentrations were observed with doses of rHu-IFN-alpha A/D up to 10(6)U (i.p.) or 7x10(5)U (i.c.v.), but increases were found following i.c.v. administration of high doses of Rat-IFN-alpha (10(3) and 5x10(3)U). The inhibitory effects of all of the IFN-alpha preparations tested were antagonized by naloxone, but the stimulatory effects of 5x10(3)U Rat-IFN-alpha were not. Injections of rHu-IFN-alpha 2A(10(4)U, i.p.) to urethane-anesthetized rats decreased the electrical activity of the majority of hypothalamic paraventricular nucleus (PVN) neurons tested, including putative corticotropin-releasing factor-(CRF)-secreting neurons antidromically identified as projecting to the median eminence. Similarly, iontophoretic application of rHu-IFN-alpha 2A decreased the electrical activity of such cells. These electrophysiological data suggest that the decreases in HPA activity evoked by IFN-alpha are mediated, at least in part, by a rapid inhibitory effect at the level of the corticotropin-releasing factor-secreting neurons.

Differential inhibition of stress-induced adrenocortical responses by 5-HT1A agonists and by 5-HT2 and 5-HT3 antagonists.

The present studies were designed to determine the effects of 5-HT1A receptor agonists and 5-HT2A/2C and 5-HT3 antagonists on adrenocortical responses to a variety of stress paradigms in conscious male rats. The following stressors were examined: acoustic stress (105 dB for 2 min); foot shock (0.2 mA, five shocks over 5 min); conditioned fear (animals placed in the foot shock chamber for 5 min, 24 h after foot shock); restraint (Plexiglas restrainer for 5 min); injection of recombinant human interleukin-1 alpha (IL-1, 20 micrograms/kg, IP); injection of cocaine hydrochloride (20 mg/kg, IP). Drug treatments consisted of intracerebroventricular (ICV) or intraperitoneal (IP) injections of the 5-HT1A agonists, 8-OH-DPAT and ipsapirone (0.1 pmol, ICV), the 5-HT2A/2C antagonist, ketanserin (2 mumol/kg, IP), and the 5-HT3 antagonist, MDL-72222 (20 nmol, ICV). The plasma corticosterone (CS) responses to foot shock and restraint stress were not affected by any of the serotonergic drugs tested. The 5-HT1A agonist, 8-OH-DPAT, was able to attenuate the adrenocortical responses to acoustic stimulation, conditioned fear, IL-1 alpha, and cocaine administration, with ipsapirone also being effective in reducing the responses to acoustic stimulation and cocaine injection. The 5-HT2 antagonist, ketanserin was able to reduce the adrenocortical response in the conditioned fear paradigm and the response to IL-1 alpha injection. The 5-HT3 antagonist, MDL-72222 was only effective in reducing the response to acoustic stimulation. Thus, adrenocortical responses to each of the applied stressors were differentially affected by the 5-HT receptor ligands tested. The results of this study indicate that 5-HT1A agonists may be efficient stress response-reducing agents. However, their efficacy depends on the lack of a somatosensory component to the applied stressor and their agonist properties suggest that their action may not involve direct effects on serotonergic pathways mediating the observed responses. In contrast, the specificity of the 5-HT2 and 5-HT3 antagonists in blocking adrenocortical responses to certain stressors suggests that these drugs exert their effects by blocking serotonergic neurotransmission in pathways mediating the adrenocortical responses to specific stimuli.

Central stimulation of adrenocortical secretion by 5-hydroxytryptamine1A agonists is mediated by sympathomedullary activation.

The mechanism by which the 5-hydroxytryptamine1A agonist, 8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT) increases adrenocortical secretion was examined using treatments designed to alter the hypothalamo-pituitary-adrenocortical axis and the autonomic nervous system. Adrenocortical responses to peripherally administered 8-OH-DPAT were preserved in animals with radiofrequency lesions of the hypothalamic paraventricular nucleus or with posterior hypothalamic deafferentations. However, adrenocortical responses to 8-OH-DPAT were abolished in hypophysectomized animals, although both acute and chronic treatment with dexamethasone were without effect. The influence of catecholamines in the central and sympathetic nervous systems was investigated using treatments designed to alter central and peripheral catecholaminergic systems. Adrenocortical responses after intracerebroventricular administration of high doses of 8-OH-DPAT were attenuated by i.v. and intra-adrenal treatment with 6-hydroxydopamine (6-OHDA), by adrenal medullectomy, by splanchnicectomy and by combined 6-OHDA-sympathectomy/medullectomy. Responses to i.p. 8-OH-DPAT were only partially attenuated by medullectomy and by 6-OHDA-sympathectomy/medullectomy. The ganglionic blocking agent, chlorisondamine also blocked the adrenocortical response after intracerebroventricular, but not i.p., administration of 8-OH-DPAT. These effects did not appear to be due to surgical damage to the adrenal cortex because the operated animals showed normal adrenocortical responses to challenge with adrenocorticotropic hormone. The data obtained indicate that adrenocortical secretion in response to i.p. and intracerebroventricular administration of the 5-hydroxytryptamine1A receptor agonist 8-OH-DPAT largely involves a sympathomedullary activation, but that additional peripheral mechanisms are also involved after i.p. administration.

Central and peripheral mechanisms in the stimulation of adrenocortical secretion by the 5-hydroxytryptamine2 agonist, (+-)-1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane.

The mechanisms by which the serotonin2A/2C (5-HT2A/2C) receptor agonist, (+-)-1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (DOI) increase adrenocortical secretion were examined. Intraperitoneal (i.p., 2 mumol/kg) and intracerebroventricular (i.c.v., 0.1 pmol -40 nmol) administration of DOI increased plasma corticosterone (CS) concentrations. Administration of the 5-HT2A/2C antagonist, ketanserin (i.c.v.) also increased adrenocortical secretion, although the more selective 5-HT2A/2C antagonist, 6-methyl-1-(1-methylethyl)-ergoline-8 beta-carboxylic acid 2-hydroxy-1-methylpropyl ester (LY 53857) did not. In pentobarbital-anesthetized rats, DOI (i.c.v.) decreased adrenocortical secretion, whereas i.p. administration increased adrenocortical secretion. Ketanserin and LY 53857 (i.c.v.) failed to block CS responses after i.p. DOI, indicating that these increases were not principally due to central 5-HT2 receptor activation; only i.p. administration of ketanserin was able to block responses to DOI. Adrenocortical responses to DOI (i.p.) were preserved after lesions of the hypothalamic paraventricular nucleus or posterior hypothalamic deafferentations. However, responses to DOI were almost abolished in hypophysectomized animals, whereas neurohypophysectomy and treatment with dexamethasone were without effect. Adrenocortical responses after DOI (i.p.) were attenuated in medullectomized and in combined 6-hydroxydopamine-sympathectomized/medullectomized animals, but responses to i.c.v. DOI were not affected by such treatment. 6-Hydroxydopamine lesions of the ventral noradrenergic ascending bundle abolished the response to i.p. DOI, indicating that activation of central noradrenergic systems mediates the increases in adrenocortical secretion. These results demonstrate that central and peripheral 5-HT2 receptors differentially regulate adrenocortical secretion, the central component being sensitive to pentobarbital anesthesia and the peripheral component being dependent upon central and peripheral noradrenergic systems.

Inhibition of neural and neuroendocrine activity by alpha-interferon: neuroendocrine, electrophysiological, and biochemical studies in the rat.

In our earlier studies we have demonstrated that recombinant human interferon-alpha 2A (rHu-IFN-alpha 2A) inhibits hypothalamo-pituitary-adrenocortical (HPA) secretion following both peripheral and central administration. Furthermore, this effect is antagonized by mu-opioid receptor antagonists, suggesting transduction by this subtype of opioid receptors. In the present studies, we demonstrate that this effect is also observed with the hybrid recombinant preparation, rHu-IFN-alpha A/D, and a leucocyte-derived rat IFN-alpha preparation. The inhibitory effects on HPA activity were observed after intraperitoneal (i.p.) injections of rHu-IFN-alpha 2A (10(3) U), rHu-IFN-alpha A/D (10(4) U), and of Rat-IFN-alpha (1 and 10 U). Similar effects were observed with intracerebroventricular (i.c.v.) administration of all three IFN-alpha preparations. No increases in plasma concentrations of corticosterone were observed with doses of rHu-IFN-alpha A/D up to 10(6) U (i.p.) or 7 x 10(5) U (i.c.v.), but increases were found following i.c.v. administration of high doses of Rat-IFN-alpha (10(3) and 5 x 10(3) U). The inhibitory effects of all of the IFN-alpha preparations tested were antagonized by naloxone, but the stimulatory effects of 5 x 10(3) U Rat-IFN-alpha were not. Injections of rHu-IFN-alpha 2A (10(4) U i.p.) to urethane-anesthetized rats decreased the electrical activity of the majority of hypothalamic paraventricular nucleus neurons tested, including putative corticotropin-releasing factor-secreting neurons antidromically identified as projecting to the median eminence. These electrophysiological data suggest that the decreases in HPA activity evoked by IFN-alpha are mediated by a rapid inhibitory effect at the level of the corticotropin-releasing factor-secreting neurons. The sensitivity of many central nervous system effects of IFN-alpha to mu-receptor antagonists strongly suggests that the cytokine serves as an endogenous opioid agonist arising from the immune system. In support of this hypothesis we have shown that SH-SY5Y human neuroblastoma cells, differentiated with retinoic acid treatment to express predominantly mu-receptors, are sensitive to rHu-IFN-alpha 2A in vitro. This sensitivity took the form of a dose-dependent inhibition of forskolin-stimulated adenylyl cyclase activity. The data yielded an IC50 (95% confidence intervals) value of 7.93 (5.70-11.04) nM for this effect. Neither undifferentiated SH-SY5Y cells nor NG108-15 mouse neuroblastoma x rat glioma hybrid cells (expressing delta-receptors) were affected by rHu-IFN-alpha 2A.(ABSTRACT TRUNCATED AT 400 WORDS)

Interactions between serotonin, thyrotropin-releasing hormone, and substance P in the CNS regulation of adrenocortical secretion.

Double-labeling immunohistochemical studies were performed to discern the morphological relationships between corticotropin-releasing factor-immunoreactive (CRF-ir) perikarya and afferent innervation in the hypothalamic paraventricular nucleus (PVN) of the rat. Attention was focussed on the local innervation by serotonin (5-hydroxytryptamine, 5-HT), thyrotropin-releasing hormone (TRH) and substance P (SP)-ir nerve terminal fibers. 5-HT-ir and SP-ir fibers were present in moderate numbers, in close apposition with CRF-ir perikarya. Sparse TRH-ir fibers were observed, but a population of TRH-ir perikarya was found in proximity with the CRF-ir cell bodies. TRH-ir perikarya in the PVN were surrounded by both 5-HT- and SP-ir fibers. Neuroendocrine studies were performed to investigate the interactions between 5-HT, TRH and SP in the regulation of hypothalamo-pituitary-adrenocortical (HPA) secretion. Male rats were prepared bearing cannulae for intracerebroventricular (ICV) or intra-PVN administration of drugs. 5-HT, at all doses tested (0.1, 40, or 80 nmol, ICV), caused increases in plasma corticosterone (CS) concentrations in tail-vein blood collected 20 min after injection. ICV injections of TRH caused dose-dependent increases in plasma CS, but did not further increase HPA responses when injected together with 5-HT. SP alone had little effect, although a significant reduction in plasma CS concentrations was observed in several individual experiments. However, SP (0.1 nmol) significantly attenuated CS responses following high doses of 5-HT (40 and 80 nmol, ICV), although the response to 0.1 nmol 5-HT was not affected. Combined injection of SP with TRH resulted in HPA responses not different from those following TRH alone. Similarly, SP did not reduce the HPA response observed with TRH and 40 nmol 5-HT in combination. Intra-PVN injections of 5-HT (0.1 or 40 nmol) and TRH also increased plasma CS concentrations. Intra-PVN injections of SP had little effect on plasma CS concentrations although a tendency toward a decrease in plasma CS was observed, as with the ICV injections. Combined intra-PVN injection of 5-HT (0.1 nmol) with TRH (0.1 nmol) did not significantly alter the response compared with that observed following TRH alone, although plasma CS concentrations were greater than with 0.1 nmol 5-HT. Combined intra-PVN injections of SP (0.1 nmol) with 5-HT (0.1 nmol) resulted in a significant decrease in plasma CS concentration compared with that following 5-HT alone, but SP did not prevent the CS response to a higher dose of 5-HT (40 nmol).(ABSTRACT TRUNCATED AT 400 WORDS)

Inhibition by the serotonin1A agonist, 8-hydroxy-2- (di-n-propylamino)tetralin, of antidromically identified paraventricular nucleus neurons in the rat.

Experiments were performed using urethane-anesthetized rats to delineate the role of serotonin (5-hydroxytryptamine, 5-HT) receptors of the 5-HT1A subtype in the neural regulation of hypothalamo-pituitary-adrenocortical (HPA) secretion. The activity of single paraventricular nucleus (PVN) neurons antidromically identified as projecting to the median eminence was recorded following injection of the 5-HT1A agonist, 8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT). Intraperitoneal administration of increasing doses of 8-OH-DPAT caused dose-dependent decreases in the spontaneous activity of all neurons tested. The data yielded an ED50 value of 0.19 mumol/kg for this effect. The decreases in PVN neuronal activity were accompanied by sustained decreases in mean arterial blood pressure, with an ED50 of 0.18 mumol/kg. These results provide evidence for an inhibitory role of 5-HT1A receptors in the regulation of PVN neuronal activity and support the suggestion that 5-HT may inhibit HPA secretory activity.

Alpha-interferon inhibits adrenocortical secretion via mu 1-opioid receptors in the rat.

The effect of recombinant human alpha-interferon on plasma corticosterone concentrations was investigated in adult male rats. Intraperitoneal (i.p.) or intracerebroventricular (i.c.v.) administration of alpha-interferon (10-10(4) U i.p., and 1-10(3) U i.c.v.) decreased basal plasma corticosterone concentrations. This effect was evident at both the peak and nadir in the circadian rhythm of hypothalamo-pituitary-adrenocortical secretory activity. The same inhibitory effect was obtained with intra-paraventricular nucleus administration of the cytokine. Furthermore, alpha-interferon attenuated the effects of stressors such as handling, 1 min of forced swimming, and sound stress in a novel environment. The effect of alpha-interferon (10(2) U i.c.v.) was blocked by prior injection of the opioid receptor antagonist, naloxone (1 mg/kg i.p.). Similarly, the effect of 10(3) U alpha-interferon administered i.p. was blocked by i.c.v. injection of naloxone (1 microgram/kg), or of the mu 1-specific receptor antagonist, naloxonazine (1 microgram). The selective delta-opioid receptor antagonist, naltrindole (1 microgram i.c.v.) and the kappa-opioid receptor antagonist, nor-binaltorphimine (1 microgram i.c.v.) both failed to prevent the inhibitory effect of alpha-interferon (10(3) U i.p.) on adrenocortical secretion. The results obtained provide further evidence for a neuromodulatory effect of alpha-interferon and that this effect is mediated by central opioid receptors of the mu 1-subtype, delta- and kappa-opioid receptors not being involved.

Electrophysiology and neuropharmacology of noradrenergic projections to rat PVN magnocellular neurons.

Responses of electrophysiologically identified tuberohypophysial paraventricular nucleus (PVN) neurons were examined following electrical stimulation of the A1, A2, A6, and C2 catecholaminergic cell groups and of the ventral noradrenergic ascending bundle (VNAB). A1, A2, and A6 stimulation evoked primarily excitatory responses from the cells recorded, but C2 stimulation yielded a greater proportion of inhibitory responses. VNAB stimulation at low frequencies (0.5/5 Hz) excited the majority of cells tested, but high-frequency (50 Hz) trains of stimulation reversed the direction of response to inhibition for approximately half of the cells excited by single-shock stimulation. Only 5-Hz stimulation had any affect on blood pressure, causing a slight increase. Treatment with alpha-methylparatyrosine, to inhibit (nor) epinephrine synthesis, reduced the proportion of excitatory responses and prevented the response reversals following 50-Hz VNAB stimulation. Treatment with 6-hydroxydopamine also reduced the proportion of cells excited by VNAB stimulation. Iontophoresis of either norepinephrine or the alpha 1-adrenoceptor agonist l-phenylephrine increased the activity of most cells tested whilst the alpha 1-antagonist ergotamine reduced the activity of most cells tested and prevented excitation elicited by VNAB stimulation. The alpha 2-agonist clonidine excited all cells tested. The beta-antagonist propranolol increased the activity of the majority of cells and prevented inhibitory responses following 50-Hz VNAB stimulation. The results confirm a role for brain stem projections in regulating PVN neuronal activity and demonstrate for the first time that the VNAB provides excitatory input to the PVN, primarily regulated by alpha 1-adrenoceptors. The effects of propranolol on spontaneous activity, and on the inhibitory responses following high-frequency VNAB stimulation, indicate the presence of an inhibitory counterbalancing beta-adrenoceptor mechanism.

Electroencephalographic changes in experimental autoimmune myasthenia gravis.

A number of reports have suggested that central disturbances of cholinergic function may occur in patients with myasthenia gravis. The present study was designed in order to examine cortical electroencephalographic (EEG) activity in Lewis rats with experimental autoimmune myasthenia gravis (EAMG). Experiments were performed on conscious rats with clinical EAMG and demonstrable antibodies against the acetylcholine receptor. The animals showed no gross changes in cortical EEG discharge in terms of cycles and durations of wake, desynchronized sleep, and synchronized sleep, as compared with control rats. However, abnormalities characterized by single spikes or waves, and by spike and wave complexes, were observed, most commonly during synchronization of the EEG. Use of computerized frequency analysis of the EEG records revealed the presence of three basic differences in EEG discharge in myasthenic animals: (1) additional high-amplitude, low frequency (< 4 Hz) activity was recorded, especially during synchronized sleep; (2) decreases in mid-range (4-7 Hz) activity were recorded, particularly during periods of wakefulness; and, (3) increases in high frequency (> 8 Hz) spike discharge were observed at all times, although this was most evident during periods of synchronized sleep. The data provide further evidence for alterations in central cholinergic function in myasthenia gravis.