A novel endogenous corticotropin release inhibiting factor.

ACTH is the major regulator of the body's adaptive response to stress and the physiological stimulus for glucocorticoid secretion. A hypothalamic corticotropin release inhibiting factor (CRIF) that inhibits ACTH synthesis and secretion has long been postulated but was not characterized until recently. We have recently identified a 22 amino acid peptide, prepro-thyrotropin releasing hormone (TRH) 178-199 that inhibits basal and stimulated ACTH synthesis and secretion in vitro and stress-induced ACTH secretion in vivo. Prepro-TRH 178-199 is abundant in several brain regions, including the external zone of the median eminence, where its concentration changes in response to stress. We propose that this peptide is a physiological regulator of ACTH production: an endogenous CRIF. Because prepro-TRH 178-199 is encoded within the same precursor as TRH, its expression is likely to be negatively regulated by thyroid hormones leading to changes in endogenous glucocorticoid levels. Streptococcal cell wall (SCW)-induced inflammation, a model of rheumatoid arthritis (RA), was alleviated after long-term thyroxine treatment. Inversely, a hypothyroid milieu led to decreased basal hypothalamic-pituitary-adrenal activity, but increased expression of IL-1 beta and MIP-1 alpha, specific markers for RA in humans. These results suggest that this putative CRIF may be an important component in the development of RA and that regulation of prepro TRH may be highly relevant to the development of other autoimmune diseases that are also exacerbated by low endogenous glucocorticoid levels.

Sexually dimorphic effects of maternal adrenalectomy on hypothalamic corticotrophin-releasing factor, glucocorticoid receptor and anterior pituitary POMC mRNA levels in rat neonates.

The stress hyporesponsive period (SHRP) occurs during the first two weeks of life (from day 4 to day 14) in the rat. SHRP may occur due to immature hypothalamic-pituitary-adrenal (HPA) regulatory mechanisms of the neonate. Decreased expression of corticotropin-releasing factor (CRF) has been observed during this period, and this decreased hypothalamic 'drive' may contribute to the manifestation of SHRP in the rat neonate. Since maternal corticosteroids are elevated toward the end of gestation they may suppress fetal CRF expression leading to a less than adequate activation of the neonatal hypothalamus in response to stress. Therefore, the purpose of the present study was to clarify the contribution of maternal glucocorticoids to the decreased CRF expression observed during SHRP in the neonatal rat. We investigated the effects of maternal adrenalectomy on hypothalamic CRF, glucocorticoid receptor (GR) and anterior pituitary proopiomelanocortin (POMC) mRNA levels in male and female neonates. Maternal adrenalectomy, or sham surgery, was performed on day 8 of gestation and the mRNA levels of CRF, GR and POMC were measured by Northern blotting in the offspring on their postnatal days 1, 7, 14, and 21. The observed changes in the mRNA levels of these genes suggests that an important developmental event occurs in the regulation of these HPA genes between neonatal days 7 and 14 corresponding to the termination of SHRP. Female offspring had significantly higher levels of CRF mRNA than males throughout this period. The lack of maternal corticosteroids evoked a gender-specific response in the neonates. In female offspring, maternal adrenalectomy resulted in a dramatic and correlated increase in mRNA levels of hypothalamic CRF and GR on day 14, with pituitary POMC expression not following this increase. There was no significant effect of maternal adrenalectomy on the expression of these genes in males. These results suggest a sex difference in response to maternal glucocorticoids in the fetus. However, the role of maternal corticosteroids in the low expression of CRF during SHRP could not be established from this study, since their removal by adrenalectomy did not advance the expression profile of CRF toward an earlier increase in the neonatal hypothalamus.

Thyroxine administration prevents streptococcal cell wall-induced inflammatory responses.

Administration of streptococcal cell wall (SCW) preparation induces an inflammatory response in susceptible animals that is a model frequently used for rheumatoid arthritis. The degree of inflammation produced by SCW is greatly enhanced by low endogenous levels of glucocorticoids due to diminished hypothalamic-pituitary-adrenal activity. Because decreased glucocorticoid production is known to occur in the hypothyroid state, we tested whether hypothyroidism would increase, and conversely, whether hyperthyroidism would decrease, the inflammatory responses to SCW. Adult female Sprague Dawley rats were fed a regular diet (control), L-T4 (T4; hyperthyroid), or 6-propyl-thiouracil (hypothyroid) in drinking water for 7 weeks. Hypothyroidism resulted in elevated plasma levels of TSH and hypothalamic preproTRH messenger RNA (mRNA) while reducing anterior pituitary POMC mRNA and plasma ACTH and corticosterone levels. In contrast, hyperthyroid rats produced opposite results: decreased measures of central thyroid function but increased pituitary-adrenal function. Three days after administration of SCW, macrophage inflammatory protein-1alpha and interleukin-1beta mRNA expression increased dramatically in controls and even further in hypothyroid animals, as measured by Northern blot analysis. In contrast, T4-treated rats showed significant inhibition of these inflammatory markers. Thus, the hyperthyroid state combined with increased endogenous glucocorticoid levels is protective against inflammatory challenges. The inverse relationship between preproTRH expression and pituitary-adrenal function suggests the possibility of a direct inhibitory link connecting the hypothalamic-pituitary-adrenal and thyroid axes, and suggests alternative sites of therapeutic intervention for rheumatoid arthritis and other inflammatory associated disorders.

Streptozotocin-induced diabetes produces a decrease in pituitary substance P content and preprotachykinin mRNA.

Complications arising from diabetes mellitus include hormonal dysfunctions such as impairment in the regulation of gonadatroph and corticotroph secretion. Preprotachykinin (PPT) mRNA encoding the peptide substance P (SP), has been localized in the anterior pituitary. The goal of this study was to determine if streptozotocin (STZ)-induced diabetes affects the SP content or PPT mRNA level in the pituitary of male rats. We injected STZ (55 mg/kg) to 6-week-old rats which developed hyperglycemia (blood glucose > 400 mg/dl) by 6 weeks post-injection. SP-like immunoreactivity in the pituitary dropped 54%. In situ hybridization was performed using a PPT-specific oligonucleotide with signal intensity differences semi-quantified using an image analysis system. Normal pituitary had a regional distribution of PPT mRNA, with no detectable signal in the posterior or intermediate lobes, while the anterior lobe displayed a distinctive pattern of labeled cells arranged in clusters. In diabetic rats there was a 23% decrease in the PPT-mRNA hybridization signal compared to controls (P < 0.05). The changes observed in PPT gene expression and SP content may be additional factors participating in the hormonal complications seen in diabetes mellitus.

Neuroendocrine and cardiovascular effects of serotonin: selective role of brain angiotensin on vasopressin.

Central serotonin (5-HT) and angiotensin (ANG II) stimulate arginine vasopressin (AVP), oxytocin (OT), and adrenocorticotropin (ACTH) secretion and increase blood pressure. Studies were conducted in conscious rats to determine whether neuroendocrine activation by 5-HT requires a brain angiotensinergic intermediate pathway. In the first study, ANG II formation was inhibited by the angiotensin-converting enzyme inhibitor enalapril before injection of the 5-HT releaser/uptake inhibitor d-fenfluramine. Fenfluramine (2 mg/kg ip) stimulated AVP, OT, corticosterone, and prolactin (PRL) secretion (P<0.01). Enalapril (60 mg/l in drinking water for 4 days and 10 mg/kg ip 2 h before the rats were killed) inhibited only the AVP response (P<0.01) to d-fenfluramine. In the second study, the effect of intracerebroventricular injection of the 5-HT2A/2C antagonist LY-53857 (10 microgram), or the ANG II AT1 antagonist DuP-753 (10 microgram), on intracerebroventricular 5-HT (10 microgram)-stimulated AVP, OT, ACTH, PRL, renin secretion, mean arterial pressure (MAP) and heart rate (HR) was tested. LY-53857 inhibited the AVP, OT, and ACTH responses to 5-HT (P<0.01), whereas DuP-753 inhibited only the AVP response (P<0.01). Intraventricular injection of 5-HT increased MAP and decreased HR. The MAP response was not affected by LY-53857 or DuP-753, and at no time did MAP decline below starting levels. The decreased HR was inhibited by LY-53857 but not by DuP-753. These results demonstrate that 5-HT-induced AVP secretion is mediated selectively via brain angiotensinergic mechanisms by way of the AT1 receptor.

Streptozotocin-induced diabetes is associated with altered expression of peptide-encoding mRNAs in rat sensory neurons.

Major complications arising from diabetes mellitus include neuropathic pain and altered peripheral inflammatory responses. Somatostatin (SOM), calcitenin gene-related peptide (CGRP), and substance P (SP) are neuropeptides that modulate pain responses transmitted by primary sensory afferents, the cell bodies of which are located in the dorsal root ganglion (DRG). Thus, the goal of the present study was to determine whether the diabetic condition is associated with altered neuropeptide gene expression in lumbar DRG of the rat. We employed an established animal model in which streptozotocin (STZ, 55 mg/kg) is administered to 6 week-old rats. The hallmark symptoms of hyperglycemia (blood glucose > 400 mg/dl), polydipsia, polyuria, and severe weight loss were maximal at 6 weeks postadministration, at which time animals were sacrificed. For determination of peptide encoding mRNAs distributed in DRG neurons, in situ hybridization histochemistry utilizing S-end-labeled oligonucleotides complimentary to sequences of preprosomatostatin (PPSOM), preprocalcitonin gene related peptide (PPCGRP), preprotachykinin (PPT), or preproneuropeptide Y (PPNPY) mRNA was performed. Silver grains were detected overlying DRG cells by autoradiography on sections of tissue counterstained with thionin. Semiquantitative analysis of differences in silver grain signal were made using an image analysis system, which expressed signals as fCi/microns2. In diabetic rats there was a significant decrease in DRG PPSOM (54%, p < 0.01), and PPCGRP (33%. p < 0.05) mRNA hybridization from the normal values PPT mRNA hybridization signal and SP-like immunoreactivity were not significantly changed in diabetic rat DRGs compared to control. In contrast, there was an increase in the number of cells labeled with PPNPY hybridization in DRG from diabetic rats. These data suggest that CGRP and SOM synthesis in primary sensory neurons is reduced in STZ-induced diabetic rats. These changes could contribute to the painful neuropathies and altered inflammatory responses seen in diabetes mellitus.

Serotonergic regulation of renin and prolactin secretion.

Drugs that, directly or indirectly produce activation of serotonin (5-HT) receptors increase plasma concentrations of both prolactin and renin. The serotonergic regulation of prolactin and renin secretion share several common characteristics. Serotonergic neurons originating in the dorsal raphe and terminating in the hypothalamus stimulate the secretion of both prolactin and renin. Destruction of cells in the hypothalamic paraventricular nucleus (PVN) inhibits both the prolactin and renin responses to 5-HT agonists and 5-HT-releasing drugs. Activation of 5-HT2 receptors increases the secretion of both prolactin and renin, while activation of other 5-HT receptor subtypes has differential effects on these hormones. However, there are also differences between the serotonergic mechanisms that regulate the secretion of prolactin and renin. Activation of 5-HT1A receptors increases the secretion of prolactin but not of renin. In addition, activation of peripheral 5-HT2 receptors stimulates the secretion of renin, while activation of peripheral 5-HT3 receptors increases plasma levels of prolactin but not renin. In humans, the effect of 5-HT-releasing drugs and 5-HT agonists on plasma prolactin concentrations has been studied to a greater extent than effects on most other hormones. In contrast, the renin response to 5-HT agonists and 5-HT releasers has not been well characterized in humans. Because of the important role of the renin-angiotensin system in cardiovascular regulation, studies on the serotonergic regulation of renin release in humans could increase our understanding of cardiovascular disorders associated with altered serotonergic function. Examples include anxiety and consequences of cocaine abuse. In conclusion, comparing the serotonergic regulation of prolactin and renin secretion indicates similarities that might shed light on common brain mechanisms that regulate neuroendocrine function.

Antisense inhibition of protein synthesis and function : rabbit retinal protein.

When considering the use of antisense technology for in vivo apphcatron, whether for therapeutic development or for the creation of animal models for human diseases, a major problem is that most extracellular compartments are constantly mixing with the blood, lymph, cerebrospinal fluid (CSF), and so forth, and thus constantly diluting the oligonucleotide. A possibly unique exception to this problem is the vitreous, a gelatinous fluid overlying the inner retinal surface. The vitreous is formed early in development and is in essence a relatively closed compartment with no active transport of fluids or ions. Forensic pathologists make use of this fact in determining the composition of electrolytes at the time of death by sampling the vitreal compartment.

Hypothalamic paraventricular, but not supraoptic neurons, mediate the serotonergic stimulation of oxytocin secretion.

The purpose of the present studies was to determine whether cells in the hypothalamic paraventricular (PVN) or supraoptic (SON) nuclei mediate the serotonergic stimulation of oxytocin secretion. The serotonergic stimulus consisted of injection of the 5-HT-releasing drug p-chloroamphetamine (8 mg/kg, IP). The validity of this approach was verified by comparing this drug with another 5-HT releaser, d-fenfluramine (5 mg/kg, IP). Both 5-HT releasers increased plasma oxytocin concentration. Furthermore, the 5-HT uptake blocker fluoxetine (10 mg/kg, IP) blocked the effects of both p-chloroamphetamine and d-fenfluramine on plasma oxytocin concentrations, suggesting that both 5-HT releasers must be taken up through the 5-HT transporter into 5-HT nerve terminals to increase oxytocin secretion. In the lesion experiments, cells in the hypothalamic PVN or SON were destroyed by injection of the cell-selective neurotoxin ibotenic acid. The PVN lesions reduced basal levels and inhibited the effect of p-chloroamphetamine (8 mg/kg, IP) on plasma oxytocin concentration. In contrast, SON lesions did not alter basal oxytocin levels and did not reduce the oxytocin response to p-chloroamphetamine, suggesting that the SON is not involved in the serotonergic stimulation of oxytocin secretion. Site specificity of the PVN lesions was confirmed when injections of ibotenic acid into the hypothalamic dorsomedial nucleus (DMN), immediately caudal to the PVN, potentiated the oxytocin response to p-chloroamphetamine, suggesting that the DMN exerts an inhibitory influence on the secretion of oxytocin. Taken together, the data suggest that the serotonergic stimulation of oxytocin secretion involves PVN, but not SON, oxytocin neurons.

Evidence that ACTH secretion is regulated by serotonin2A/2C (5-HT2A/2C) receptors.

The present study characterized the serotonin (5-HT) receptor subtypes mediating adrenal corticotropic hormone (ACTH) and corticosterone responses to 5-HT agonists in conscious rats. The 5-HT2A/5-HT2C agonist (+/-(-)1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane HC1 (DOI) increased plasma ACTH and corticosterone in a dose-dependent manner. The 5-HT2A/5-HT2C antagonist ritanserin (0.01 and 0.1 mg/kg sc) inhibited the DOI-induced increase in plasma ACTH, but not corticosterone. Low doses of spiperone (0.01 and 0.1 mg/kg sc) significantly reduced the ACTH response to DOI. Because spiperone has a higher affinity for 5-HT2A than 5-HT2C receptors, these data suggest that DOI stimulates ACTH secretion through 5-HT2A receptors. 5-methoxy-3-[1,2,3,4-tetrahydro-4-pyridinyl]-1H-indole (RU 24969) is a potent 5-HT1A/1B and moderate 5-HT2C agonist that also has been suggested to release 5-HT. However, p-chlorophenylalanine (PCPA) did not reduce the effect of RU 24969 on plasma ACTH, suggesting that RU 24969 only acts as a direct agonist. 6-methyl-1-[1-methylethyl]ergoline-8-carboxylic acid (LY53857) injected into the lateral cerebral ventricles (i.c.v.) inhibited the ACTH, but not corticosterone response to peripheral injection of RU 24969, suggesting that central 5-HT2A/2C receptors mediate the ACTH response. LY53857 injection (i.c.v.) also inhibited the effect of p-chloroamphetamine (i.c.v.) on plasma ACTH. However, the corticosterone response was not inhibited by LY53857, suggesting a distinct location of 5-HT receptors regulating corticosterone secretion.(ABSTRACT TRUNCATED AT 250 WORDS)

Central stimulation of renin secretion through serotonergic, noncardiovascular mechanisms.

The aims of the study were to determine; (1) whether activation of serotonin (5-HT) receptors in the brain increases renin secretion, and (2) whether the hypertensive effects of a 5-HT agonist and 5-HT releaser obscure their ability to stimulate renin release. Various drugs that increase serotonergic neuro-transmission can activate the secretion of renin from the kidneys. Many of these drugs can also elevate blood pressure. Changes in blood pressure can alter renin secretion by activating renal baroreceptor mechanisms, so that a decrease in perfusion pressure will increase renin secretion and vice versa. To address the first objective, the 5-HT agonist RU 24969 (0, 10, 100 and 200 micrograms/kg) and the 5-HT releaser p-chloroamphetamine (0, 50, 500 and 1,000 micrograms/kg) were injected intracerebroventricularly (ICV) at doses lower than those that are peripherally effective. ICV injection of RU 24969 dose-dependently increased plasma levels of renin. ICV injection of the 5-HT2A/5-HT2C antagonist LY53857 (50 micrograms/kg) inhibited the renin response to peripherally injected RU 24969 (0, 1, 5 and 10 mg/kg i.p.), suggesting that 5-HT2A/5-HT2C receptors in the brain mediate the effect of peripherally injected RU 24969 on renin secretion. In contrast, ICV injection of p-chloroamphetamine decreased renin secretion. To determine whether hypertensive actions could account for the differences between RU 24969 and p-chloroamphetamine, we measured the effects of both p-chloroamphetamine and RU 24969 on blood pressure and heart rate. ICV injection of p-chloroamphetamine (1,000 micrograms/kg) produced a large rise of 44 mm Hg at 2 min and 25 mm Hg at 5 min after injection, while ICV injection of RU 24969 (200 micrograms/kg) caused a slower and smaller blood pressure elevation of 18 mm Hg at 5 min after injection. To determine whether the hypertensive effects of both RU 24969 and p-chloroamphetamine could mask their effects on renin secretion, rats were pretreated with the alpha 1 antagonist prazosin. Administration of prazosin (1 mg/kg s.c.), which prevents the hypertensive effects of p-chloroamphetamine, exposed a stimulatory effect of ICV-injected p-chloroamphetamine (500 micrograms/kg) on renin secretion and potentiated the effect of RU 24969 (5 mg/kg i.p.) on renin release. In conclusion, these data suggest that both RU 24969 and p-chloroamphetamine increase renin secretion through central 5-HT receptors, and that these effects are partially obscured by their hypertensive actions.

ICV injection of the serotonin 5-HT1B agonist CP-93,129 increases the secretion of ACTH, prolactin, and renin and increases blood pressure by nonserotonergic mechanisms.

This study tested whether a new serotonin (5-HT1B) agonist, 3-(1,2,5,6-tetrahydro-4-pyridyl)-5-propoxy-pyrrolo[3,2-b]pyridine (CP-93,129), could be used to study the potential role of 5-HT1B receptors in the secretion of adrenocorticotropic hormone (ACTH), prolactin, and renin. CP-93,129 has a high affinity for 5-HT1B receptors but low affinity for other 5-HT receptor subtypes. In addition, CP-93,129 does not readily cross the blood-brain barrier. The secretion of ACTH, prolactin, and renin is known to be increased after activation of 5-HT receptors. ICV injections of CP-93,129 (100 micrograms/kg) increased the plasma concentrations of ACTH, prolactin, and renin. CP-93,129 also increased blood pressure and reduced heart rate. To determine whether these effects of CP-93,129 are centrally mediated, we compared them with IP injection of the same dose of CP-93,129. IP-injected CP-93,129 did not alter blood pressure or heart rate and did not elevate plasma prolactin and renin concentrations. To determine whether 5-HT1B receptors mediate the central effects of CP-93,129, rats were pretreated with the 5-HT antagonists l-propranolol or metergoline prior to ICV injections of doses of CP-93,129 (0-100 micrograms/kg). The 5-HT1A/1B/2A/2C antagonist metergoline (0.5 mg/kg, IP) failed to inhibit the CP-93,129-induced elevation of ACTH, prolactin, or renin concentrations. In contrast, the 5-HT1A/1B/beta antagonist l-propranolol (20 micrograms/kg, ICV) inhibited the renin but not the ACTH or prolactin responses to ICV CP-93,129.(ABSTRACT TRUNCATED AT 250 WORDS)

Influence of repeated cocaine exposure on the endocrine and behavioral responses to stress in rats.

Previous studies have determined that chronic cocaine exposure inhibits the serotonergic stimulation of hormone secretion. The present experiments were conducted to determine whether the endocrine responses to stress could be a useful approach to assess the influence of cocaine exposure on neuronal function. Male rats received twice daily injections of cocaine (1-15 mg/kg, IP) for 7 days. Animals were subsequently exposed to different stressors, i.e. conditioned emotional stress utilizing a low (0.5 mA) or high (1.5 mA) intensity footshock during training, or to immobilization stress. Immediately after the stress procedures, blood samples were collected for radioimmunoassay of plasma corticosterone, prolactin, and renin concentrations. Repeated cocaine exposure attenuated the stress-induced elevations of corticosterone and prolactin secretion, and attenuated some of the behavioral effects of the low intensity conditioned emotional stress. When exposed to the high intensity conditioned emotional stress, cocaine did not alter the endocrine or behavioral effects of stress. Finally, repeated cocaine exposure modified the immobilization stress-induced elevation of renin secretion; low doses of cocaine (1 or 5 mg/kg) attenuated, while higher doses (10 mg/kg) potentiated the renin response to immobilization stress. Thus, the influence of repeated cocaine exposure on the endocrine and behavioral responses to stress appears to depend upon the type and intensity of the stressor. Compared with previous studies which found altered neuroendocrine responses to serotonin releasers and agonists following cocaine exposure, the hormonal responses to stress are less consistently modified by cocaine.

Repeated exposure to cocaine produces long-lasting deficits in the serotonergic stimulation of prolactin and renin, but not adrenocorticotropin secretion.

To determine whether cocaine-induced deficits in serotonergic function are long-lasting, the neuroendocrine responses to the serotonin (5-hydroxytryptamine, 5-HT) releaser, p-chloroamphetamine were evaluated 1-8 weeks subsequent to 7 days of cocaine exposure (15 mg/kg b.i.d.). In cocaine-pretreated rats, the p-chloroamphetamine-induced elevations of prolactin and renin secretion were significantly reduced for 8 and 4 weeks, respectively. In contrast, the p-chloroamphetamine-induced elevation of adrenocorticotropic hormone (ACTH) secretion was at control values 1 week after cocaine exposure. The data suggest that some cocaine-induced deficits in serotonergic function are long-lasting.

Long-term treatment with the antidepressants fluoxetine and desipramine potentiates endocrine responses to the serotonin agonists 6-chloro-2-[1-piperazinyl]-pyrazine (MK-212) and (+-)-1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane HCl (DOI).

Various endocrine responses to 5-hydroxytryptamine (serotonin, 5-HT) agonists were used to assess serotonergic receptor function after chronic treatment with the antidepressants fluoxetine (10 mg/kg), a 5-HT uptake blocker and the norepinephrine uptake blocker desipramine (DMI, 5 mg/kg). Both were injected (i.p.) once a day for 21 days. DOI (5-HT1C/2 agonist, 0-5 mg/kg i.p.) and 6-chloro-2-[1-piperazinyl]-pyrazine (MK-212) (less selective, but predominantly a 5-HT1C agonist, 0-20 mg/kg i.p.) were administered 18 hr after the final antidepressant injection and 30 min before decapitation. Chronic treatment with both fluoxetine and DMI produced a potentiation in most hormone responses to the 5-HT agonists (+-)-1-(2,5-dimethoxy-4-iodophenyl)-2-amino-propane HCl (DOI) and MK-212, although there were several differences in individual hormone responses to the two 5-HT agonists. Fluoxetine and DMI potentiated the MK-212- and DOI-induced increase of plasma oxytocin levels and potentiated the effect of DOI on plasma adrenocorticotropic hormone (corticotropin) and prolactin levels. In contrast, the effect of the high dose of MK-212 on plasma prolactin concentration was reduced by both antidepressants. Only MK-212 increased vasopressin levels and this effect was potentiated by fluoxetine, but not by DMI. Fluoxetine also significantly increased the resting level of plasma vasopressin. DMI potentiated the effect of MK-212 on plasma renin concentration. Pretreatment with fluoxetine significantly increased (38%) the Bmax for the 5-HT1C/2 agonist sites ([125I]DOI) in the hypothalamus.(ABSTRACT TRUNCATED AT 250 WORDS)

Neurons in the hypothalamic paraventricular nucleus mediate the serotonergic stimulation of prolactin secretion via 5-HT1c/2 receptors.

These studies examined the hypothalamic site and receptor subtype mediating the serotonergic (5-HT) control of PRL secretion in conscious male rats. Initially, we characterized the pharmacology of the 5-HT releaser and 5-HT agonists that increase PRL release. Subsequently, we performed lesion experiments to locate the 5-HT receptors involved in PRL secretion. p-Chloroamphetamine, a 5-HT releaser, is postulated to enter serotonergic nerve terminals through the 5-HT uptake mechanism, which can be inhibited by fluoxetine. p-Chloroamphetamine (8 mg/kg, ip) increased the plasma PRL concentration approximately 6-fold. The 5-HT uptake inhibitor fluoxetine almost completely prevented this increase, demonstrating that p-chloroamphetamine increases PRL release via a serotonergic mechanism. The 5-HT1C/5-HT2 agonist +(-)-1-(2,5-dimethoxy-4-iodophenyl)2-aminopropane HCl (ip) produced a strong (30-fold) dose-dependent elevation of plasma PRL, which was virtually eliminated by 0.1 mg/kg (sc) ritanserin, a 5-HT1C/5-HT2 antagonist. +(-)-1-(2,5-Dimethoxy-4-iodophenyl)2-aminopropane HCl injected intracerebroventricularly (icv) in doses below those that were peripherally effective also produced a significant (8-fold) increase in PRL secretion that was again attenuated by icv pretreatment with ritanserin (2 micrograms/kg). RU 24969 (5-methoxy-3-[1,2,3,4-tetrahydro-4-pyridinyl]1H-indole) was reported to act as both a 5-HT releaser and a direct postsynaptic 5-HT agonist. To test whether RU 24969 releases 5-HT to increase PRL secretion, we depleted 5-HT stores with the 5-HT synthesis inhibitor p-chlorophenylalanine. The ability of RU 24969 (0.5, 1, 5, and 10 mg/kg, ip) to elevate PRL secretion was not inhibited by pretreatment with p-chlorophenylalanine, suggesting that RU 24969 stimulates PRL secretion only through activation of postsynaptic 5-HT receptors. To test whether RU 24969 acts centrally, it was injected either icv, through chronic icv cannulae, or peripherally (ip). RU 24969 injected icv significantly stimulated PRL secretion (11-fold) at doses 500-fold lower than the peripherally effective doses (10 micrograms/kg vs. 5 mg/kg), suggesting a role for central 5-HT receptors in the regulation of PRL secretion. In addition, rats pretreated with the 5-HT1C/5-HT2 antagonist LY53857 (icv) significantly inhibited the PRL response if RU 24969 was injected ip, but not icv. The results of these experiments suggest that 5-HT1C or 5-HT2 receptors in the brain participate in the serotonergic stimulation of PRL secretion.(ABSTRACT TRUNCATED AT 400 WORDS)

Investigation of the role of 5-HT3 receptors in the secretion of prolactin, ACTH and renin.

The potential roles of central and peripheral 5-HT3 receptors in the secretion of prolactin, adrenocorticotropic hormone (ACTH), corticosterone and renin was investigated. Male rats received the 5-HT3 antagonist ondansetron (0, 0.1 or 1 mg/kg i.p.), 30 min prior to injections of the serotonin (5-HT) releaser, p-chloroamphetamine (PCA; 0, 3 or 8 mg/kg i.p.). Blood samples were collected 60 min after PCA for radioimmunoassays of plasma prolactin, ACTH, corticosterone and renin concentrations. PCA significantly elevated secretion of each of these hormones. Pretreatment with the 5-HT3 antagonist, ondansetron, significantly attenuated the PCA-induced elevation of prolactin secretion, suggesting that 5-HT3 receptors contribute to the serotonergic stimulation of prolactin secretion. Ondansetron did not modify effects of PCA on ACTH, corticosterone or renin secretion. To determine whether the 5-HT3 receptor role in prolactin secretion is mediated in the brain, the endocrine effects of intracerebroventricular (i.c.v.) injections of 5-HT (30 micrograms/kg) or the 5-HT3 agonist, 2-methylserotonin (1, 20 or 200 micrograms/kg) were evaluated. Both 5-HT and 2-methylserotonin significantly elevated plasma prolactin levels 15 min postinjection. However, ondansetron (1 mg/kg i.p.) did not antagonize these actions. Both 5-HT and 2-methylserotonin also increased plasma ACTH and corticosterone concentrations. Finally, 5-HT suppressed, while 2-methylserotonin stimulated renin secretion. None of the hormonal effects of i.c.v. injected 5-HT or 2-methylserotonin were altered by ondansetron. Thus, the results suggest that peripheral, but not central 5-HT3 receptors are involved in the stimulation of prolactin secretion. Furthermore, 5-HT3 receptors do not mediate the serotonergic stimulation of ACTH, corticosterone, or renin secretion.

Ibotenic acid lesions in the bed nucleus of the stria terminalis attenuate conditioned stress-induced increases in prolactin, ACTH and corticosterone.

The contribution of the bed nucleus of the stria terminalis (BST) to the expression of stress-induced increases in ACTH/corticosterone, prolactin and renin secretion was assessed. Neurons in the lateral part of the BST were destroyed with bilateral injections of the cell-selective neurotoxin ibotenic acid (1.5 micrograms in 0.1 microliter of solution per side). Two weeks later, the rats were stressed using an immobilization or conditioned stress paradigm. Rats with lesions in the lateral part of the BST showed attenuated ACTH and corticosterone responses to conditioned stress. Bilateral ablation of lateral BST significantly reduced the prolactin secretory response to conditioned stress. The same lesions had no effect upon plasma increases in renin that occur in response to conditioned stress. Also, destruction of neurons in the BST did not affect immobilization-induced increases in ACTH, corticosterone, prolactin or renin. Previous studies have demonstrated that ibotenic acid lesions in the central amygdala reduce corticosterone and renin response to conditioned stress. Thus, both the BST and central amygdala are important for the adrenocortical response to conditioned stress. Neurons in the central nucleus of the amygdala are part of the circuitry that mediates renin responses to conditioned stress. Neurons in the BST are important for the full expression of prolactin responses to conditioned stress. The neuronal circuitry and stressor specificity in the mediation of prolactin, renin and ACTH/corticosterone responses are discussed.

Neurons in the hypothalamic paraventricular nucleus mediate the serotonergic stimulation of renin secretion.

The aim of the present study was to resolve which hypothalamic nucleus is necessary for the serotonergic control of renin secretion. RU 24969 is considered a serotonin (5-HT1A/5-HT1B) agonist, while p-chloroamphetamine is a 5-HT releaser. Both drugs reliably elevate plasma levels of renin when injected peripherally. Previous studies suggest that serotonergic neurons, projecting to the hypothalamus, mediate the effect of p-chloroamphetamine on renin secretion. Discrete cell-selective lesions were made with ibotenic acid in three hypothalamic sites: the paraventricular, the dorsomedial or the ventromedial nuclei. Two weeks after surgery rats were injected with RU 24969 (5 mg/kg, i.p.) or p-chloroamphetamine (8 mg/kg, i.p.). The renin response to both RU 24969 and p-chloroamphetamine was significantly reduced in rats with histologically verified paraventricular lesions compared to vehicle treated controls. In contrast, the renin response to p-chloroamphetamine remained unchanged in rats with either dorsomedial or ventromedial hypothalamic lesions. Thus, these results are consistent with the hypothesis that 5-HT receptors located on cell bodies in the paraventricular nucleus mediate the renin response to a serotonin agonist and releaser. Furthermore, they confirm previous studies that suggest that 5-HT neurons regulate renin secretion through central receptors.

Neuroendocrine responses to the serotonin2 agonist DOI are differentially modified by three 5-HT1A agonists.

Evidence suggests that activation of 5-HT1A receptors leads to inhibition of 5-HT2-mediated behavior. The purpose of this study was to investigate the interaction between 5-HT1A and 5-HT2 receptor-mediated hormone secretion. Rats were pretreated with the 5-HT1A agonists buspirone (0, 0.5, 2.0 mg/kg, i.p.), 8-OH-DPAT (0, 0.05, 0.2 mg/kg, s.c.) or ipsapirone (0, 1.0, 2.5 mg/kg, i.p.), 45 min before decapitation. The 5-HT2 agonist DOI was administered (0-10 mg/kg, i.p.) 15 min after injection of the 5-HT1A agonists. The three 5-HT1A agonists differentially altered the DOI-induced increase of concentrations of hormone in plasma. None of the three 5-HT1A agonists influenced the basal levels of renin, ACTH and prolactin but 8-OH-DPAT and buspirone increased the basal level of corticosterone in plasma. Also, 8-OH-DPAT increased the effects of DOI on the concentration of ACTH in plasma but ipsapirone and buspirone did not. None of the 5-HT1A agonists significantly affected DOI-induced increase of concentration of corticosterone in plasma. Buspirone and 8-OH-DPAT potentiated the effect of DOI on prolactin in plasma, but ipsapirone did not. Ipsapirone potentiated the effect of DOI on the concentration of renin in plasma but this effect was not observed in 8-OH-DPAT- and buspirone-pretreated rats. The results do not support the hypothesis for a functional interaction between 5-HT1A and 5-HT2 receptors, since the three 5-HT1A agonists did not have the same influence on the hormonal effects of DOI.