Effect of L-dopa and bilateral nephrectomy on the aldosterone response to metoclopramide

The dopaminergic antagonist, metoclopramide (MCP) causes an increase in plasma aldosterone (PA) by a processnot well delineated. To investigate the mechanism of action of metoclopramide (MCP), studies were performed in rats after pre-treatment with L-dihydroxy-phenylalanine (L-dopa) and after bilateral nephrectomy. Intra-arterial MCP (200 μg/kg) resulted in a significant elevation in PA and prolactin (PRL) at 5 min and plasma renin activity (PRA) at 10 min without altering serum potassium levels. Pre-administration of L-dopa (30 mg/kg) delayed and markedly blunted PA, PRL and PRA resonses to MCP. In 7 rats, studied 30 hours after bilateral nephrectomy, the PRA was measurable (2.5 ± 0.4 ng/ml h^?1) but displayed no response to MCP. In contrast, the PA and PRL responses to MCP were not significantly affected. L-dopa induced suppression of PRA and PA was prevented by pre-administration of MCP. These results suggest that dopaminergic modulation of PA secretion occurs independently of the renin-angiotensin system.     

The effect of mescaline, 3, 4-dimethoxyphenethylamine and 2, 5-dimethoxy-4-methylamphetamine on rat plasma prolactin: evidence for serotonergic mediation.

Mescaline, 3,4-dimethoxyphenethylamine (DMPEA) or 2,5-dimethoxy-4-methylamphetamine (DOM) was administered to male Sprague-Dawley rats, alone or in combination with para-chlorophenylalanine (PCPA), an inhibitor of serotonin (5-HT) synthesis, or methysergide, a 5-HT receptor blocker. All three compounds increased plasma prolactin (PRL) levels. These increases were potentiated by PCPA and blocked by methysergide. Pretreatment with alpha-methylparatyrosine (AMPT), an inhibitor of catecholamine synthesis, resulted in an increase in plasma PRL equal to the additive effects of the independent administration of mescaline, DMPEA, or DOM plus the AMPT-induced response. The results suggest that mescaline, DMPEA and DOM may be exerting their effects on rat plasma PRL through direct stimulation of serotonin receptors. These results further demonstrate the ability of PCPA to rapidly induce supersensitivity of the 5-HT receptors which stimulate PRL secretion.     

Prolactin receptors on human T and B lymphocytes: antagonism of prolactin binding by cyclosporine

Prolactin (PRL) receptors have been identified recently on human peripheral blood mononuclear cells (MNC) and may be involved in the regulation of cell-mediated immunity. Cyclosporine (CsA), an immunosuppressive cyclic endecapeptide utilized to prolong graft survival in human organ transplant patients, affects PRL binding to MNC. At concentrations of CsA from 10(-10) through 10(-8) M, the amount of PRL bound to MNC markedly increased to ca. 400% of controls, whereas CsA concentrations of 10(-6) and 10(-5) M totally inhibited PRL binding to lymphocytes. The ability of low concentrations of CsA to enhance PRL binding was temperature-dependent and did not occur when binding assays were conducted at 4 degrees C. PRL displaced [3H]CsA from lymphocytes with ca. 50% displacement at 10(-9) M PRL and total displacement at concentrations of 10(-7), 10(-6), and 10(-5) M. Growth hormone did not displace [3H]CsA in similar experiments. CsA also did not alter the binding of a beta-receptor antagonist to MNC, again suggesting that CsA was specific in its antagonism of PRL binding. A CsA analog with no immunosuppressive action, cyclosporin H, did not alter PRL binding to MNC. Furthermore, PRL receptors were demonstrated on four cell lines of human and mouse origin. Finally, PRL receptors were identified on purified populations of T and B lymphocytes isolated from human spleens, and CsA again inhibited PRL binding at concentrations of 10(-7) and 10(-6) M. The presence of PRL receptors on T and B lymphocytes suggests that PRL may be involved in the regulation of humoral and cell-mediated immunity, and that one effect of CsA on immune function may be its ability to inhibit the effects of PRL action on these lymphocytes.     

Dopamine control of aldosterone secretion in end-stage renal failure

The role of the tonic inhibitory effect of dopamine on aldosterone secretion has been investigated in 10 patients with chronic renal failure (CRF) on hemodialysis, in 8 normotensive renal transplant recipients (Tx) with normal renal function and in 8 normotensive volunteers (NV). The following tests were performed: the response of plasma aldosterone (PA) to metoclopramide administration; the response of plasma prolactin (PRL) to TRH administration, and the changes induced by Lisuride (a dopaminergic agonist, on the values of PA and PRL). The basal values of PA and PRL were higher in CRF than in NV and Tx. The inverse was true for plasma renin activity (PRA) values. The response of PA and PRL to metoclopramide showed blunted increases in CRF when compared to NV, in the absence of changes of PRA, cortisol and potassium. After TRH administration, PRL increase in CRF was also inferior. Lisuride induced a decrease of both PA and PRL both in CRF and NV. In Tx, basal values of PA and PRL were similar to NV. Nevertheless, the response to metoclopramide and TRH were partially blunted when compared to that of NV. These results point to the existence of a deranged dopaminergic regulation of aldosterone secretion in end-stage renal failure patients. The alterations are partially corrected by a well-functioning kidney graft.     

Effects of venlafaxine on extracellular 5-HT,dopamine and noradrenaline in the hippocampus and on peripheral hormone concentrations in the rat in vivo

The purpose of the present study was to study the effect of an acute dose of the serotonin (5-HT) - noradrenaline (NA) reuptake inhibitor venlafaxine on extracellular concentrations of 5-HT, NA and dopamine (DA) in the hippocampus and on the peripheral hormone concentrations in freely moving rats. Blood obtained from a catheter placed in the vena femoralis was analyzed for adrenocorticotropin (ACTH), beta-endorphins, prolactin (PRL), growth hormone (GH) and cortisol. Collections are referred to pre and post injection of 20 mg/kg of venlafaxine. Extracellular hippocampal NA and 5-HT as determined with in vivo microdialysis increased significantly after drug injection. PRL and ACTH were significantly affected by the drug. At the selected dose venlafaxine is able to increase the release of 5-HT but also of NA in rat hippocampus. Due to the dual reuptake properties of the drug and the functional interconnection of the NA and the 5-HT systems, the observed effects on peripheral hormones are possibly mediated by a combined action of these 2 systems.     

Possible direct effect of serotonin on pituitary prolactin secretion: in vivo and in vitro studies

In this work we analyze the possibility of serotonin (5-HT)-releasing prolactin (PRL) through a direct action at the pituitary level. 5-HT (2 mg/kg i.v.) stimulates PRL secretion in hypophysectomized autotransplanted animals (HAG) significantly and this effect was not influenced by pretreatment with the dopaminergic antagonist domperidone. In perifused pituitaries, 5-HT administration (0.01, 0.1 and 1 microM for 90 min, or 1, 10, 100 microM for 15 min) was ineffective in stimulating PRL release. In pituitaries obtained from animals previously treated with the neurotoxic 5,7-dihydroxytryptamine (5,7-DHT) or vehicle and incubated in the presence of 5-HT (2.5, 5 and 10 microM), no response in PRL secretion was observed. These results suggested that 5-HT does not release PRL through a direct pituitary action, and that the effect observed in HAG animals could be mediated through the release of a PRL-releasing factor after 5-HT administration.     

Novel class of arylpiperazines containing N-acylated amino acids: their synthesis, 5-HT1A, 5-HT2A receptor affinity, and in vivo pharmacological evaluation

Novel arylpiperazines with N-acylated amino acids, selected on the basis of a preliminary screening of two libraries previously synthesized on SynPhase Lanterns, were prepared in solution and their affinity for 5-HT(1A), 5-HT(2A), and D(2) receptors was evaluated. The compounds bearing (3-acylamino)pyrrolidine-2,5-dione (19-26) and N-acylprolinamide (29-34) moieties showed high affinity for 5-HT(1A) (K(i)=3-47 nM), high-to-low for 5-HT(2A) (K(i)=4.2-990 nM), and low for D(2) receptors (K(i)=0.77-21.19 microM). All the new o-methoxy derivatives of (3-acylamino)pyrrolidine-2,5-diones tested in vivo revealed agonistic activity at postsynaptic 5-HT(1A) receptors, while m-chloro derivatives were classified as antagonists of these sites; similar relations were observed for o-methoxy (29) and m-chlorophenylpiperazine derivatives of N-acylprolinamides. The reported results show that the amino acid-derived terminal fragment modified the in vivo functional profile. Finally, the selected compounds 19 and 20, a 5-HT(1A) partial agonist and a full agonist, respectively, and 26, a mixed 5-HT(1A)/5-HT(2A) antagonist, were evaluated in preclinical animal models of depression and anxiety. The project allowed selecting the lead compound 20 which exhibited an anxiolytic-like effect in the four-plate test in mice and revealed distinct antidepressant-like effects in the forced swimming and tail suspension tests in mice.     

Mechanisms for the stimulatory effects of opioidergic and serotonergic input signals on prolactin in pregnant rats.

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

Plasma aldosterone, renin activity, and cortisol responses to heat exposure in sodium depleted and repeleted subjects.

The effect of 90-min heat exposure (46 degrees C, 35 mbar) on plasma aldosterone (PA) patterns was studied and the respective roles of plasma renin activity (PRA), adrenocorticotropin (ACTH), Na+ and K+ concentrations in the control of PA response were in investigated in eight subjects on a low sodium diet and in five subjects on a high sodium diet. In all subjects, transitory PA increases of varying importance were observed, which were not related to sweat losses (less than 1% bodyweight) or to rectal temperature rise. In sodium-repleted subjects, basal PA and PRA levels as well as heat-induced rises were low (mean PA peak level = 12.62 +/- 1.15 ng/100 ml). They were enhanced by sodium depletion and PA reached a mean peak level of 34.07 +/- 2.73 ng/100 ml. But, in both conditions, the heat-induced PA peaks were 3-times higher than the initial levels. PA correlated with PRA in all but one of the sodium-repleted subjects and in 6 of the 8 sodium-depleted subjects. ACTH release, as measured by plasma cortisol (PC) levels, occurred in those subjects who noted an increased feeling of annoyance and discomfort. Thus, PA correlated positively with PC in 4 sodium-depleted subjects. A high sodium intake improved heat-tolerance. Plasma K+ and Na+ concentrations were not significantly modified by exposure to heat. PA increases can occur without concomitant changes in PRA, PC, K+ or Na+, which suggests that an additional factor may play a role in aldosterone regulation during acute heat exposure.     

The 5-hydroxytryptamine stimulated formation of inositol phosphate is inhibited in platelets from alcoholics

The accumulation of inositol monophosphate (IP1) was measured after stimulation of 5-hydroxytryptamine2 (5-HT2) receptors on platelets from alcoholics and healthy controls. In controls, 5-HT induced a dose-dependent response with an EC50 = 2 x 10(-6) M and a maximal response at 10(-5) M. Ritanserin, a selective 5-HT2 antagonist, markedly reduced the accumulation. The IP1 formation after stimulation by 10(-5) M 5-HT was significantly impaired in platelets from alcoholics as compared to controls. This study indicates that the 5-HT2 receptor function is inhibited in alcoholics. It also illustrates the possibility of using IP1 formation in peripheral cells as a mean of studying receptor function in disease.     

Fenfluramine-induced pulmonary vasoconstriction: role of serotonin receptors and potassium channels.

The anorexic agent fenfluramine considerably increases the risk of primary pulmonary hypertension. The mechanism of this effect is unknown. The appetite-reducing action of fenfluramine is mediated by its interaction with the metabolism of serotonin [5-hydroxytryptamine (5-HT)] in the brain. We tested the hypothesis that the pulmonary vasoconstrictive action of fenfluramine is at least in part mediated by 5-HT receptor activation. In addition, we sought to determine whether pharmacological reduction of voltage-gated potassium (K(V)) channel activity would potentiate the pulmonary vascular reactivity to fenfluramine. Using isolated rat lungs perfused with Krebs-albumin solution, we compared the inhibitory effect of ritanserin, an antagonist of 5-HT(2) receptors, on fenfluramine- and 5-HT-induced vasoconstriction. Both 5-HT (10(-5) mol/l) and fenfluramine (5 x 10(-4) mol/l) caused significant increases in perfusion pressure. Ritanserin at a dose (10(-7) mol/l) sufficient to inhibit >80% of the response to 5-HT reduced the response to fenfluramine by approximately 50%. A higher ritanserin dose (10(-5) mol/l) completely abolished the responses to 5-HT but had no more inhibitory effect on the responses to fenfluramine. A pharmacological blockade of K(V) channels by 4-aminopyridine (3 x 10(-3) mol/l) markedly potentiated the pulmonary vasoconstrictor response to fenfluramine but was without effect on the reactivity to 5-HT. These data indicate that the pulmonary vasoconstrictor response to fenfluramine is partly mediated by 5-HT receptors. Furthermore, the pulmonary vasoconstrictor potency of fenfluramine is elevated when the K(V)-channel activity is low. This finding suggests that preexisting K(V)-channel insufficiency may predispose some patients to the development of pulmonary hypertension during fenfluramine treatment.     

Mutagenesis analysis of the serotonin 5-HT2C receptor and a Caenorhabditis elegans 5-HT2 homologue: conserved residues of helix 4 and helix 7 contribute to agonist-dependent activation of 5-HT2 receptors

An alignment of serotonin [5-hydroxytryptamine (5-HT)] G protein-coupled receptors identified a lysine at position 4.45 (helix 4) and a small polar residue (serine or cysteine) at 7.45 (helix 7) that occur exclusively in the 5-HT2 receptor family. Other serotonin receptors have a hydrophobic amino acid, typically a methionine, at 4.45 and an invariant asparagine at 7.45. The functional significance of these class-specific substitutions was tested by site-directed mutagenesis of two distantly related 5-HT2 receptors, Caenorhabditis elegans 5-HT2ce and rat 5-HT2C. Residues 4.45 and 7.45 were each mutated to a methionine and asparagine, respectively, or an alanine and the resulting constructs were tested for activity. A K4.45M mutation decreased serotonin-dependent activity (Emax) of the rat 5-HT2C receptor by 60% and that of the C. elegans homologue by 40%, as determined by a fluorometric plate-based calcium assay. The rat mutant also exhibited nearly sixfold higher agonist binding affinity and significantly lower constitutive activity compared with wildtype. Mutagenesis of S7.45 in the C. elegans receptor increased serotonin binding affinity by up to 25-fold and decreased Emax by up to 65%. The same mutations of the cognate C7.45 in rat 5-HT2C produced a smaller fourfold change in the affinity for serotonin and decreased agonist efficacy by up to 50%. Substitutions of S/C7.45 did not produce a significant change in the basal activity of either receptor. All mutants tested exhibited levels of receptor expression similar to the corresponding wildtype based on measurements of specific [3H]-mesulergine binding or flow cytometry analyses. Taken together, these results suggest that K4.45 and S/C7.45 play an important role in the conformational rearrangements leading to agonist-induced activation of 5-HT2 receptors.     

Evidence for a complex influence of nicotinic acetylcholine receptors on hippocampal serotonin release

The effects of nicotine on 5-hydroxytryptamine (5-HT) release from serotonergic nerve endings in rat dorsal hippocampal slices were studied. Nicotine (50-500 microM:) caused a concentration-dependent increase in 5-HT release. This effect was antagonised by mecamylamine (0.5 microM:), indicating an action at nicotinic receptors. Nicotine-evoked 5-HT release was not affected by tetrodotoxin (3 microM:), cadmium chloride (0.1 mM:), or the absence of Ca(2+) or Na(+) in the superfusion medium. Unexpectedly, higher concentrations of mecamylamine alone (1-50 microM:) increased 5-HT release. This suggested the presence of inhibitory input to 5-HT neurones and that these inhibitory neurones possess tonically active nicotinic receptors. The effect of mecamylamine (50 microM:) on 5-HT release was reduced by the muscarinic M(1) receptor agonist, McN-A-343 (100 microM:), but pirenzepine (0.005-1 microM:), which blocks M(1) receptors, alone increased 5-HT release. Hippocampal serotonergic neurones are known to possess both excitatory nicotinic receptors and inhibitory M(1) receptors. Although there may be several explanations for our results, one possible explanation is that nicotine stimulates 5-HT release by activating nicotinic heteroreceptors on 5-HT terminals. Mecamylamine (0.5 microM:) antagonises this effect, but higher concentrations increase 5-HT release indirectly by blocking the action of endogenous acetylcholine on nicotinic receptors situated on cholinergic neurones that provide muscarinic inhibitory input to 5-HT neurones.     

Pulmonary vasoconstriction by serotonin is inhibited by S-nitrosoglutathione

Nitric oxide (NO) functions as an endothelium-derived relaxing factor by activating guanylate cyclase to increase cGMP levels. However, NO and related species may also regulate vascular tone by cGMP-independent mechanisms. We hypothesized that naturally occurring NO donors could decrease the pulmonary vascular response to serotonin (5-HT) in the intact lung through chemical interactions with 5-HT(2) receptors. In isolated rabbit lung preparations and isolated pulmonary artery (PA) rings, 50-250 microM S-nitrosoglutathione (GSNO) inhibited the response to 0.01-10 microM 5-HT. The vasoconstrictor response to 5-HT was mediated by 5-HT(2) receptors in the lung, since it could be blocked completely by the selective inhibitor ketanserin (10 microM). GSNO inhibited the response to 5-HT by 77% in intact lung and 82% in PA rings. In PA rings, inhibition by GSNO could be reversed by treatment with the thiol reductant dithiothreitol (10 mM). 3-Morpholinosydnonimine (100-500 microM), which releases NO and O simultaneously, also blocked the response to 5-HT. Its chemical effects, however, were distinct from those of GSNO, because 5-HT-mediated vasoconstriction was not restored in isolated rings by dithiothreitol. In the intact lung, neither NO donor altered the vascular response to endothelin, which activates the same second-messenger vasoconstrictor system as 5-HT. These findings, which did not depend on guanylate cyclase, are consistent with chemical modification by NO of the 5-HT(2) G protein-coupled receptor system to inhibit vasoconstriction, possibly by S-nitrosylation of the receptor or a related protein. This study demonstrates that GSNO can regulate vascular tone in the intact lung by a reversible mechanism involving inhibition of the response to 5-HT.     

Serotoninergic control of somatostatin and gastrin release from the isolated rat stomach

The influence of exogenous serotonin on the secretion of gastric somatostatin and gastrin was investigated under in vitro conditions using an isolated, vascularly perfused rat stomach preparation. Serotonin stimulated gastrin release, maximal effects were observed at 10(-6) M which increased gastrin levels by 78%; on the contrary, somatostatin secretion was inhibited (maximal inhibition of 56% at 10(-6) M). Changes in hormone secretion in response to serotonin were reversed by combined blockade of 5-HT1 and 5-HT2 receptors by methysergide and blockade of 5-HT2 receptors by ketanserin (10(-5) and 10(-6) M, respectively), and of cholinoreceptors by atropine (10(-5) M). It is concluded that in rats in vitro serotonin inhibits release of gastric somatostatin and stimulates gastrin secretion via specific serotonin receptors but muscarinic cholinergic receptors are also involved.     

Effect of acute and chronic exercise on plasma amino acids and prolactin concentrations and on [3H]ketanserin binding to serotonin2A receptors on human platelets

The neurotransmitter serotonin (5-hydroxytryptamine, 5-HT) has been shown to modulate various physiological and psychological functions such as fatigue. Altered regulation of the serotonergic system has been suggested to play a role in response to exercise stress. In the present study, the influence was investigated of acute endurance exercise and short-term increase in the amount of training on the concentrations of the 5-HT precursor tryptophan (TRP), of prolactin (PRL) and of branched-chain amino acids (BCAA) in the blood, as well as on the binding of [3H]ketanserin to the serotonin-2A (5-HT2A) receptors on platelets. Nine healthy endurance-trained men were tested the day before (I) and after (II) a 9-day training programme. Samples of venous blood were drawn after an overnight fast and following 5 h of cycling. Fasted and post-exercise plasma concentrations of free TRP, BCAA and free TRP:BCAA ratio did not differ between I and II. A significant decrease of plasma BCAA (P < 0.01) and significant augmentations of plasma free TRP, free TRP:BCAA ratio and PRL (P < 0.01) were found post-exercise. The increase in plasma PRL was smaller in II compared with I. Acute endurance exercise reduced the density of platelet 5-HT2A receptor [3H]ketanserin binding sites at I and II (P < 0.05). The basal density of the binding sites and the affinity of [3H]ketanserin for these binding sites were unaffected by an increase in the amount of training. The present results support the hypothesis that acute endurance exercise may increase 5-HT availability. This was reflected in the periphery by increased concentration of the 5-HT precursor free TRP, by increased plasma PRL concentration, and by a reduction of 5-HT2A receptors on platelets. It remains to be resolved whether these alterations in the periphery occur in parallel with an increase in the availability of 5-HT in the brain.     

The activation of 5-HT receptors in prefrontal cortex enhances dopaminergic activity

Atypical antipsychotics show preferential 5-HT 2A versus dopamine (DA) D2 receptor affinity. At clinical doses, they fully occupy cortical 5-HT2 receptors, which suggests a strong relationship with their therapeutic action. Half of the pyramidal neurones in the medial prefrontal cortex (mPFC) express 5-HT 2A receptors. Also, neurones excited through 5-HT 2A receptors project to the ventral tegmental area (VTA). We therefore hypothesized that prefrontal 5-HT 2A receptors can modulate DA transmission through excitatory mPFC-VTA inputs. In this study we used single unit recordings to examine the responses of DA neurones to local (in the mPFC) and systemic administration of the 5-HT 2A/2C agonist 1-[2,5-dimethoxy-4-iodophenyl-2-aminopropane] (DOI). Likewise, using microdialysis, we examined DA release in the mPFC and VTA (single/dual probe) in response to prefrontal and systemic drug administration. The local (in the mPFC) and systemic administration of DOI increased the firing rate and burst firing of DA neurones and DA release in the VTA and mPFC. The increase in VTA DA release was mimicked by the electrical stimulation of the mPFC. The effects of DOI were reversed by M100907 and ritanserin. These results indicate that the activity of VTA DA neurones is under the excitatory control of 5-HT 2A receptors in the mPFC. These observations may help in the understanding of the therapeutic action of atypical antipsychotics.     

Functional characterization of the novel antipsychotic iloperidone at human D2, D3, alpha 2C, 5-HT6, and 5-HT1A receptors

Iloperidone has demonstrated an interesting monoamine receptor profile in radioligand binding studies, with nanomolar affinity for certain noradrenaline, dopamine, and serotonin receptors. In this study, the agonist/antagonist activity of iloperidone was determined in cell lines expressing recombinant human D(2A), D(3), alpha(2C), 5-HT(1A), or 5-HT(6) receptors. With the exception of 5-HT(6) receptors, these receptors are negatively coupled to cyclase. Thus, after stimulation with forskolin, the agonists dopamine (at D(2A) and D(3)), noradrenaline (at alpha(2C)), or 8-OH-DPAT (at 5-HT(1A)) induced a reduction in cAMP accumulation. Conversely, activation of the 5-HT(6) receptor by 5-HT led to an increase in cAMP accumulation. Iloperidone alone was devoid of significant agonist activity but inhibited the agonist response in all 5 cell lines in a surmountable and concentration-dependent fashion. Iloperidone was most potent at D(3) receptors (pK(B) 8.59 +/- 0.20; n = 6), followed by alpha(2C) (pK(B) 7.83 +/- 0.06; n = 15), 5-HT(1A) (pK(B) 7.69 +/- 0.18; n = 10), D(2A) (pK(B) 7.53 +/- 0.04; n = 11) and 5-HT(6) (pK(B) 7.11 +/- 0.08; n = 11) receptors.     

Activating Mutations of the Serotonin 5-HT2C Receptor

Abstract: Site-directed mutagenesis was performed to create a mutant serotonin 5-HT_2C receptor that would mimic the active conformation of the native receptor. Structural alteration of receptor conformation was achieved by changing amino acid no. 312 from serine to phenylalanine (S312F) or lysine (S312K). After expression in COS-7 cells, the binding affinity of 5-HT for [³H]-mesulergine-labeled 5-HT_2C receptors increased from 203 n M (native) to 76 n M for S312F and 6.6 n M for S312K mutant receptors. 5-HT potency for stimulation of phosphatidylinositol (PI) hydrolysis increased from 70 n M (native) to 28 n M for S312F and 2.7 n M for S312K mutant receptors. The mutant receptors were constitutively active, stimulating PI hydrolysis in the absence of agonist. S312F and S312K mutations resulted in twofold and five-fold increases, respectively, in basal levels of PI hydrolysis. Mianserin and mesulergine displayed inverse agonist activity by decreasing basal levels of PI hydrolysis stimulated by S312K mutant receptors. [³H]5-HT and [³H]-mesulergine labeled the same number of S312K mutant receptors and 5'-guanylylimidodiphosphate had no effect on [³H]5-HT binding. These results indicate that serine → lysine mutation at amino acid no. 312 produces an agonist high-affinity state of the 5-HT_2C receptor that spontaneously couples to G proteins and stimulates PI hydrolysis in the absence of agonist.     

Brain neurotransmitter receptor-binding characteristics in rats after oral administration of haloperidol, risperidone and olanzapine

The present study was conducted to characterize the binding of neurotransmitter receptors (dopamine D(2), serotonin 5-HT(2), histamine H(1), adrenaline alpha(1) and muscarine M(l) receptors) in the rat's brain after the oral administration of haloperidol, risperidone, and olanzapine. Haloperidol at 1 and 3 mg/kg displayed significant activity to bind the D(2) receptor (increase in the Kd value for [(3)H]raclopride binding) in the corpus striatum with little change in the activity toward the 5-HT(2) receptor (binding parameters for [(3)H]ketanserin). In contrast, risperidone (0.1-3 mg/kg) showed roughly 30 times more affinity for the 5-HT(2) receptor than D(2) receptor. Also, olanzapine (1-10 mg/kg) was most active toward the H(1) receptor in the cerebral cortex, corpus striatum, and hippocampus, was less active in binding 5-HT(2) and D(2) receptors, and showed the least affinity for alpha(1) and M(1) receptors. In conclusion, haloperidol and risperidone administered orally selectively bind D(2) and 5-HT(2) receptors, respectively, in the rat brain, while olanzapine binds H(1), 5-HT(2), and D(2) receptors more than alpha(1) and M(1) receptors.