Mediation of the cardiovascular response to spinal gamma-aminobutyric acid(B) receptor stimulation by adenosine A(1) receptors in anesthetized rats

Cardiovascular inhibitory effects induced by intrathecal (i.t.) administration of adenosine A(1) receptor agonist and its modulation by gamma-aminobutyric acid(B) (GABA(B)) receptor was suggested by our previous report. In this experiment, we examined the mediation of cardiovascular effects of GABA(B) receptor stimulation by adenosine A(1) and A(2) in the spinal cord. I.t. administration of GABA(B) receptor agonist, baclofen (30, 60 and 100 nmol) produced a dose dependent decrease of blood pressure and heart rate. Pretreatment with adenosine A(1) receptor antagonist, 8-cyclopentyl-1,3-dimethylxanthine (50 nmol), attenuated the depressor and bradycardiac effects of baclofen (100 nmol), but not with adenosine A(2) receptor antagonist, 3, 7-dimethyl-1-propargylxanthine (25 nmol). These results suggest that GABA(B) receptors in the spinal cord play an inhibitory role in the central cardiovascular regulation and that the depressor and bradycardiac actions are mediated by adenosine A(1) receptors.     

Involvement of nitric oxide production via kynurenic acid-sensitive glutamate receptors in DOPA-induced depressor responses in the nucleus tractus solitarii of anesthetized rats

We have proposed the hypothesis that L-3,4-dihydroxyphenylalanine (DOPA) plays a role of neurotransmitter of the primary baroreceptor afferents terminating in the nucleus tractus solitarii (NTS). In the present study, we tried to clarify whether glutamate receptors and/or nitric oxide (NO), important modulators for central cardiovascular regulation, are involved in the DOPA-induced cardiovascular responses in the nucleus. Male Wistar rats were anesthetized with urethane and artificially ventilated. Compounds or antisense oligos (17-mer) for neuronal NO synthase were microinjected into depressor sites of the unilateral nucleus. DOPA 30-300 pmol microinjected into the nucleus dose-dependently induced depressor and bradycardic responses. Prior injection of kynurenic acid (600 pmol) suppressed DOPA (300 pmol)-induced responses by approximately 80%. Prior injection of N(G)-monomethyl-L-arginine 100 nmol, a potent NO synthase inhibitor, reversibly attenuated by approximately 90% DOPA-induced responses, while the D-isomer 100 nmol produced no effect. Furthermore, prior injection of neuronal NO synthase antisense oligos (20 pmol) reversibly reduced by approximately 70% responses to DOPA. Sense or scrambled oligos produced no effect. A NO precursor L-arginine (30 nmol) induced depressor and bradycardic responses, but these responses were not affected by kynurenic acid. These results suggest important roles for glutamate receptors and NO in DOPA induced-depressor and bradycardic responses in the NTS.     

Cardiovascular responses to microinjection of nociceptin and endomorphin-1 into the nucleus tractus solitarii in conscious rats

Increasing evidence suggests an active participation of nociceptinergic transmission in the central control of cardiovascular activity and reflex. In this study, the role of the classic opioid mu receptor and the nociceptin/orphanin FQ receptor, a novel opioid receptor, in the nucleus tractus solitarii (NTS) in the regulation of cardiovascular activity was investigated and compared in chronically cannulated and freely moving conscious rats. Microinjections of nociceptin, an endogenous ligand for the nociceptin receptor, into the relatively rostral NTS produced dose-related (0.04, 0.2, and 1 nmol) increases in blood pressure and heart rate. Intra-NTS injection of the selective nociceptin receptor antagonist [Nphe(1)]Nociceptin(1-13)NH(2) (NOR-AN) at 1 nmol blocked the increases in blood pressure and heart rate induced by nociceptin. In contrast, pretreatment with the nonselective opioid receptor antagonist naloxone (5 nmol) had no effects on the cardiovascular responses to nociceptin. Like nociceptin, microinjection of endomorphin-1 (EM-1), an endogenous ligand for the opioid mu receptor, into the rostral NTS increased blood pressure and heart rate in a dose-dependent manner (0.04, 0.2, and 1 nmol). Pretreatment with naloxone (5 nmol), but not NOR-AN, blocked cardiovascular responses elicited by EM-1. Neither NOR-AN nor naloxone alone had significant effects on the baseline blood pressure and heart rate. Injection of excitatory amino acid l-glutamate (1 nmol) into the same sites caused the typical depressor and bradycardic responses. In the caudal NTS areas, nociceptin and EM-1 seemed to induce opposite responses: hypotension and bradycardia. These results suggest that the novel nociceptin receptors and traditional opioid receptors in the NTS may be independently involved in the regulation of cardiovascular activity.     

Nitric oxide in the gracile nucleus mediates depressor response to acupuncture (ST36)

The purpose of these studies was to determine the role of gracile nucleus and the effects of l-arginine-derived nitric oxide (NO) synthesis in the nucleus on the cardiovascular responses to electroacupuncture (EA) stimulation of "Zusanli" (ST36). Arterial blood pressure and heart rate were monitored during EA stimulation of ST36 following microinjections of agents into gracile nucleus. EA ST36 produced depressor and bradycardiac responses in anesthetized Sprague-Dawley rats. The cardiovascular responses to EA ST36 were blocked by bilateral microinjection of lidocaine into gracile nucleus. Microinjection of L-arginine into gracile nucleus facilitated the hypotensive and bradycardiac responses to EA ST36. The cardiovascular responses to EA ST36 were attenuated by bilateral microinjection of neuronal NO synthase (nNOS) antisense oligos into gracile nucleus. Microinjection of nNOS sense oligos into gracile nucleus did not alter the cardiovascular response to EA ST36. The results demonstrate that a blockade of neuronal conduction in the gracile nucleus inhibits the cardiovascular responses to EA ST36. The hypotensive and bradycardiac responses to EA ST36 are modified by influences of L-arginine-derived NO synthesis in the gracile nucleus. We conclude that NO plays an important role in mediating the cardiovascular responses to EA ST36 through gracile nucleus.     

Seizure suppression by adenosine A(2A) receptor activation in a rat model of audiogenic brainstem epilepsy

Adenosine is known to suppress seizure activity mainly by activation of adenosine A(1) receptors. However, little is known about the potential involvement of other types of adenosine receptors in seizure suppression. It was now tested whether activation of adenosine A(2A) receptors would be effective in the suppression of generalized brainstem seizures. Genetically epilepsy-prone rats were intraperitoneally injected with increasing doses of the A(2A) receptor agonist, 5'-(N-cyclopropyl)-carboxamido-adenosine (CPCA), and, for comparison, with the A(1) receptor agonist, 2-chloro-N(6)-cyclopentyladenosine (CCPA). Both CPCA and CCPA were effective in suppressing generalized brainstem seizures with minimal effective concentrations of 2.5 and 1.5 mg/kg, respectively. Seizure suppression was maintained when CPCA was co-injected with the peripherally acting adenosine receptor antagonist 8-(p-sulphophenyl)theophylline, suggesting that central activation of A(2A) receptors effectively contributes to seizure suppression.     

The ventrolateral periaqueductal gray is involved in the cardiovascular response evoked by l-glutamate microinjection into the lateral hypothalamus of anesthetized rats

Microinjection of l-glutamate (l-glu: 1, 3, 10 and 30nmol/100nL) into the lateral hypothalamus (LH) caused dose-related depressor and bradycardiac responses. The cardiovascular response to l-glu stimulation of the LH was blocked by pretreatment of the ventrolateral portion of the periaqueductal gray matter (vlPAG) with CoCl2 (1mM/100nL), indicating the existence of a synaptic relay of the hypotensive pathway in that area. Furthermore, the response to l-glu was blocked by pretreatment of the vlPAG with 2nmol/100nL of the selective NMDA-receptor antagonist LY235959 and was not affected by pretreatment with 2nmol/100nL of the selective non-NMDA-receptor antagonist NBQX, suggesting a mediation of the hypotensive response by NMDA receptors in the vlPAG. In conclusion, our results indicate that the hypotensive pathway activated by microinjection of l-glu into the LH involves a NMDA synaptic relay in the vlPAG.     

Nitrergic stimulation of the locus coeruleus modulates blood pressure and heart rate in the anaesthetized rat.

To investigate whether nitric oxide is involved in the cardiovascular responses mediated via the locus coeruleus, the effects of microinjections of L-arginine and L-glutamate into the locus coeruleus on blood pressure and heart rate were investigated in sodium pentobarbitone-anaesthetized rats. Unilateral microinjection of L-arginine (25, 50 nmol) elicited dose-related depressor (-17 +/- 4, -25 +/- 4 mmHg) and bradycardic (13 +/- 3, 24 +/- 6 b.p.m.) effects. Furthermore, these effects were attenuated by prior local microinjection of N(G)-nitro-L-arginine (40 nmol). Peripheral muscarinic receptor blockade with atropine methyl nitrate (1 mg/kg, i.v.) attenuated the bradycardic but not the depressor responses to L-arginine. L-Glutamate (2 nmol) microinjections also mediated depressor (-27 +/- 6 mmHg) and bradycardic (53 +/- 23 b.p.m.) effects that were attenuated by microinjections of dizocilpine maleate (1 nmol) into the locus coeruleus. In addition, pretreatment with N(G)-nitro-L-arginine (40 nmol) also significantly attenuated the depressor response elicited by L-glutamate. These results suggest that nitrergic and glutamatergic pathways are operative within the locus coeruleus to modulate cardiovascular function, and also that a functional interaction may exist between the nitrergic and glutamatergic systems within the rat locus coeruleus.     

Nitric oxide and GABA mediate bi-directional cardiovascular effects of orexin in the nucleus tractus solitarii of rats

The present study investigated the cardiovascular effects of orexin (OX)-A and OX-B in the nucleus tractus solitarii (NTS) and delineated the engagement of nitric oxide (NO) and GABA in OX-induced cardiovascular responses. In adult male Sprague-Dawley rats maintained under propofol anesthesia, microinjection bilaterally into the NTS of OX-A or OX-B evoked bi-directional cardiovascular effects in a dose-dependent manner. At a lower dose (5 pmol), OX-A or OX-B decreased systemic arterial pressure (SAP), heart rate (HR), and power density of the vasomotor components of SAP signals, our experimental index for sympathetic neurogenic vasomotor tone. At higher doses (>20 pmol), these two compounds elicited cardiovascular excitatory responses. These bi-directional cardiovascular effects of OX were abolished by co-injection of an OX(1) receptor antagonist, 1-(2-methylbenzoxazol-6-yl)-3-[1,5]naphthyridin-4-yl-urea hydrochloride (SB-334867, 0.75 nmol) or the OX(2) receptor antiserum (1:20). In addition, the vasodepressor effects of low dose (5 pmol) OX-A or OX-B in the NTS were attenuated by a nitric oxide synthase (NOS) inhibitor, N(G)-nitro-l-arginine methyl ester (l-NAME, 5 nmol), a neuronal nitric oxide synthase (nNOS) inhibitor, 7-nitroindazole (2.5 pmol) or the soluble guanylate cyclase (sGC) inhibitor, 1H-[1,2,4]oxadiazole[4,3-alpha]quinoxalin-1-one (250 pmol). The vasopressor effects of high dose (200 pmol) OX were reversed by co-administration with GABA(A) or GABA(B) receptor antagonist, bicuculline methiodine (10 pmol) or 2-hydroxy saclofen (100 pmol), or l-NAME (5 nmol). Our results indicate that OX-A or OX-B elicited bi-directional cardiovascular effects via OX receptor-dependent mechanisms. The vasodepressor effects of OX were induced by the nNOS-derived NO and activation of sGC-associated signaling pathway, whereas the vasopressor effects were mediated by interaction with GABAergic or nitrergic neurotransmission in the NTS.     

Neuronal Nitric Oxide Synthase Activation Is Involved in Insulin-Mediated Cardiovascular Effects in the Nucleus Tractus Solitarii of Rats

Neuronal nitric oxide synthases (nNOS) is distributed throughout the central nervous system (CNS) and has been proposed to modulate neuronal activity in the nucleus tractus solitarii (NTS). Here, we investigated whether the activation of nNOS is involved in insulin-induced cardiovascular responses in the NTS. Insulin (100 IU/ml) was unilaterally microinjected into the NTS, and the cardiovascular effects were evaluated before and after microinjection of the nNOS inhibitors 7-nitroindazole (7-NI) (5 pmol) and N(5)-(1-imino-3-butenyl)-l-ornithine (vinyl-L-NIO) (600 pmol). Western blot and immunohistochemical analyses were performed to determine nNOS phosphorylation levels after insulin or phosphoinositide 3-kinase (PI3K) inhibitor LY294002 microinjection into the NTS. Unilateral microinjection of insulin into the NTS produced prominent depressor and bradycardic effects in WKY rats. Pretreatment with the nNOS inhibitors 7-NI and Vinyl-L-NIO attenuated the cardiovascular response evoked by insulin in Wistar-Kyoto (WKY) rats. Moreover, Western blot analysis showed a significant increase in nNOS (16.5±0.4-fold; P<0.05; n=4) phosphorylation after insulin injection, whereas the PI3K inhibitor LY294002 abolished the insulin-induced effects. In situ nNOS phosphorylation was found to be increased in the NTS after insulin injection. Furthermore, co-immunoprecipitation assay showed Akt and nNOS can bind to each other as detected by phospho-Akt^S473 and phospho-nNOS^S1416 antibodies. In vitro kinase assay showed insulin activated Akt can directly phosphorylate nNOS^S1416. These results demonstrated that nNOS may couple with the activation of the insulin receptor, via the liberation of NO, in order to participate in central cardiovascular regulation of WKY rats.     

The effects of A1 and A2A adenosine receptor agonists on kainic acid excitotoxicity in the guinea pig cochlea

The present study aimed to clarify the protective effect of adenosine receptors against the excitotoxicity of cochlear afferent dendrites. The effects of 2-chloro-N6-cyclopentyladenosine (CCPA), an A1 adenosine receptor agonist, and 5′-N-cyclopropyl-carboxamidoadenosine (CPCA), an A2A adenosine receptor agonist, on cochlear excitotoxicity induced by kainic acid (KA) were examined using guinea pigs. KA was applied to the round window membrane at a concentration of 10 mM for 30 min. CCPA or CPCA was given at the onset of KA application. KA morphologically induced the swelling of cochlear afferent dendrites and significantly elevated the threshold of the compound action potential (CAP) of the cochlea. CCPA inhibited the KA-induced CAP threshold shift and swelling of the cochlear afferent dendrites. However, CPCA did not affect cochlear excitotoxicity induced by KA. The results suggest that adenosine A1 receptor activation could prevent the excitotoxicity of cochlear afferent dendrites.     

Involvement of the purinergic system in central cardiovascular modulation at the level of the nucleus ambiguus of anaesthetized rats

Anatomical studies have demonstrated the existence of purinergic P2 receptors in the nucleus ambiguus (NA), a site containing cardiac vagal motoneurons. However, very little is known about the functional role of these receptors in central cardiac vagal regulation. The aims of our study were to evaluate the following: (1) the blood pressure and heart rate responses following purinoceptor activation within the NA; (2) the role of purinoceptors and excitatory amino acid (EAA) receptors in mediating the cardiovascular responses evoked by ATP and L-glutamate stimulation of NA; and (3) the role of NA purinoceptors in mediating the cardiovascular responses of the Bezold-Jarisch reflex. In anaesthetized rats, microinjection of L-glutamate (5.0 nmol/50 nl) into the NA induced a marked and immediate onset bradycardia with minimal change in arterial pressure. Microinjection of ATP into the NA induced a dose-dependent (0.31-6.0 nmol/50 nl) bradycardia and pressor responses. It is noteworthy that the bradycardia occurred either before or simultaneously with a pressor response (when present), indicating that it was not a baroreceptor reflex mediated response due to the rise in arterial pressure. The pressor response was prevented by α(1)-adrenergic blockade with prazosin, whereas muscarinic blockade with methyl-atropine abolished the evoked bradycardia. Ipsilateral microinjection of PPADS (a P2 receptor antagonist; 500 pmol/100 nl) into the NA significantly attenuated the ATP-induced bradycardia but spared the pressor response. In contrast, PPADS in the NA had no effect on the L-glutamate-evoked bradycardic response. Ipsilateral injection of kynurenic acid (a non-selective EAA receptor antagonist; 10 nmol/50 nl) into the NA totally blocked the bradycardia induced by l-glutamate and partly attenuated the ATP induced bradycardia. Finally, both the depressor and the bradycardic responses of the Bezold-Jarisch reflex were attenuated significantly (P < 0.01 and P < 0.05, respectively) following bilateral microinjection of PPADS into the NA. These results identify ATP and purinergic P2 receptors within the ventrolateral medulla as excitatory to cardiovagal neurons. Additionally, our data show that P2 receptors within the ventrolateral medulla are integral to the cardiovascular responses of the Bezold-Jarisch reflex.     

侧脑室注射肾上腺髓质素激活大鼠脑内儿茶酚胺神经元

利用Fos蛋白和酪氨酸羟化酶(TH)的双重免疫组化方法,观察侧脑室注射肾上腺髓质素(ADM)对大鼠心血管相关核团中儿茶酚胺神经元c-fos表达的影响,以探讨ADM的中枢效应是否通过激活脑内儿茶酚胺能神经元而诱发。侧脑室注射ADM引起最后区(AP)、孤束核(NTS)、巨细胞旁外侧核(PGL)和蓝斑核(LC)内Fos-TH双标神经元明显增加;降钙素基因相关肽受体拮抗剂CGRP8-37(30nmol/kg)可明显减弱ADM的效应。结果表明,ADM可兴奋脑内多个心血管相关核团的神经元,其中枢效应通过激活儿茶酚胺能神经元而诱发,降钙素基因相关肽受体可能介导这一效应。     

The medial amygdaloid nucleus modulates cardiovascular responses to acute restraint in rats

The medial amygdaloid nucleus (MeA) modulates several physiological and behavioral processes and among them, the cardiovascular correlates of behavioral responses to stressful stimuli. Acute restraint evokes cardiovascular responses, which are characterized by both elevated blood pressure (BP) and intense heart rate (HR) increase. We presently report effects of MeA pharmacological manipulations on BP and HR responses evoked by acute restraint in rats. Bilateral microinjection of 100 nL of the unspecific synaptic blocker CoCl₂ (1 mM) into the MeA increased HR response to acute restraint, without significant effect on the BP response. This result indicates an inhibitory influence of MeA on restraint-evoked HR changes. Injections of the non-selective muscarinic receptor antagonist atropine (3 nmol); the inhibitor of choline uptake hemicholinium (2 nmol) or the selective M₁-receptor antagonist pirenzepine (6 nmol) caused effects that were similar to those caused by cobalt. These results suggest that local cholinergic neurotransmission and M₁-receptors mediate the MeA inhibitory influence on restraint-related HR responses. Pretreatment with the M3 receptor antagonist 4-DAMP (4-Diphenylacetoxy-N-methylpiperidine methiodide-2 nmol) did not affect restraint-related cardiovascular responses, reinforcing the idea that M₁-receptors mediate MeA-related inhibitory influence on restraint-evoked HR increase.     

Evidence that neurons of the central amygdaloid nucleus directly project to the site concerned with circulatory and respiratory regulation in the ventrolateral nucleus of the cat: a WGA-HRP study.

Projections from the central amygdaloid nucleus (ACE) to the ventrolateral nucleus (VLN) of the medulla oblongata were studied using the wheat germ agglutinin-horseradish peroxidase (WGA-HRP) method. WGA-HRP was injected into the rostral VLN, where a pressor response associated with bradycardiac and apneustic responses was elicited by microinjection of glutamate, and into the caudal VLN, where a depressor response associated with bradycardiac and apneustic responses was elicited. In both experiments, HRP-labeled cells were found in the ACE. These results suggest that the ACE may send direct projections to the VLN and may play a role in cardiovascular and/or respiratory control mechanisms.     

Evidence for gamma-aminobutyric acid receptor-mediated modulation of the aortic baroreceptor reflex in the nucleus tractus solitarii of the rat

Microinjections of the gamma-aminobutyric acid (GABA) receptor antagonist bicuculline into the medial area of the nucleus tractus solitarii of the rat enhanced the depressor and bradycardiac responses to aortic nerve stimulation whereas the glycine receptor antagonist strychnine did not affect them. The GABA receptor agonist muscimol and the GABA uptake inhibitor nipecotic acid reduced the responses to aortic nerve stimulation. These results provide evidence suggesting GABA receptor-mediated modulation of the aortic baroreceptor reflex in the nucleus tractus solitarii of the rat.     

Nitric oxide mediates noradrenaline-induced hypothermic responses and opposes prostaglandin E2-induced fever in the rostromedial preoptic area

Noradrenaline (NA) microinjected into the rostromedial preoptic area (POA) elicits heat loss responses and opposes prostaglandin E₂-induced fever. Here, I tested the hypothesis that local synthesis and release of nitric oxide (NO) mediates the NA-induced effects. The unilateral microinjection of the NO donor sodium nitroprusside (SNP, 8.4 nmol), but not that of saline solution, into the NA-sensitive site elicited an increase in tail skin temperature and decreases in the whole-body O₂ consumption rate, heart rate, and colonic temperature simultaneously in urethane–chloralose-anesthetized rats. Pretreatment with SNP greatly attenuated the thermogenic, tachycardic, and hyperthermic effects of prostaglandin E₂ (140 fmol) microinjected into the same site. Furthermore, the NA-induced hypothermic responses were largely blocked by a prior microinjection of an NO synthase inhibitor N^G-monomethyl-l-arginine (l-NMMA, 5 nmol), but not by that of its inactive enantiomer, N^G-monomethyl-d-arginine (d-NMMA, 5 nmol), at the same site. These results suggest that the hypothermic and antipyretic effects of NA are mediated by NO in the rostromedial POA.     

Adenosine postsynaptically modulates supraoptic neuronal excitability

Effects of adenosine on the excitability of supraoptic nucleus neurons were investigated in whole cell patch-clamp experiments conducted in horizontal slices of rat hypothalamus. Adenosine (10-100 muM) inhibited all neurons tested by reducing or abolishing spontaneous or evoked discharge. Large hyperpolarizations were seen, averaging -6.08 +/- 0.83 mV below resting membrane potential, and action potential durations were significantly reduced by 134 +/- 41 mus in the presence of 100 muM adenosine. The A(1) receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine (DPCPX, 1 muM) blocked these effects, whereas the A(1) agonists N(6)-cyclopentyladenosine (CPA) and N(6)-cyclohexyladenosine (CHA) mimicked the actions of adenosine. A(2) receptor contributions to excitability were assessed by application of an A(2) agonist, carboxamidoadenosine (CPCA). This resulted in membrane depolarizations (3.56 +/- 0.65 mV) and maintenance of firing. The presence of endogenous adenosine in the slice was revealed by both the application of the adenosine uptake inhibitor dilazep (1-100 muM), which resulted in a strong inhibition of firing activity, and the application of DPCPX, which induced firing in cells silenced by negative current injection. We tested for postsynaptic actions of adenosine by blocking G protein activation via GDP-beta-S infusion into recorded neurons. Under these conditions, the adenosinergic inhibition of firing and reduction of spike duration were blocked, suggesting the effects were mediated by postsynaptic adenosine receptors. That the effects on excitability could be due to direct activation of adenosine A(1) receptors on supraoptic neurons was further explored immunocytochemically via the co-labeling of magnocellular neurons with polyclonal antibodies raised against the A(1) receptors. It is concluded that adenosine, acting at postsynaptic A(1) receptors, exhibits a powerful inhibitory influence on supraoptic magnocellular activity and is an important endogenous regulator of magnocellular neuroendocrine function.     

Risperidone attenuates the increase of extracellular nitric oxide and glutamate levels in serotonin syndrome animal models

Serotonin (5-hydroxytryptamine; 5-HT) syndrome is a potentially life-threatening neurotoxic condition provoked by pharmacologically induced excess serotonergic activity. Several studies report that nitric oxide (NO) and glutamate play a role in psychostimulant-induced hyperthermia related to neurotoxicity. In the present study, the involvement of NO and glutamate, as well as the effect of risperidone, a potent 5-HT_2A and D₂ (and a less potent D₁) receptor antagonist, were investigated in animal models of 5-HT syndrome. Two 5-HT syndrome animal models were utilized. The first model was induced by administration of tranylcypromine, a nonselective monoamine oxidase (MAO) inhibitor, and fluoxetine, a selective 5-HT reuptake inhibitor. The second model was induced by the administration of clorgyline, an MAO-A inhibitor, and 5-hydroxy-l-tryptophan, a precursor of 5-HT. Changes in the level of NO metabolites and glutamate in the anterior hypothalamus were measured using microdialysis. In both models, NO metabolite levels significantly increased, and this increase was significantly attenuated by risperidone pretreatment. Extracellular levels of glutamate were increased only in the tranylcypromine and fluoxetine model, and this increase was significantly attenuated by risperidone pretreatment. These results indicate that NO and glutamate may be involved in the development of 5-HT syndrome and that risperidone may be effective against neurotransmitter abnormalities in 5-HT syndrome.     

Regulation of adenosine receptor subtypes during cultivation of human monocytes: role of receptors in preventing lipopolysaccharide-triggered respiratory burst

Adenosine is a potent anti-inflammatory agent that modulates the function of cells involved in the inflammatory response. Here we show that it inhibits lipopolysaccharide (LPS)-induced formation of reactive oxygen intermediates (ROI) in both freshly isolated and cultured human monocytes. Blocking of adenosine uptake and inactivation of the adenosine-degrading enzyme adenosine deaminase enhanced the inhibitory action of adenosine, indicating that both pathways regulate the extracellular adenosine concentration. Adenosine-mediated inhibition could be reversed by XAC (xanthine amine congener), an antagonist of the adenosine receptor A(2A), and MRS 1220 [N-9-chloro-2-(2-furanyl)[1, 2, 4]-triazolo[1,5-c]quinazolin-5-benzeneacetamide], an A(3) receptor antagonist, in both cell populations, while DPCPX (1,3-dipropyl-8-cyclopentylxanthine), an A(1) receptor antagonist, had no effect. Similar to what was seen with adenosine, CGS 21680, an A(2A) and A(3) receptor agonist, and IB-MECA, a nonselective A(1) and A(3) receptor agonist, dose dependently prevented ROI formation, indicating the involvement of A(3) and probably also A(2A) in the suppressive effect of adenosine. Pretreatment of monocytes with adenosine did not lead to changes in the LPS-induced increase in intracellular calcium levels ([Ca(2+)](i)). Thus, participation of [Ca(2+)](i) in the action of adenosine seems unlikely. The adenosine-mediated suppression of ROI production was found to be more pronounced when monocytes were cultured for 18 h, a time point at which changes in the mRNA expression of adenosine receptors were observed. Most prominent was the increase in the A(2A) receptor mRNA. These data demonstrate that cultivation of monocytes is accompanied by changes in the inhibitory action of adenosine mediated by A(3) and probably also the A(2A) receptor and that regulation of adenosine receptors is an integral part of the monocyte differentiation program.     

Nitric oxide inhibits the firing activity of hypothalamic paraventricular neurons that innervate the medulla oblongata: role of GABA

Nitric oxide (NO) has been shown to modulate autonomic function by acting both peripherally and centrally. A growing body of evidence indicates that the paraventricular nucleus of the hypothalamus (PVN), an important site for autonomic and endocrine homeostasis, constitutes an important locus mediating central NO actions. However, the cellular targets and mechanisms mediating NO actions within the PVN are not completely understood. Here, we examined whether NO influences the firing activity of identified PVN neurons that innervate two functionally different autonomic centers, the dorsal vagal complex (DVC) and the rostral ventrolateral medulla (RVLM). Perforated patch-clamp recordings were performed in hypothalamic slices containing retrogradely labeled PVN neurons innervating the DVC or the RVLM. Application of the NO donors dyethylamine- or 1-propanamine, 3-(2-hydroxy-2-nitroso-1-propylhydrazino) NONOate inhibited the firing activity of both DVC- and RVLM-projecting PVN neurons. Furthermore, application of 2-(4-carboxypheny)-4,4,5,5,-tetramethilimidazoline-1-oxyl-3-oxide (carboxy-PTIO), or the relatively selective neuronal nitric oxide synthase (nNOS) inhibitor 7-nitroindazole alone, increased their basal firing activity, suggesting the presence of an endogenous NO inhibitory tone. GABAergic synaptic activity in PVN neurons was potentiated by NO donors, an action that involved a presynaptic mechanism. Furthermore, the NO-mediated inhibition of firing activity was blocked by the GABA(A) receptor antagonist bicuculline, suggesting that NO-inhibitory actions involved potentiation of local GABAergic synaptic activity. Immunohistochemical studies showed that approximately 25% of DVC- and RVLM-projecting PVN neurons express nNOS, suggesting that a proportion of these medullary-projecting PVN neurons contribute to the cellular source of NO within the PVN.In summary, NO has been identified as an important molecule controlling autonomic function under physiological and pathological conditions. Here, we provide information on the cellular mechanisms mediating central NO actions. Our results demonstrate for the first time that NO modulates the activity of identified populations of PVN neurons that innervate the medulla oblongata, an action that is likely mediated by enhancing synaptic GABAergic function. This work suggests that NO-GABA interaction in PVN neurons that innervate the medulla constitutes an efficient cellular mechanism mediating NO central regulation of autonomic function.