Activation of nucleus tractus solitarius 5-HT2A but not other 5-HT2 receptor subtypes inhibits the sympathetic activity in rats.

Our first aim was to elucidate the mechanisms underlying the hypotensive response elicited by 5-HT(2) receptor activation in the nucleus tractus solitarius (NTS). In pentobarbitone-anaesthetized rats, intra-NTS administration of 2,5-dimethoxy-4-iodoamphetamine (DOI), a wide spectrum 5-HT(2) receptor agonist, but not an antagonist of selective 5-HT(2B) and 5-HT(2C) receptors, produced a decrease in blood pressure and heart rate. The maximal cardiovascular changes obtained by DOI (0.5 pmol) could be almost completely abolished by prior intra-NTS microinjection (10 pmol) of MDL-100907, a selective 5-HT(2A) receptor antagonist, but not by 5-HT(2B) or 5-HT(2C) receptor antagonists. In addition, using extracellular recordings we found that the large majority of identified cardiovascular rostroventrolateral medulla (RVLM) neurons were almost totally inhibited by NTS 5-HT(2A) receptor stimulation. We then investigated whether intra-NTS administration of a subthreshold dose (0.05 pmol) of DOI, known to facilitate the cardiovagal component of the baroreflex, could also modulate the sympathoinhibitory component of this reflex. These experiments showed that neither the decrease in the activity of the cardiovascular RVLM neurons and lumbar sympathetic nerve activities produced by aortic occlusion (gain of the baroreflex), nor the hypotensive response elicited by aortic nerve stimulation, were potentiated by the microinjection of DOI under such conditions. These data show that activation of 5-HT(2A), but not 5-HT(2B) or 5-HT(2C), receptors, located on NTS neurons, elicits depressor and bradycardic responses, and that this 5-HT(2A)-mediated hypotension is produced via the inhibition of RVLM cardiovascular neurons. In addition, NTS 5-HT(2A) receptor activation facilitates the cardiac but not the sympathetic baroreflex response.

Cardiac baroreflex facilitation evoked by hypothalamus and prefrontal cortex stimulation: role of the nucleus tractus solitarius 5-HT2A receptors.

We previously showed that serotonin (5-HT2) receptor activation in the nucleus of the tractus solitarius (NTS) produced hypotension, bradycardia, and facilitation of the baroreflex bradycardia. Activation of the preoptic area (POA) of the hypothalamus, which is involved in shock-evoked passive behaviors, induces similar modifications. In addition, previous studies showed that blockade of the infralimbic (IL) part of the medial prefrontal cortex, which sends projections to POA, produced an inhibitory influence on the baroreflex cardiac response. Thus, to assess the possible implication of NTS 5-HT2 receptors in passive cardiovascular responses, we analyzed in anesthetized rats the effects of NTS inhibition and NTS 5-HT2 receptor blockade on the cardiovascular modifications induced by chemical (0.3 M D,L-homocysteic acid) and electrical (50 Hz, 150-200 microA) stimulation of IL or POA. Intra-NTS microinjections of muscimol, a GABAA receptor agonist, prevented the decreases in blood pressure and heart rate normally evoked by IL or POA activation. In addition, we found that intra-NTS microinjection of R(+)-alpha-(2,3-dimethoxyphenyl)-1-[2-(4-fluorophenylethyl)]-4-piperidine-methanol, a specific 5-HT2A receptor antagonist, did not affect the decreases in cardiovascular baseline parameters induced by IL or POA stimulation but prevented the facilitation of the aortic baroreflex bradycardia normally observed during IL (+65 and +60%) or POA (+70 and +69%) electrical and chemical stimulation, respectively. These results show that NTS 5-HT2A receptors play a key role in the enhancement of the cardiac response of the baroreflex but not in the changes in basal heart rate and blood pressure induced by IL or POA stimulation.

Functional interaction between nucleus tractus solitarius NK1 and 5-HT3 receptors in the inhibition of baroreflex in rats.

OBJECTIVE: Previous data showed that in the nucleus tractus solitarius (NTS), 5-HT(3) receptors are critically involved in the inhibition of cardiac baroreceptor reflex response occurring during the defense reaction. Since stimulation of NTS NK(1) receptors has been found to inhibit the baroreflex bradycardia, we examined in this study whether this reflex response is inhibited during the defense reaction via an interaction between NK(1) and 5-HT(3) receptors. METHODS: For this purpose, we analyzed in urethane-anaesthetized rats the effects of intra-NTS GR205171, a selective NK(1) receptor antagonist, on the baroreflex bradycardia inhibition observed either during the defense reaction triggered by electrical stimulation of the dorsal periaqueductal grey matter (dPAG) or after NTS 5-HT(3) receptor activation. RESULTS: Intra-NTS GR205171, reversed, in dose-dependent manner, the inhibitory effect of dPAG stimulation on baroreflex bradycardia. This reversion was of 49% when both sinus carotid and aortic baroreceptors were stimulated by phenylephrine, and of 84% when aortic depressor nerve was stimulated. Similarly, intra-NTS GR205171 reversed partially or almost totally the inhibitory effect of local microinjections of phenylbiguanide, a 5-HT(3) receptor agonist, on baroreflex bradycardia induced either by phenylephrine administration or aortic nerve stimulation, respectively. CONCLUSION: These results strongly suggest that NK(1) receptors contribute downstream to the 5-HT(3) receptor-mediated inhibition of the aortic but not carotid cardiac baroreflex response occurring during the defense reaction, therefore implying that baroreceptor afferent inputs may be differentially modulated depending on their origin. This differentiation may be useful for a better understanding of baroreflex dysfunction in disease-induced conditions.

The Key Role of Medullary 5-HT3 Receptors in the Serotonin-Mediated Neural Control of Cardiovascular Function.

The baroreceptor reflex plays a crucial role in the homeostatic control of cardiovascular parameters. In the central nervous system, the nucleus of the tractus solitarius (NTS) is critically involved in cardiovascular reflex control because it is both the first site of termination of glutamatergic baroreceptor afferent fibres and an important integrative area for the sensory afferent signals reaching the brainstem. In addition to glutamate, the NTS contains numerous neurotransmitters that could participate in the modulations of the baroreceptor reflex sensitivity which occur under various physiological conditions. In particular, a large body of evidence indicates that serotonin plays a modulatory role in the central control of blood pressure, especially at the level of the NTS, which is innervated by both central and peripheral serotonergic fibres. Indeed, serotonin exerts multiple cardiovascular influences through the activation of several receptors in the NTS. Actually, the NTS is the central area endowed with the highest density of serotonin3 (5-HT3) receptors whose stimulation triggers all the adaptive cardiovascular changes normally associated with behavioural responses to various stressful conditions. In this review, we first assess the current knowledge about the mechanisms underlying the cardiovascular effects of the specific activation of serotonergic receptors in the NTS. Secondly, we describe evidence that, in the NTS, 5-HT3 receptors play a key role in one of the crucial homeostatic responses that characterise the defence reaction: the inhibitory modulation of the parasympathetic cardiac component of the baroreceptor reflex. The possible functional interactions of 5-HT3 receptors with GABAA, NK1 and NMDA receptors within the NTS are also discussed.

Role of nucleus tractus solitarius 5-HT3 receptors in the defense reaction-induced inhibition of the aortic baroreflex in rats.

Different stressful conditions elicit a typical behavior called the defense reaction. Our aim was to determine whether 5-HT3 receptors in the nucleus tractus solitarius (NTS) are involved in 1) the inhibition of the baroreflex bradycardia and 2) the rise in blood pressure, which are known to occur during the defense reaction. In urethane-anesthetized rats, the defense reaction was elicited by electrical stimulation of the dorsomedial nucleus of the hypothalamus (DMH) or the dorsal part of the periaqueductal gray (dPAG). Direct electrical stimulation of the aortic depressor nerve was used to trigger the typical baroreflex responses. Aortic stimulation at high (100-150 microA) and low (50-90 microA) intensity produced a decrease in heart rate of -39 to -44% (relative to baseline, Group 1 responses, n = 113) and -19 to -24% (Group 2 responses, n = 43), respectively. In spontaneously breathing rats, Group 1 and Group 2 bradycardiac responses were inhibited during DMH (-75 +/- 4% and -96 +/- 4%, n = 38 and n = 11, respectively), as well as dPAG (-81 +/- 3% and -95 +/- 4%, n = 36 and n = 10, respectively) stimulation. The aortic baroreflex bradycardia was hardly affected by DMH or dPAG stimulation when bicuculline (5 pmol), a specific GABAA receptor antagonist, had previously been microinjected into the NTS. Likewise, NTS microinjections of granisetron, a specific 5-HT3 receptor antagonist, prevented, in a dose-dependent manner, the baroreflex bradycardia inhibition. In addition, intra-NTS granisetron did not affect the rise in blood pressure induced by either site stimulation. These data show that 5-HT3 receptors in the NTS are involved in the GABAergic inhibition of the aortic baroreflex bradycardia, but not in the rise in blood pressure, occurring during the defense reaction elicited by DMH or dPAG stimulation.

Serotonin(3) receptor stimulation in the nucleus tractus solitarii activates non-catecholaminergic neurons in the rat ventrolateral medulla.

The present study was performed to determine whether or not the increased arterial pressure triggered by 5-HT(3) receptor stimulation in the nucleus tractus solitarii and underlain by a sympathoexcitation is associated with the activation of ventromedullary cells known to be involved in vascular regulation, i.e. the C1 and A1 catecholaminergic cells. For this purpose, double immunohistochemical labeling for tyrosine hydroxylase and c-fos protein was performed all along the ventrolateral medulla after microinjection of 1-(m-chlorophenyl)-biguanide, a selective and potent 5-HT(3) receptor agonist, into the nucleus tractus solitarii of alpha-chloralose/urethane-anaesthetized rats. This treatment produced a significant elevation of arterial pressure ( approximately +35 mm Hg). Concomitantly, a significant increase in the number of c-fos expressing neurons was observed in the rostral ventrolateral medulla (+63%), in particular in its most anterior part (+78%), and in the medullary region surrounding the caudal part of the facial nucleus (+91%). Retrograde labeling with gold-horseradish peroxidase complex showed that at least some of these activated c-fos expressing cells project to the spinal cord. However, the number of double-stained neurons, i.e. c-fos and tyrosine hydroxylase positive neurons, did not increase at any level of the ventrolateral medulla. In contrast, under the same alpha-chloralose/urethane anesthesia, systemic infusion of sodium nitroprusside appeared to produce a hypotension and a marked increase in the density of such double c-fos and tyrosine hydroxylase expressing cells in the rostral ventrolateral medulla and the caudal medullary region surrounding the caudal part of the facial nucleus. These data indicate that medullary catecholaminergic C1 and A1 neurons are not involved in the pressor effect elicited by 5-HT(3) receptor stimulation in the nucleus tractus solitarii. However, this 5-HT(3) receptor-mediated effect is clearly associated with the excitation of (non-catecholaminergic) neurons within the pressor region of the ventral medulla.

Stimulation of 5-HT2 receptors in the nucleus tractus solitarius enhances NMDA receptor-mediated reflex-evoked bradycardiac responses in the rat.

The modulation by 5-HT2 receptors in the nucleus tractus solitarius of the reflex bradycardia evoked by stimulation of peripheral baroreceptors and cardiopulmonary chemoreceptors, and their possible functional interactions with local NMDA receptors, were investigated in pentobarbital- and urethane-anaesthetized rats, respectively. Microinjection of the 5-HT2 receptor agonist, 2,5-dimethoxy-4-iodoamphetamine (0.1-0.5 pmol), into the nucleus tractus solitarius elicited a dose-dependent hypotension and bradycardia. Bilateral microinjections at the same site of a subthreshold dose of 2,5-dimethoxy-4-iodoamphetamine (0.05 pmol) significantly enhanced the aforementioned reflex-evoked bradycardiac responses. In contrast, local bilateral microinjections of the NMDA receptor antagonist, 2-amino-5-phosphonopentanoic acid (500 and 1000 pmol), reduced, in a dose-dependent manner, both reflex-evoked responses. The facilitatory effect of 2,5-dimethoxy-4-iodoamphetamine upon these reflex-evoked bradycardiac responses was prevented by prior local microinjection of low doses of either the selective 5-HT2 receptor antagonist, ketanserin (10 pmol), or 2-amino-5-phosphonopentanoic acid (100 pmol), which, on their own, did not affect the reflex-associated bradycardia. These data suggest that 5-HT2 receptors within the nucleus tractus solitarius participate in a facilitatory modulation of the reflex control of heart rate, probably through functional interactions with local NMDA receptors.

Microinjection of a 5-HT3 receptor agonist into the NTS of awake rats inhibits the bradycardic response to activation of the von Bezold-Jarisch reflex.

In the present study we investigated the effects of bilateral microinjection into the lateral commissural nucleus tractus solitarius (NTS) of 2-methyl-5-HT, a 5-HT3 receptor agonist, on the bradycardic response of the von Bezold-Jarisch reflex of awake rats. We evaluated mainly the bradycardic response because in previous studies we documented that the hypotensive response of the von-Bezold-Jarisch reflex in awake rats is secondary to the intense bradycardic response. The Bezold-Jarisch reflex was activated by intravenous injection of serotonin (8 microg/kg) in awake rats before and 1, 3, 10, 20 and 60 min after bilateral microinjection of 2-methyl-5-HT (5 nmol/50 nl, n = 8) into the NTS. Microinjections of 2-methyl-5-HT into the NTS produced a significant increase in basal mean arterial pressure [(MAP), 97 +/- 4 vs. 114 +/- 4 mmHg), no changes in basal heart rate and a significant reduction in bradycardic (-78 +/- 19; -94 +/- 24 and -107 +/- 21 bpm) and hypotensive (-16 +/- 4; -10 +/- 5 and -17 +/- 4 mmHg) responses to activation of the von Bezold-Jarisch reflex at 3, 10 and 20 min, respectively, when compared with the control value (-231 +/- 13 bpm and -43 +/- 4 mmHg). The data of the present study suggest that serotonin acting on 5-HT3 receptors in the NTS may play an important inhibitory neuromodulatory role in the bradycardic response to activation of the von Bezold-Jarisch reflex.

Activation of GABA receptors in the NTS of awake rats reduces the gain of baroreflex bradycardia.

In the present study we evaluated the effects of bilateral microinjection of muscimol (a GABA(A) receptor agonist) and baclofen (a GABA(B) receptor agonist) into the lateral commissural nucleus tractus solitarii (NTS) of awake rats on the gain of the baroreflex (BG) activated by a short duration (10-15 s) infusion of phenylephrine (Phe, 2.5 microg/0.05 ml, i.v.). Microinjection of muscimol (50 pmol/50 nl, n=8) into the NTS produced a significant increase in baseline mean arterial pressure ((MAP) 122+/-6 vs. 101+/-2 mmHg), no changes in baseline heart rate (HR) and a reduction in BG (-1.59+/-0. 1 vs. -0.69+/-0.1 beats/mmHg). Microinjection of baclofen (6.25 pmol/50 nl, n=6) into the NTS also produced a significant increase in baseline MAP (138+/-5 vs. 103+/-2 mmHg), no changes in baseline HR and a reduction in BG (-1.54+/-0.3 vs. -0.53+/-0.2 beats/mmHg). Considering that the reduction in BG could be secondary to the increase in MAP in response to microinjection of muscimol (n=6) or baclofen (n=7) into the NTS, in these two groups of rats we brought the MAP back to baseline by infusion of sodium nitroprusside (NP, 3.0 microg/0.05 ml, i.v.). Under these conditions, we verified that the BG remained significantly reduced after muscimol (-1.49+/-0.2 vs. -0.35+/-0.2 beats/mmHg) and after baclofen (-1.72+/-0.2 vs. -0.33+/-0.2 beats/mmHg) when compared to control. Reflex tachycardia was observed during the normalization of MAP by NP infusion and, in order to prevent the autonomic imbalance from affecting BG, we used another group of rats treated with atenolol (5 mg/kg, i.v.), a beta1 receptor antagonist. In rats previously treated with atenolol and submitted to NP infusion, we verified that BG remained reduced after microinjection of muscimol or baclofen into the NTS. The data show that activation of GABA(A) and GABA(B) receptors, independently of the changes in the baseline MAP or HR, inhibited the neurons of the NTS involved in the parasympathetic component of the baroreflex.

Bradycardic responses to microinjection of N-methyl-D-aspartate into the nucleus tractus solitarius are inhibited by local activation of 5-HT(3) receptors.

Previous reports have described that glutamate ionotropic receptors in the nucleus tractus solitarius (NTS) are involved in the reflex control of heart rate, and that such a control can be inhibited by NTS-5-HT(3) receptor stimulation. In the present study, we examined in urethane anaesthetized rats the effects of intra-NTS microinjection of 1-(m-chlorophenyl)-biguanide (CPBG), a potent and selective 5-HT(3) receptor agonist, on the cardiovascular responses to local administration of glutamate ionotropic receptor agonists. Intra-NTS microinjection of CPBG reduced the atropine-sensitive bradycardia elicited by local microinjection of NMDA without affecting the cardiovascular responses to intra-NTS microinjections of AMPA or kainic acid. The reduction by CPBG of the NMDA-evoked cardiac response was blocked by prior intra-NTS microinjection of granisetron, a 5-HT(3) receptor antagonist, as well as bicuculline, a GABA(A) receptor antagonist. These results suggest that the stimulation of NTS 5-HT(3) receptors specifically reduces, via a GABA-dependent mechanism, the cardiac response to local NMDA administration.

Activation of GABAA but not GABAB receptors in the NTSblocked bradycardia of chemoreflex in awake rats.

In the present study we analyzed effects of bilateral microinjections of muscimol (a GABAA agonist) and baclofen (a GABAB agonist) into the nucleus tractus solitarius (NTS) on bradycardic and pressor responses to chemoreflex activation (potassium cyanide, 40 micrograms/rat iv) in awake rats. Bilateral microinjections of muscimol (25 and 50 pmol/50 nl) into the NTS increased baseline mean arterial pressure (MAP): 119 +/- 8 vs. 107 +/- 2 mmHg (n = 6) and 121 +/- 8 vs. 103 +/- 3 mmHg (n = 6), respectively. Muscimol at 25 pmol/50 nl reduced the bradycardic response to chemoreflex activation 5 min after microinjection; with 50 pmol/50 nl the bradycardic response to chemoreflex activation was reduced 5, 15, 30, and 60 min after microinjection. Neither muscimol dose produced an effect on the pressor response of the chemoreflex. Effects of muscimol (50 pmol/50 nl) on basal MAP and on the bradycardic response of the chemoreflex were prevented by prior microinjection of bicuculline (a GABAA antagonist, 40 pmol/50 nl) into the NTS. Bilateral microinjections of baclofen (12.5 and 25 pmol/50 nl) into the NTS produced an increase in baseline MAP [137 +/- 9 vs. 108 +/- 4 (n = 7) and 145 +/- 5 vs. 105 +/- 2 mmHg (n = 7), respectively], no changes in basal heart rate, and no effects on the bradycardic response; 25 pmol/50 nl only attenuated the pressor response to chemoreflex activation. The data show that activation of GABAA receptors in the NTS produces a significant reduction in the bradycardic response, whereas activation of GABAB receptors produces a significant reduction in the pressor response of the chemoreflex. We conclude that 1) GABAA but not GABAB plays an inhibitory role in neurons of the lateral commissural NTS involved in the parasympathetic component of the chemoreflex and 2) attenuation of the pressor response of the chemoreflex by activation of GABAB receptors may be due to inhibition of sympathoexcitatory neurons in the NTS or may be secondary to the large increase in baseline MAP produced by baclofen.

Dorsal medullary 5-HT3 receptors and sympathetic premotor neurones in the rat.

1. Our aim was to determine whether the cardiovascular neurones in the rostro-ventrolateral medulla (CV-RVLM neurones) were involved in the sympathoexcitation induced by stimulation of 5-HT3 receptors in the region of the nucleus tractus solitarii (NTS). Experiments were performed in pentobarbitone-anaesthetized rats, artificially ventilated and paralysed with pancuronium bromide. 2. Using extracellular recordings, different types of RVLM neurones were characterized: cardiovascular (CV), ventilation-related and baroreflex-insensitive (unidentified) neurones. The CV-RVLM cells were further subdivided into three populations according to their axonal conduction velocities: A (1.2 +/- 0.1 m s-1), B (2.5 +/- 0.2 m s-1) and C (6.8 +/- 1.1 m s-1). 3. Only the CV-RVLM neurones of the A and B categories were partially inhibited (-30 %) by a hypotensive dose (2.5 microg kg-1 i.v.) of clonidine. 4. Microinjections into the region of the commissural NTS of 1-(m-chlorophenyl)-biguanide (CPBG, 2 nmol), a selective 5-HT3 receptor agonist, elicited an increase in both lumbar sympathetic nerve discharge (SND) and arterial pressure. In addition, this treatment produced a marked excitation of CV-RVLM neurones of the A and B categories, without affecting those of the C type, as well as ventilation-related and unidentified RVLM cells. 5. The activity of the CV neurones in the caudo-ventrolateral part of the medulla oblongata (CV-CVLM) was not modified by 5-HT3 receptor stimulation in the NTS. 6. Prior intra-NTS microinjections of ondansetron (300 pmol, a selective 5-HT3 receptor antagonist) into the region of the commissural NTS prevented the excitation of A and B CV-RVLM neurones induced by CPBG. 7. Intracarotid administration of saline saturated with CO2 (chemoreceptor activation) elicited both an increase in the SND and an excitation of the clonidine-insensitive CV-RVLM neurones of the C type, without affecting A and B neurones. 8. In conclusion, the sympathoexcitation elicited following 5-HT3 receptor stimulation in the region of the commissural NTS of pentobarbitone-anaesthetized rats seems to result from the excitation of two different pools of clonidine-sensitive CV-RVLM neurones. These neurones are apparently not involved in the sympathetic component of the chemoreceptor reflex.

c-Fos induction in the rostroventrolateral medulla by 5-HT3 receptor activation in the nucleus tractus solitarius.

The nucleus tractus solitarius (NTS) plays a crucial role in the reflex control of sympathetic tone through direct projections to the caudal and rostral regions of the ventrolateral medulla (CVLM and RVLM respectively). 5-HT3 receptor stimulation in the NTS increases both blood pressure and sympathetic tone. In order to investigate whether the ventrolateral medulla participates in these effects, the expression of c-Fos protein, a marker of neuronal activation, was examined at this level after intra-NTS microinjections of 1-(m-chlorophenyl)-biguanide, a selective 5-HT3 receptor agonist. A marked increase (93%) in the density of c-Fos-immunoreactive neurons was restricted to the most rostral part of the RVLM after this treatment. This effect could be prevented by prior intra-NTS microinjection of ondansetron, a selective 5-HT3 antagonist. These results suggest the participation of sympathoexcitatory neurons in the rostral RVLM in the pressor response to 5-HT3 receptor stimulation in the NTS.

Cardiovascular effects of microinjection of low doses of serotonin into the NTS of unanesthetized rats.

In the present study, we analyzed in conscious rats the effects of microinjections of serotonin (5-HT; pmol range) into the nucleus of the solitary tract (NTS) on basal mean arterial pressure (MAP) and heart rate (HR) and also on the reflex bradycardia induced by the activation of the baro- and chemoreflex evaluated 1 min after 5-HT microinjection into the NTS. The data show that unilateral microinjection of 5-HT in the picomolar range into the NTS of unanesthetized rats produced a dose-dependent decrease in MAP and HR, which was blocked by previous microinjection of ketanserin (250 pmol/50 nl) into the NTS. The changes in MAP and HR induced by 5-HT were of very short duration, with a return to baseline values a few seconds later. The cardiovascular responses to baro- or chemoreflex activation 1 min after 5-HT microinjection into the NTS did not differ from the control, indicating that low doses of 5-HT produced no effect on the cardiovascular reflexes tested at that time. The present data show that, as also observed in anesthetized rats, the microinjection of picomolar doses of 5-HT into the NTS elicits the typical cardiovascular responses to baroreceptor activation. These effects, hypotension and bradycardia, seem to be mediated by 5-HT2 receptors because both were blocked by a selective 5-HT2 receptor antagonist. However, since microinjection of 5-HT (1 pmol) into the NTS produced no changes in the cardiovascular responses to the baro- and chemoreflex activated 1 min later, the role of 5-HT2 receptors in the processing of the cardiovascular afferent messages in the NTS remains to be elucidated.

Microinjection of a serotonin3 receptor agonist into the NTS of unanesthetized rats inhibits the bradycardia evoked by activation of the baro- and chemoreflexes.

In the present study we investigated the effects of microinjection into the commissural nucleus tractus solitarius (NTS) of unanesthetized rats of 2-methylserotonin (2-methyl-5-HT), a 5-HT3 receptor agonist, on the cardiac component of the baro- and chemoreflexes. The study was performed in conscious freely moving rats in order to avoid the possible effects of anesthetics on the cardiovascular responses to microinjection of neuroactive substances into the NTS. The baroreflex (phenylephrine, 0.5-2.0 micrograms/kg, i.v.) and the chemoreflex (potassium cyanide, 40 micrograms/rat, i.v) were activated in different groups of rats before and after bilateral microinjection of 2-methyl-5-HT into the NTS. Microinjections of 2-methyl-5-HT (5 nmol/50 nl) into the NTS produced a significant increase in basal mean arterial pressure (101 +/- 3 versus 125 +/- 8 mmHg), no changes in basal HR and a significant reduction in the reflex bradycardia triggered by baroreflex activation at 3 (-28 +/- 7 bpm), 10 (-35 +/- 4 bpm) and 20 min (-34 +/- 5 bpm) in comparison with the control value (-68 +/- 9 bpm). A similar reduction in the bradycardic response to chemoreflex activation was observed at 3 (-94 +/- 35 bpm), 10 (-98 +/- 38 bpm) and 20 min (-110 +/- 29 bpm) after 2-methyl-5-HT in comparison with the control value (-178 +/- 19 bpm). The effect of 2-methyl-5-HT on the basal mean arterial pressure and on the bradycardia evoked by stimulation of the baro- and chemoreflexes was blocked by pretreatment with granisetron bilaterally microinjected (500 pmol/50 nl) into the NTS. The data show that the stimulation of 5-HT3 receptors in the NTS of unanesthetized rats elicits a significant increase in basal mean arterial pressure and decreases the bradycardic response to baro- or chemoreflex activation.

Role of serotonin3 receptors in the nucleus tractus solitarii on the carotid chemoreflex.

The effects of serotonin3 (5-HT3)-receptor stimulation in the nucleus tractus solitarii (NTS) on the cardiovagal, sympathetic, and respiratory responses to activation of carotid body chemoreceptors were investigated in anesthetized rats. The chemoreflex responses were triggered by an intravenous administration of KCN (40 microg/kg) in spontaneously breathing urethan-chloralose-anesthetized rats or by an intracarotid administration of saline saturated with 100% CO2 in pancuronium bromide-paralyzed and artificially ventilated urethan-anesthetized rats. Microinjections of 5-HT (2.5-5 nmol) or the 5-HT3 agonist 1-(m-chlorophenyl)-biguanide (CPBG, 300-1,200 pmol) into the commissural NTS blocked in a dose-dependent manner the atropine-sensitive chemoreflex bradycardia elicited by KCN. However, neither 5-HT nor CPBG affected the KCN-induced increase in respiratory volume and the CO2-induced increases in blood pressure and lumbar sympathetic nerve discharge. The inhibitory effect of 5-HT or CPBG on KCN-induced bradycardia was blocked by prior intra-NTS microinjection of a 5-HT3 antagonist, such as zacopride (100 pmol) or ondansetron (100 pmol), or the A-type gamma-aminobutyric acid (GABA(A)) antagonist bicuculline (10 pmol). In contrast, local microinjections of antagonists acting at 5-HT1 and 5-HT2 receptors, such as methysergide (100 pmol) and ketanserin (10 pmol), respectively, did not prevent the actions of 5-HT or CPBG. These data show that the stimulation of 5-HT3 receptors in the NTS exerted an inhibitory influence, probably through the activation of a local GABAergic system, on the cardiovagal component of the chemoreflex. Because similar effects of 5-HT3-receptor stimulation in the NTS were previously found on the baroreflex and Bezold-Jarisch reflex responses, it can be inferred that NTS 5-HT3 receptors play a key modulatory role in the reflex control of the heart rate.

Medullary pathways of cardiovascular responses to 5-HT2 and 5-HT3 receptor stimulation in the rat nucleus tractus solitarius.

As previously found for the baroreceptor reflex, microinjection of kynurenic acid (KYN, 2 nmol), a glutamate receptor antagonist, into the caudal ventrolateral medulla (CVL) blocked the hypotension and bradycardia elicited by microinjections of a 5-HT2 receptor agonist, 2,5-dimethoxy-4-iodoamphetamine (DOI, 0.5 pmol) into the nucleus tractus solitarius (NTS). In addition, as previously observed with the sympathetic chemoreflex response, microinjections of KYN into the RVI, blocked the increase in blood pressure elicited by microinjections of 1-m-(chlorophenyl)-biguanide (1200 pmol), a 5-HT1 receptor agonist, into the NTS. These results suggest that medullary pathways involved in the responses to 5-HT2 and 5-HT3 receptor stimulation in the NTS are similar to those that mediate the baroceptor and chemoreceptor reflex responses, respectively.

Stimulation of 5-HT3 receptors in the NTS inhibits the cardiac Bezold-Jarisch reflex response.

Intra-atrial administration of phenylbiguanide has been shown to trigger, through the stimulation of vagal afferent C-fibers, reflex bradycardia, hypotension, and sympathoinhibition classically known as the Bezold-Jarisch (B-J) reflex (O. Krayer. Naunyn-Schmiedeberg's Arch. Exp. Pathol. Pharmacol. 240: 361-368, 1961). The effects of microinjections, into the nucleus tractus solitarius (NTS), of serotonin (5-HT) and 1-(m-chlorophenyl)-biguanide (CPBG), a potent 5-HT3 receptor agonist, on these reflex responses were studied in urethananesthetized rats. 5-HT (600 and 900 pmol) and CPBG (10-150 pmol) produced a dose-dependent inhibition of the atropine-sensitive bradycardiac component of the B-J reflex. The effect of both agonists was reversed by prior local microinjection of the 5-HT3 receptor antagonists zacopride (100 pmol) and ondansetron (100 pmol), but not by that of the 5-HT2 receptor antagonist ketanserin (10 pmol) or the mixed 5-HT1/5-HT2 receptor antagonist methysergide (100 pmol). In contrast, CPBG (150 pmol) did not affect the B-J reflex inhibition of lumbar sympathetic nerve discharge. These results show that stimulation of NTS 5-HT3 receptors produced an inhibition of the cardiovagal component of the B-J reflex without affecting its sympathetic component. Because the stimulation of these receptors also inhibits the cardiac component of the baroreflex, the present data suggest the participation of NTS 5-HT3 receptors in the mechanisms that modulate cardiac reflex responses elicited by messages from different vagal afferents.

Cardiovascular effects induced by the stimulation of neuropeptide FF receptors in the dorsal vagal complex: an autoradiographic and pharmacological study in the rat.

Previous studies have suggested that central administration of neuropeptide FF-related peptides may modulate cardiovascular parameters in the rat. In the present study, we investigated the role of dorsal vagal complex neuropeptide FF receptors in the central regulation of cardiovascular parameters. The fate of neuropeptide FF receptors in normal and nodose ganglionectomized rats was investigated using an autoradiographic approach with 125I-[DTyr1, (NMe)Phe3]NPFF as ligand for these receptors. We showed that neuropeptide FF binding sites are preferentially located postsynaptically with respect to the vagal afferent fibers in the nucleus tractus solitarius. Thus, ganglionectomy reduced by only 30% and 17% the density of peptide binding sites in the rostral and caudal regions of this nucleus, respectively. Bilateral microinjection of neuropeptide FF (1 nmol) into the commissural nucleus tractus solitarius produced an increase in blood pressure (+13.8 +/- 0.8 mmHg, n = 6), bradycardia (-29.0 +/- 3.2 bpm) and a significant inhibition (-47.6 +/- 3.1%) of the cardiac component of the baroreceptor reflex. Further studies with doses below 1 nmol indicate that NTS microinjections of the neuropeptide produced a dose-dependent decrease in heart rate. Similar cardiovascular effects were observed after bilateral NTS microinjections of one analog neuropeptide FF receptor agonist, [DTyr1, (NMe)Phe3]NPFF (1 nmol). Pretreatment with prazosin (100 micrograms/kg), an alpha 1-adrenoreceptor antagonist, inhibited the neuropeptide FF-evoked blood pressure effect. In addition, the neuropeptide FF-induced heart rate decrease was abolished by pretreatment with atropine (30 micrograms/kg), a muscarinic receptor antagonist. Taken together, these anatomical and pharmacological data suggest that neuropeptide FF receptors within the nucleus tractus solitarius, preferentially located on the postsynaptic component, are involved in the central reflex regulation of cardiovascular parameters.

Mechanisms underlying the cardiovascular responses to peripheral administration of NPFF in the rat.

In the present study, we examined the possibility of the presence of the Phe-Leu-Phe-Gln-Pro-Gln-Arg-Phe-NH2 (NPFF) system in the rat heart as well as the effects of drugs affecting noradrenergic transmission upon the cardiovascular responses elicited by peripheral administration of NPFF. The presence of NPFF receptors on heart sections and of NPFF-immunoreactivity in heart tissue was demonstrated with autoradiographic and radioimmunoassay procedures, respectively. Intravenous administration of NPFF (100-300 micrograms/kg) produced a dose-dependent increase in blood pressure and heart rate without affecting plasma noradrenaline and adrenaline levels. These effects of NPFF were also observed, although attenuated, in catecholamine-depleted rats and in rats pretreated with a ganglionic blocking agent, hexamethonium (10 mg/kg, i.v.). Prazosin (100 micrograms/kg, i.v.), an alpha1 adrenergic receptor antagonist, reduced the NPFF-induced blood pressure response by 50%. In contrast, propranolol (2 mg/kg, i.v.) and metroprolol (0.5 mg/kg, i.v.), beta- and beta1 adrenergic receptor antagonists, respectively, reduced the NPFF-induced heart rate response by 50%. Surprisingly, the alpha2 adrenergic receptor antagonists, idazoxan (2 mg/kg, i.v.) and yohimbine (2 mg/kg, i.v.), both produced a drastic increase in the NPFF-induced heart rate response. These data, which demonstrate the presence of the NPFF system in the rat heart, suggest that the cardiovascular responses of peripheral administration of NPFF are mediated by the stimulation of peripheral NPFF receptors. In addition, the present data show that the aforementioned NPFF-induced responses are also mediated by catecholamine-dependent mechanisms and suggest a functional interaction between adrenergic and NPFF systems.