Synaptic profile of nucleus tractus solitarius neurons involved with the peripheral chemoreflex pathways.

The glomus cells in the carotid bodies (CB) detect alterations in pH and pCO2 and low pO2 level in arterial blood. The carotid sinus nerve conveys the information related to the oxygen level to 2nd-order neurons in the nucleus tractus solitarius (NTS) via tractus solitarius (TS), which is part of the chemoreflex pathways. It has been demonstrated that in 2nd-order NTS neurons receiving inputs from the aortic depressor nerve (ADN), the TS stimulation presents high temporal fidelity. However, the temporal properties of synaptic activity in NTS neurons receiving inputs from CB were not yet fully investigated. Herein using patch-clamp recordings in NTS brainstem slices, we studied TS-evoked excitatory postsynaptic currents (TS-eEPSCs) on morphologically identified 2nd-order NTS neurons that receive afferent inputs from the CB and compared with 2nd-order ADN-NTS neurons recorded in the same experimental conditions. The amplitudes of TS-eEPSCs were similar in both groups, but the latencies and standard deviation (SD) of latency were significantly higher in the CB-NTS neurons (latency: 4±0.2 ms, SD: 0.49±0.03 ms) than in ADN-NTS neurons (latency: 3.3±0.3 ms, SD: 0.19±0.02 ms; P=0.049 for latency and P<0.001 for SD of latency). In a series of double-labeling experiments, we confirmed that some CB-NTS 2nd-order neurons send direct projections to the rostral ventrolateral medulla (RVLM). We conclude that: (a) CB-NTS 2nd-order neurons present temporally distinct postsynaptic currents when compared with ADN-NTS 2nd-order neurons; (b) low SD of latency of TS-eEPSCs is not necessarily a characteristic of all 2nd-order neurons in the NTS; and (c) the presence of direct connections between these 2nd-order neurons in the NTS and RVLM is indicative that these synaptic properties of CB-NTS neurons are relevant for the processing of respiratory and autonomic responses to chemoreflex activation.

Multiple pontomedullary mechanisms of respiratory rhythmogenesis.

Mammalian central pattern generators producing rhythmic movements exhibit robust but flexible behavior. However, brainstem network architectures that enable these features are not well understood. Using precise sequential transections through the pons to medulla, it was observed that there was compartmentalization of distinct rhythmogenic mechanisms in the ponto-medullary respiratory network, which has rostro-caudal organization. The eupneic 3-phase respiratory pattern was transformed to a 2-phase and then to a 1-phase pattern as the network was physically reduced. The pons, the retrotrapezoid nucleus and glycine mediated inhibition are all essential for expression of the 3-phase rhythm. The 2-phase rhythm depends on inhibitory interactions (reciprocal) between Bötzinger and pre-Bötzinger complexes, whereas the 1-phase-pattern is generated within the pre-Bötzinger complex and is reliant on the persistent sodium current. In conditions of forced expiration, the RTN region was found to be essential for the expression of abdominal late expiratory activity. However, it is unknown whether the RTN generates or simply relays this activity. Entrained with the central respiratory network is the sympathetic nervous system, which exhibits patterns of discharge coupled with the respiratory cycle (in terms of both gain and phase of coupling) and dysfunctions in this coupling appear to underpin pathological conditions. In conclusion, the respiratory network has rhythmogenic capabilities at multiple levels of network organization, allowing expression of motor patterns specific for various physiological and pathophysiological respiratory behaviors.

Intermittent activation of peripheral chemoreceptors in awake rats induces Fos expression in rostral ventrolateral medulla-projecting neurons in the paraventricular nucleus of the hypothalamus.

Despite the well-established sympathoexcitation evoked by chemoreflex activation, the specific sub-regions of the CNS underlying such sympathetic responses remain to be fully characterized. In the present study we examined the effects of intermittent chemoreflex activation in awake rats on Fos-immunoreactivity (Fos-ir) in various subnuclei of the paraventricular nucleus of the hypothalamus (PVN), as well as in identified neurosecretory preautonomic PVN neurons. In response to intermittent chemoreflex activation, a significant increase in the number of Fos-ir cells was found in autonomic-related PVN subnuclei, including the posterior parvocellular, ventromedial parvocellular and dorsal-cap, but not in the neurosecretory magnocellular-containing lateral magnocellular subnucleus. No changes in Fos-ir following chemoreflex activation were observed in the anterior PVN subnucleus. Experiments combining Fos immunohistochemistry and neuronal tract tracing techniques showed a significant increase in Fos-ir in rostral ventrolateral medulla (RVLM)-projecting (PVN-RVLM), but not in nucleus of solitarii tract (NTS)-projecting PVN neurons. In summary, our results support the involvement of the PVN in the central neuronal circuitry activated in response to chemoreflex activation, and indicate that PVN-RVLM neurons constitute a neuronal substrate contributing to the sympathoexcitatory component of the chemoreflex.

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.

Antagonism of adenosine A(1) receptors in the NTS does not affect the chemoreflex in awake rats.

The possible involvement of adenosine A(1) receptors in neurotransmission of the sympathoexcitatory component of the chemoreflex in the nucleus tractus solitarii (NTS) of awake rats was evaluated. Unilateral microinjection of increasing doses of adenosine (0.01, 0.06, 0.12, 1.25, 2.5, and 5.0 nmol/50 nl) into the lateral aspect of the commissural NTS produced a long-lasting increase in baseline mean arterial pressure (MAP) and no changes in baseline heart rate (HR). Microinjection of adenosine at 1.25 nmol/50 nl (ED(50)) into the NTS (n = 9) produced a significant increase in baseline MAP (119 +/- 3, 122 +/- 4, and 117 +/- 4 mmHg at 30 s, 1 min, and 2 min, respectively) compared with control (102 +/- 3 mmHg) but no significant changes after previous microinjection of 8-cyclopentyl-1,3-dipropylxanthine (DPCPX), an adenosine A(1) receptor antagonist (107 +/- 3, 107 +/- 3, and 106 +/- 3 mmHg at 30 s, 1 min, and 2 min, respectively) compared with control (102 +/- 3 mmHg). Microinjection of adenosine before and after DPCPX into the same site of the lateral commissural NTS produced no changes in baseline HR. In another group of rats (n = 8), microinjection of DPCPX (0.285 nmol/50 nl) into lateral and midline aspects of the commissural NTS produced no significant changes in pressor (+46 +/- 4 vs. +47 +/- 2 mmHg) or bradycardic responses (-216 +/- 9 vs. -226 +/- 12 beats/min) to chemoreflex activation with intravenous potassium cyanide compared with control responses. These data show that microinjection of adenosine into the NTS produced a small and long-lasting pressor response by activating A(1) receptors and that blockade of these receptors produced no changes in cardiovascular responses to chemoreflex activation. We conclude that adenosine A(1) receptors are not involved in processing of the chemoreflex afferents at the NTS level.

Pressor response to unilateral carotid chemoreceptor activation is not affected by ipsilateral antagonism of excitatory amino acid receptors in the rostral ventrolateral medulla of awake rats.

The importance of the integrity of the ipsilateral rostral ventrolateral medulla (RVLM) in the pressor response to activation of unilateral arterial chemoreceptors was evaluated. To achieve this goal, the left carotid body artery was ligated prior to the experiment and a guide cannula was implanted in the direction of the right RLVM, i.e. the side where the carotid body artery was kept intact. On the day of the experiment, the chemoreflex was activated with potassium cyanide (KCN, i.v.) before and after unilateral microinjection of kynurenic acid into the rostral or caudal aspect of the RVLM. The data indicated that microinjection of kynurenic into the rostral or caudal aspect of the RVLM produced no effect on the pressor response of chemoreflex activation. These data suggest that chemoreflex activation excites a neuronal network in which the processing of the sympatho-excitatory component of the chemoreflex is not restricted to an excitatory projection from the nucleus tractus solitarii to the ipsilateral RVLM.

Changes in regional vascular resistance in response to microinjection of L-glutamate into different antero-posterior coordinates of the RVLM in awake rats.

Changes in mean arterial pressure (MAP) and in regional vascular resistance (RVR, hindquarter, mesenteric and renal) induced by microinjection of L-glutamate into three different antero-posterior coordinates of the rostral ventrolateral medulla (RVLM) [1,200-1,600 (microm (n=10), 1,601-2,000 microm (n=12) and 2,001-2,500 microm (n=6) rostral to the obex] were investigated in unanesthetized rats. Guide cannulas directed towards the RVLM were implanted 4 days prior to the experiments. Doppler probes were implanted around the superior mesenteric, inferior abdominal aorta and left renal arteries and a catheter was inserted into the femoral artery and vein 1 day prior to the experiments. Insertion of the injector into the RVLM produced an increase in baseline MAP, which was back to control levels 2 min later, when L-glutamate was microinjected. Microinjection of L-glutamate (1 nmol/30 nl) into the three antero-posterior coordinates of the RVLM produced an increase in MAP associated with a similar increase in hindquarter, mesenteric and renal vascular resistance, which were back to control 1 min later. Saline into the RVLM produced negligible effects on MAP and RVR. These findings suggest that the sympathetic vasomotor neurons involved in the regulation of the regional vascular resistance in rats are not topographically distributed in the antero-posterior coordinates of the RVLM. However, the experimental methods used to evaluate the topographic distribution of sympatho-vasomotor neurons in the RVLM and the measurement of the regional blood flow may not be precise enough to detect any possible differences.

Neurotransmission of the cardiovascular reflexes in the nucleus tractus solitarii of awake rats.

Chemoreflex activation with potassium cyanide (i.v.) produces pressor and bradycardic responses in awake rats. Microinjection of AP-5, a selective NMDA receptor antagonist, into the nucleus tractus solitarii (NTS) produced a dose-dependent blockade of the bradycardic response; while microinjection of DNQX, a selective non-NMDA receptor antagonist, or kynurenic acid, a nonselective ionotropic receptor antagonist, produced only a partial reduction in the pressor response, indicating that the bradycardic component of the chemoreflex is mediated by NMDA receptors and that the sympathoexcitatory component may involve neurotransmitters other than excitatory amino acids. With respect to the baroreflex, we verified that the gain of baroreflex bradycardia in response to phenyleprine (Phe) infusion was significantly reduced in a dose-dependent manner by microinjection of AP-5 into the NTS, indicating that the parasympathetic component of the baroreflex is mediated mainly by NMDA receptors. However, in a series of experiments involving the electrical stimulation of the aortic depressor nerve (ADN) we observed that the maximal bradycardic response was almost blocked by the combination of microinjection of NMDA and non-NMDA receptor antagonists into the NTS, while the depressor response was only partially reduced. These data indicate that the bradycardic response produced by the activation of the baroreflex with Phe is mediated by mechanisms differing from those in response to the electrical stimulation of the ADN because phenylephrine also activates carotid and aortic baroreceptors, while unilateral electrical stimulation of the ADN involves only one specific set of baroreceptor afferents. These data also indicate that the sympatho inhibitory component of this response may involve neurotransmitters other than L-glutamate. We discuss the possibility that two different afferent systems of arterial baroreceptors are involved in the modulation of parasympathoexcitation and sympathoinhibition: one activated within the normal range of pulsatile arterial pressure (on a pulse-to-pulse basis) and the other acting under circumstances of challenge to the pulsatile arterial pressure above the normal range.

Involvement of the paraventricular nucleus of the hypothalamus in the pressor response to chemoreflex activation in awake rats.

Chemoreflex activation with potassium cyanide (KCN, i.v.) produces pressor and bradycardic responses in awake rats in addition to the tachypneic response. In the present study we evaluated the role of the paraventricular nucleus of the hypothalamus (PVN) in the cardiovascular responses to chemoreflex activation in awake rats. Bilateral electrolytic lesion of the PVN was performed 1 day before chemoreflex activation and the results were compared to those obtained with sham-lesioned rats. Bilateral electrolytic lesion of the PVN (n=6) produced a significant reduction in both the magnitude (+51+/-5 vs. +22+/-2 mmHg) and duration (+26+/-5 vs. +6+/-2 s) of the pressor response to chemoreflex activation when compared to sham-lesioned rats (n=10). The bradycardic response to chemoreflex activation in rats with bilateral lesion of the PVN was not significantly different from the response of sham-lesioned rats (-229+/-20 vs. -88+/-76 bpm). Unilateral or partial bilateral lesion of the PVN (n=10) produced no significant changes in the pressor response (+51+/-5 vs. +49+/-3 mmHg), in the duration of the response (+26+/-5 vs. +18+/-3 s) or in the bradycardic response (-229+/-20 vs. -230+/-27 bpm) compared to sham-lesioned rats. The data show that effective bilateral lesion of the PVN produced a significant reduction in the magnitude and duration of the pressor response, indicating that the PVN plays a key role in the processing of the sympathoexcitatory component of the chemoreflex.

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.

Blockade of NK-1 receptors in the lateral commissural nucleus tractus solitarii of awake rats had no effect on the cardiovascular responses to chemoreflex activation

The neurotransmission of the chemoreflex in the nucleus tractus solitarii (NTS), particularly of the sympatho-excitatory component, is not completely understood. There is evidence that substance P may play a role in the neurotransmission of the chemoreflex in the NTS. Microinjection of substance P (50 pmol/50 nl, N = 12, and 5 nmol/50 nl, N = 8) into the commissural NTS of unanesthetized rats produced a significant increase in mean arterial pressure (101 +/- 1 vs 108 +/- 2 and 107 +/- 3 vs 115 +/- 4 mmHg, respectively) and no significant changes in heart rate (328 +/- 11 vs 347 +/- 15 and 332 +/- 7 vs 349 +/- 13 bpm, respectively) 2 min after microinjection. Previous treatment with WIN, an NK-1 receptor antagonist (2.5 nmol/50 nl), microinjected into the NTS of a specific group of rats, blocked the pressor (11 +/- 5 vs 1 +/- 2 mmHg) and tachycardic (31 +/- 6 vs 4 +/- 3 bpm) responses to substance P (50 pmol/50 nl, N = 5) observed 10 min after microinjection. Bilateral microinjection of WIN into the lateral commissural NTS (N = 8) had no significant effect on the pressor (50 +/- 4 vs 42 +/- 6 mmHg) or bradycardic (-230 +/- 16 vs -220 +/- 36 bpm) responses to chemoreflex activation with potassium cyanide (iv). These data indicate that the activation of NK-1 receptors by substance P in the NTS produces an increase in baseline mean arterial pressure and heart rate. However, the data obtained with WIN suggest that substance P and NK-1 receptors do not play a major role in the neurotransmission of the chemoreflex in the lateral commissural NTS

Changes in regional vascular resistance in response to microinjection of L-glutamate into different antero-posterior coordinates of the RVLM in awake rats.

Changes in mean arterial pressure (MAP) and in regional vascular resistance (RVR, hindquarter, mesenteric and renal) induced by microinjection of L.-glutamate into three different antero-posterior coordinates of the rostral ventrolateral medulla (RVLM) [1,200-1,600 microm (n=10), 1,601-2,000 microm (n=12) and 2,001-2,500 microm (n=6) rostral to the obex] were investigated in unanesthetized rats. Guide cannulas directed towards the RVLM were implanted 4 days prior to the experiments. Doppler probes were implanted around the superior mesenteric, inferior abdominal aorta and left renal arteries and a catheter was inserted into the femoral artery and vein 1 day prior to the experiments. Insertion of the injector into the RVLM produced an increase in baseline MAP, which was back to control levels 2 min later, when L-glutamate was microinjected. Microinjection of L-glutamate (1 nmol/30 nl) into the three antero-posterior coordinates of the RVLM produced an increase in MAP associated with a similar increase in hindquarter, mesenteric and renal vascular resistance, which were back to control 1 min later. Saline into the RVLM produced negligible effects on MAP and RVR. These findings suggest that the sympathetic vasomotor neurons involved in the regulation of the regional vascular resistance in rats are not topographically distributed in the antero-posterior coordinates of the RVLM. However, the experimental methods used to evaluate the topographic distribution of sympatho-vasomotor neurons in the RVLM and the measurement of the regional blood flow may not be precise enough to detect any possible differences.

Blockade of NK-1 receptors in the lateral commissural nucleus tractus solitarii of awake rats had no effect on the cardiovascular responses to chemoreflex activation.

The neurotransmission of the chemoreflex in the nucleus tractus solitarii (NTS), particularly of the sympatho-excitatory component, is not completely understood. There is evidence that substance P may play a role in the neurotransmission of the chemoreflex in the NTS. Microinjection of substance P (50 pmol/50 nl, N = 12, and 5 nmol/50 nl, N = 8) into the commissural NTS of unanesthetized rats produced a significant increase in mean arterial pressure (101 +/- 1 vs 108 +/- 2 and 107 +/- 3 vs 115 +/- 4 mmHg, respectively) and no significant changes in heart rate (328 +/- 11 vs 347 +/- 15 and 332 +/- 7 vs 349 +/- 13 bpm, respectively) 2 min after microinjection. Previous treatment with WIN, an NK-1 receptor antagonist (2.5 nmol/50 nl), microinjected into the NTS of a specific group of rats, blocked the pressor (11 +/- 5 vs 1 +/- 2 mmHg) and tachycardic (31 +/- 6 vs 4 +/- 3 bpm) responses to substance P (50 pmol/50 nl, N = 5) observed 10 min after microinjection. Bilateral microinjection of WIN into the lateral commissural NTS (N = 8) had no significant effect on the pressor (50 +/- 4 vs 42 +/- 6 mmHg) or bradycardic (-230 +/- 16 vs -220 +/- 36 bpm) responses to chemoreflex activation with potassium cyanide (iv). These data indicate that the activation of NK-1 receptors by substance P in the NTS produces an increase in baseline mean arterial pressure and heart rate. However, the data obtained with WIN suggest that substance P and NK-1 receptors do not play a major role in the neurotransmission of the chemoreflex in the lateral commissural NTS.

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.

Neurotransmission of autonomic components of aortic baroreceptor afferents in the NTS of awake rats.

The effect of sequential blockade of N-methyl-D-aspartic acid (NMDA) receptors with DL-2-amino-5-phosphonopentanoic acid (AP-5) and non-NMDA receptors with 6,7-dinitroquinoxaline-2,3 dione (DNQX) in the nucleus tractus solitarii (NTS) on the cardiovascular responses to electrical stimulation (ES) of the aortic depressor nerve (ADN) was evaluated in awake rats. Two protocols were used. In protocol 1, bilateral microinjection of AP-5 into the NTS (n = 7) reduced the hypotensive response to ES of the ADN; subsequent microinjection of DNQX produced additional reduction in this response. AP-5 reduced the bradycardic response, and DNQX almost abolished this response. In protocol 2, bilateral microinjection of DNQX into the NTS (n = 6) reduced the hypotensive response, and subsequent microinjection of AP-5 significantly reduced this response. DNQX produced a significant reduction in bradycardic response, and AP-5 abolished this response. The data indicate that processing of the parasympathetic component of the NTS aortic baroreceptor afferents is mediated by both NMDA and non-NMDA receptors, whereas processing of the sympathoinhibitory component seems to be only partially mediated by ionotropic receptors.

Bradycardic and hypotensive responses to microinjection of L-glutamate into the lateral aspect of the commissural NTS are blocked by an NMDA receptor antagonist.

Baroreflex activation by phenylephrine infusion produces a bradycardic response while microinjection of L-glutamate into the most lateral aspect of the commissural nucleus tractus solitarius (NTS, 0.8 mm lateral to the midline) produces bradycardic and hypotensive responses. In the present study we investigated the role of NMDA receptors in the lateral aspect of the commissural NTS (0.8 mm lateral to the midline) in the bradycardic and hypotensive responses to microinjection of L-glutamate as well as in the processing of the bradycardic response to the baroreflex activation. The hypotensive and bradycardic responses to L-glutamate microinjection into the NTS were blocked by methyl-atropine (intravenous, i.v.), indicating that the hypotensive response was secondary to the bradycardia. Microinjection of L-glutamate (1 nmol/50 nl) into the NTS was performed before and after microinjection of increasing doses of phosphonovaleric acid (AP-5, a selective NMDA antagonist) at the same site. The microinjection of AP-5 [0.5 (n = 9), 2.0 (n = 8) and 10.0 nmol/50 nl (n = 7)] into the NTS (0.8 mm lateral to the midline) produced a dose-dependent blockade of the bradycardic and hypotensive responses to L-glutamate. In a specific group of rats the microinjection of 10 nmol/50 nl of AP-5 produced a significant reduction in baroreflex sensitivity 2 min after microinjection into the lateral NTS [gain = -1.48 +/- 0.12 vs. -0.5 +/- 0.2 beats/mmHg, (n = 5)], which was reversible. The data show that the bradycardic responses produced by microinjection of L-glutamate into the most lateral aspect of the commissural NTS or by activation of the baroreflex were blocked by microinjection of AP-5, indicating that the neurotransmission of the parasympathetic component of the baroreflex in the neurons of the lateral aspect of the commissural NTS involves NMDA receptors.

The gain of the baroreflex bradycardia is reduced by microinjection of NMDA receptor antagonists into the nucleus tractus solitarii of awake rats.

The baroreflex activation with phenylephrine infusion produces a bradycardic response. In the present study, the role of NMDA receptors in the nucleus tractus solitarii (NTS) in the processing of the parasympathetic component of the baroreflex was evaluated using acid phosphonivaleric (AP-5), a selective NMDA receptor antagonist. Baroreflex activation was performed before and after bilateral microinjection of AP-5 into the intermediate commissural NTS (0.5 mm lateral to the midline). Microinjection of the vehicle (saline, 0.9%) or a dose of 2 nmol/50 nl of AP-5 into the NTS produced no effect on the gain of the baroreflex while a dose of 10 nmol/50 nl of AP-5 produced a significant reduction in the gain of the baroreflex 2 min after microinjection [-1.43+/-0.22 vs. -0. 43+/-0.03 bpm/mmHg, (n=6)], with a return to control levels 10 min after the microinjections. The dose of 10 nmol/50 nl was selective for NMDA receptors considering that the cardiovascular responses to microinjection of AMPA (0.05 pmol/50 nl), a non-NMDA receptor agonist, were not affected by this dose of AP-5 and the responses to microinjection of NMDA (2 nmol/50 nl) were blocked. The data show that the bradycardic response to baroreflex activation was blocked by AP-5 microinjected into the NTS, indicating that the neurotransmission of the parasympathetic component of the baroreflex is mediated by NMDA receptors in the NTS.

Hemodynamic responses to electrical stimulation of the aortic depressor nerve in awake rats.

Changes in mean arterial pressure (MAP), heart rate (HR), and vascular resistance (hindquarter and mesenteric territories) in response to electrical stimulation (ES) of the aortic depressor nerve (ADN) were evaluated in conscious freely moving rats. Platinum electrodes were implanted into the ADN of all rats studied, and some of these animals were also implanted with miniaturized Doppler probes around the superior mesenteric artery and inferior abdominal aorta (hindquarter). In both groups, the femoral artery and vein were catheterized one day before the experiments. In the first group of rats (n = 7), the control ES of the ADN in the range from 0.5 to 3.0 V (50 Hz, 10 ms) produced bradycardia and hypotension in an intensity-dependent manner, and treatment with methylatropine (intravenously) blocked the bradycardia but produced no significant changes in the hypotensive response. In a second group (n = 6), ES of the ADN was performed with the intensity fixed at 3 V and the frequency of the stimuli varying from 10 to 50 Hz. In this group, the hypotensive response was frequency dependent, whereas the bradycardic response was not. In a third group of rats (n = 6), ES of the ADN (2.5 V) produced hypotension (-35 +/- 4 mmHg), minor changes in the mesenteric (+5 +/- 14%), and vasodilation in hindquarter (-32 +/- 6%) vascular beds. The data show that 1) ES of the ADN produces a fall in pressure, bradycardia, vasodilation in the hindquarter, and no changes in the mesenteric vascular resistance, 2) methylatropine blocked the bradycardia and produced no effect on the hypotensive response to ES of the ADN, and 3) the baroreceptor afferent fibers involved in the hypotensive response to ES of ADN are sensitive to the variation of the frequency of the stimuli, whereas the fibers involved in the bradycardic response are not.