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Am J Physiol Regul Integr Comp Physiol 280: R1285-R1291, 2001;
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Vol. 280, Issue 5, R1285-R1291, May 2001

Glutamate release via NO production evoked by NMDA in the NTS enhances hypotension and bradycardia in vivo

Isamu Matsuo, Yoshitaka Hirooka, Kiyoshi Hironaga, Kenichi Eshima, Hideaki Shigematsu, Miwako Shihara, Koji Sakai, and Akira Takeshita

Department of Cardiovascular Medicine, Cardiovascular Science, Graduate School of Medical Sciences, Kyushu University, Fukuoka 812-8582, Japan


    ABSTRACT
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES

Nitric oxide (NO) in the nucleus tractus solitarii (NTS) plays an important role in regulating sympathetic nerve activity. The aims of this study were to determine whether the activation of N-methyl-D-aspartate (NMDA) receptors in the NTS facilitates the release of L-glutamate (Glu) via NO production, and, if so, to determine whether this mechanism is involved in the depressor and bradycardic responses evoked by NMDA. We measured the production of NO in the NTS as NO<UP><SUB>2</SUB><SUP>−</SUP></UP> and NO<UP><SUB>3</SUB><SUP>−</SUP></UP> (NOx) or Glu levels by in vivo microdialysis before, during, and after infusion of NMDA in anesthetized rats. We also examined effects of Nomega -nitro-L-arginine methyl ester (L-NAME) on the changes in these levels. NMDA elicited depressor and bradycardic responses and increased the levels of NOx and Glu. L-NAME abolished the increases in the levels of NOx and Glu and attenuated cardiovascular responses evoked by NMDA. These results suggest that NMDA receptor activation in the NTS induces Glu release through NO synthesis and that Glu released via NO enhances depressor and bradycardic responses.

nitric oxide synthase; central nervous system; blood pressure; amino acid; microdialysis; nucleus tractus solitarii; N-methyl-D-aspartate


    INTRODUCTION
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES

IT HAS BEEN SUGGESTED THAT nitric oxide (NO) in the nucleus tractus solitarii (NTS) plays an important role in regulating blood pressure and sympathetic nerve activity (6, 10, 15, 19, 32). The NTS is the site of termination of primary afferent fibers arising from many cardiovascular receptors (5, 13). The excitatory amino acid L-glutamate (Glu) is considered the neurotransmitter of baroreceptor primary afferent fibers (5, 13, 28). In addition, neuronal NO synthase (nNOS)-positive neurons have been demonstrated in the NTS by NADPH diaphorase staining or immunohistochemistry for nNOS (15, 17, 34). We and others have previously demonstrated that microinjection of an inhibitor of NO synthase (NOS), NG-monomethyl-L-arginine (L-NMMA), in the NTS increases blood pressure and sympathetic nerve activity in anesthetized rabbits and rats (6, 10, 19, 32). On the other hand, administration of the substrate of NO, L-arginine, into the NTS has been reported to elicit significant decreases in blood pressure and sympathetic nerve activity in rats (19, 32). Furthermore, using single-unit extracellular recordings of NTS neurons in rat brain stem slices, we have shown that both L-arginine and an NO donor, sodium nitroprusside, increase neuronal activity in the NTS via activation of soluble guanylate cyclase (31). However, the mechanism(s) of NO release and the effect of NO on synaptic transmission in the NTS in vivo are not known. It has been shown using brain slice preparations that activation of Glu receptors, particularly those of the N-methyl-D-aspartate (NMDA) subtype, induces the synthesis of NO in cerebellum and hippocampus (3, 8, 33). It has been proposed that NO acts in an ultrashort loop feedback system, in which release of Glu results in activation of nNOS and production of NO (8). This NO in turn reaches presynaptic terminals where it influences subsequent release of Glu in response to neuronal activation. In addition, both NMDA and non-NMDA receptors in the NTS are known to be involved in processing of baroreceptor afferent information (27). Moreover, there are no studies examining both release of NO or Glu and cardiovascular responses evoked by NMDA receptor activation.

The aim of this study was thus to determine whether activation of NMDA receptors of neurons in the NTS releases NO and, if so, to determine whether NMDA-induced NO stimulates the release of Glu and whether this mechanism is involved in the depressor and bradycardic responses evoked by activation of NMDA receptors in the NTS in anesthetized rats. For this purpose, we employed in vivo microdialysis to assess the effects of NMDA receptor activation in the NTS on the release of NO and Glu with blood pressure and heart rate monitoring.


    METHODS
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES

General Procedures

Experiments were performed on adult male Wistar-Kyoto rats (300-350 g) anesthetized with pentobarbital sodium (50 mg/kg ip; followed by 10-20 mg · kg-1 · h-1 iv). The trachea was cannulated, and catheters were inserted in the femoral artery and vein for recording arterial blood pressure and the administration of drugs. Blood pressure was monitored throughout the experiments. When a stable and adequate level of anesthesia was achieved, the rat was placed in a stereotaxic frame, and the dorsal surface of the medulla was exposed. The rat was mechanically ventilated with room air supplemented with oxygen and then paralyzed with tubocurarine (0.25 mg · kg-1 · h-1 iv) to avoid cardiovascular effects secondary to blood gas changes. The adequacy of anesthesia without paralysis was verified by the absence of a withdrawal response to nociceptive stimulation of a hindpaw and during paralysis by a stable arterial blood pressure and heart rate. As shown in Fig. 1, a microdialysis probe (A-I-12-01, 1 mm length; Eicom, Kyoto, Japan) was inserted in the unilateral NTS (0.6 mm rostral and 0.6 mm lateral to calamus scriptorius and 1 mm below the dorsal surface of the medulla). At the end of experiments, rats were deeply anesthetized with an overdose of pentobarbital sodium (100 mg/kg), and intracardiac perfusion of saline followed by phosphate buffer solution containing 4% paraformaldehyde was carried out. The brain stem was removed, and 50-µm-thick coronal sections of the medulla were cut on a microtome. The location of the dialysis probe was histologically confirmed by cresyl violet staining after each experiment. This study was reviewed and approved by the Committee on Ethics of Animal Experiments, Faculty of Medicine, Kyushu University.


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  		Fig. 1.   Photomicrograph stained by neutral red (A) and an illustration of the position (arrow) and the size of the microdialysis probe in the nucleus tractus solitarii (NTS; B). IX, hypoglossal nucleus; DMV, dorsal motor nucleus of vagus; AP, area postrema; 4th, 4th ventricle.

Measurement of NOx and Glu Levels by In Vivo Microdialysis

The NTS was perfused with Ringer solution (140 mM NaCl, 4 mM KCl, 1.26 mM CaCl2, and 1.15 mM MgCl2, pH 7.4) at a constant flow rate of 2 µl/min through a microdialysis probe. To measure NO<UP><SUB>2</SUB><SUP>−</SUP></UP> and NO<UP><SUB>3</SUB><SUP>−</SUP></UP> (NOx) levels in the dialysates, the perfused dialysates were collected every 10 min in the sample loop of an automated sample injector connected to an automated NO detector-HPLC system (ENO-10; Eicom; see Refs. 35 and 36). NOx in the dialysate was separated by a reverse-phase separation column packed with polystyrene polymer (NO-PAK, 4.6 × 50 mm; Eicom). NO<UP><SUB>2</SUB><SUP>−</SUP></UP> was mixed with a Griess reagent to form a purple azo dye in a reaction coil. The separation and reduction columns and the reaction coil were placed in a column oven that was set at 30°C. The absorbance of the color of the product dye at 540 nm was measured by a flow-through spectrophotometer (NOD-10; Eicom). The mobile phase, which was delivered by a pump at a rate of 0.33 ml/min, was 10% methanol containing 0.15 M NaCl/NH4Cl and 0.5 g/l 4Na-EDTA. The Griess reagent, which was 1.25% HCl containing 5 g/l sulfanilamide with 0.25 g/l N-naphthylethylenediamine, was delivered at a rate of 0.1 ml/min (23). Glu level was measured by HPLC. The perfused dialysates were collected every 30 min. Collected samples were analyzed with an HPLC system with an electrochemical detector (ECD-300; Eicom; see Ref. 23). The mobile phase for Glu detection consisted of 75% 0.1 M PBS (pH 6.5) and 25% methanol.

Experimental Protocols

Protocol 1. We examined the effects of infusion of NMDA in the NTS on the level of NOx in the dialysates (n = 4 or 5). After confirming that the level of NOx in the dialysates was stable, 0.001, 0.01, 0.1, or 1 mM NMDA dissolved in Ringer solution was infused for 10 min through a dialysis probe. We chose the dose of 1 mM NMDA for subsequent experiments, since depressor and bradycardic responses were apparent with it. Use of this dose of NMDA has been reported for in vivo microdialysis (35, 36). We also examined the effects of intracisternal injection of Nomega -nitro-L-arginine methyl ester (L-NAME, 5 µmol), an inhibitor of NOS, on the level of NOx resulting from infusion of NMDA (1 mM) in the NTS.

Protocol 2. We examined the effects of infusion of NMDA (1 mM) in the NTS on the level of Glu in dialysates (n = 5). After confirmation that the level of Glu in the dialysates was stable, 1 mM NMDA dissolved in Ringer solution was infused for 30 min through a dialysis probe. We also examined the effects of intracisternal injection of L-NAME (5 µmol) on the level of Glu resulting from infusion of NMDA in the NTS (n = 5).

Protocol 3. We examined the effect of the NO donor, S-nitroso-N-acetylpenicillamine (SNAP, 10 mM), on the level of Glu in dialysates. The basal level of Glu was determined from measurements of two consecutive stable dialysate samples immediately before infusion of SNAP (n = 5). To confirm that the released Glu originated from nerve terminals by exocytosis, we performed the same experiments but with removal of Ca2+ from the infusion solution (n = 5).

Protocol 4. To confirm the effects of L-NAME in the NTS on the depressor and bradycardic responses evoked by NMDA, we examined the depressor and bradycardic responses evoked by microinjection of NMDA (40 pmol) in the NTS before and after microinjection of L-NAME in the NTS (n = 5).

Protocol 5. To rule out the possibility that blood pressure elevation caused by intracisternal injection of L-NAME might have affected our results, we examined the depressor and bradycardic effects of microinjection of NMDA (40 pmol) in the NTS before and during intravenous infusion of phenylephrine (2-3 µg · kg-1 · min-1) for ~10 min to achieve a level of blood pressure similar to that obtained with intracisternal L-NAME (n = 5).

Statistical Analysis

One-way ANOVA for repeated measures was used to compare baseline values. Two-way ANOVA followed by Bonferroni's multiple comparison was used to compare any two mean values. All values are expressed as means ± SE, and differences were considered significant at P < 0.05.


    RESULTS
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES

Effect of NMDA Receptor Activation in the NTS on Level of NOx in Dialysates

Figure 2A shows typical chromatograms exhibiting the effect of infusion of NMDA in the NTS on level of NOx in dialysates. Infusion of NMDA (0.001, 0.01, and 0.1 mM, n = 4 for each, or 1 mM, n = 5) increased dose dependently the level of NOx in dialysates (3.7 ± 1.4, 4.8 ± 2.3, 5.3 ± 1.0, and 7.8 ± 1.3 pmol/20 µl, respectively, P < 0.01 for each). Intracisternal injection of L-NAME attenuated the magnitude of increase in the level of NOx in dialysates evoked by infusion of NMDA (Fig. 2B).


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  		Fig. 2.   A: typical chromatograms showing the effect of infusion of N-methyl-D-aspartate (NMDA) in the NTS on level of NO<UP><SUB>2</SUB><SUP>−</SUP></UP> and NO<UP><SUB>3</SUB><SUP>−</SUP></UP> (NOx) in dialysate. B: effect of NMDA (1 mM) on level of NOx in dialysate (n = 5). Basal level was determined from measurements of two consecutive stable dialysate samples obtained immediately before infusion of NMDA. Values are expressed as means ± SE. *P < 0.05 and **P < 0.01 vs. basal level. dagger P < 0.01 vs. before Nomega -nitro-L-arginine methyl ester (L-NAME).

Effect of NMDA Receptor Activation in the NTS on Level of Glu in Dialysates

Infusion of NMDA in the NTS increased the level of Glu in dialysates (3.4 ± 0.5 to 5.3 ± 0.5 pmol/15 µl, P < 0.01; Fig. 3). Intracisternal injection of L-NAME abolished the increase in the level of Glu in dialysates evoked by NMDA infusion (Fig. 3).


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  		Fig. 3.   A: typical chromatograms showing the effect of infusion of NMDA on level of L-glutamate (Glu) in dialysate. B: effect of NMDA (1 mM) on level of Glu in dialysate (n = 5 for each). Basal level was determined from measurements of two consecutive stable dialysate samples obtained immediately before infusion of NMDA. Values are expressed as means ± SE of percentages of basal levels. **P < 0.01 vs. basal level. dagger P < 0.01 vs. control.

Effect of NO Donor in the NTS on Level of Glu in Dialysates With or Without Ca2+

Infusion of SNAP in the NTS increased the level of Glu in dialysates (Fig. 4A). However, deprivation of Ca2+ from Ringer solution significantly attenuated SNAP-induced Glu release (P < 0.05; Fig. 4B).


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  		Fig. 4.   Effect of S-nitroso-N-acetylpenicillamine (SNAP, 10 mM) on level of Glu in dialysate. A: SNAP infused with normal Ringer solution (n = 5). B: SNAP infused with Ca2+-free Ringer solution (n = 5). Basal level was determined from measurements of two consecutive stable dialysate samples obtained immediately before infusion of SNAP. Values are expressed as means ± SE. *P < 0.05 vs. basal level. dagger P < 0.05 vs. control.

Effects of NMDA Receptor Activation in the NTS on Blood Pressure and Heart Rate

Infusion of NMDA in the NTS decreased blood pressure and heart rate, as shown in Fig. 5A. Prior intracisternal injection of L-NAME increased baseline blood pressure (106 ± 4 to 137 ± 3 mmHg, P < 0.01) and tended to decrease heart rate (254 ± 10 to 228 ± 14 beats/min, P = 0.17) and also attenuated the depressor and bradycardic responses evoked by NMDA (Fig. 5B). In addition, microinjection of NMDA in the NTS resulted in depressor and bradycardic responses (-55 ± 4 mmHg and -70 ± 13 beats/min, respectively). These effects were significantly attenuated by injection of L-NAME directly in the NTS (-42 ± 4 mmHg and -44 ± 10 beats/min, respectively, P < 0.05). Continuous intravenous infusion of phenylephrine increased blood pressure (from 108 ± 3 to 135 ± 5 mmHg). The magnitude of decreases in blood pressure and heart rate evoked by microinjection of NMDA in the NTS during infusion of phenylephrine did not differ from those before infusion of phenylephrine [39 ± 6 vs. 45 ± 3 mmHg, 53 ± 18 vs. 54 ± 16 beats/min, respectively, n = 5, not significant (NS)].


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  		Fig. 5.   A: example of the effect of L-NAME on the depressor and bradycardic responses evoked by infusion of NMDA in the NTS. AP, arterial pressure; HR, heart rate; bpm, beats/min. B: effects of stimulation of NMDA receptors within the NTS before and after intracisternal injection of L-NAME on depressor and bradycardic responses (n = 5). Values are expressed as means ± SE. *P < 0.05 and **P < 0.01 vs. control.


    DISCUSSION
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES

The major findings of this study are that 1) NMDA receptor activation in the NTS increased the levels of NOx and Glu, and both of these increases were attenuated by NOS blockade, and 2) the depressor and bradycardic responses evoked by infusion of NMDA in the NTS were attenuated by NOS blockade. These results suggest that NMDA receptor activation in the NTS induces NO release, which in turn facilitates release of Glu from presynaptic terminals and thus augments the depressor and bradycardic effects of NMDA receptor activation in anesthetized rats. This is the first study that demonstrates that facilitatory release of Glu via NO production evoked by NMDA in the NTS contributes to the depressor and bradycardic responses in vivo to measure levels of NOx or Glu, by in vivo microdialysis, and blood pressure and heart rate.

In this study, we demonstrated that NO production occurs after NMDA receptor activation in the NTS using in vivo brain microdialysis in anesthetized rats. NO production in the brain has been measured by other laboratories using this technique (20, 35, 36). In the present study, the NO production caused by NMDA receptor activation was markedly attenuated by prior intracisternal injection of L-NAME, an NOS inhibitor, suggesting that the increase in NOx level after NMDA receptor activation is caused by activation of NOS in the NTS. In the preliminary experiments, we repeated NMDA infusion at this dose (1 mM) to measure the NOx level and found the same levels of increases in NOx and the depressor responses evoked by NMDA (changes in NOx levels: 11.4 ± 2.8 vs. 9.4 ± 3.4 pmol/20 µl, changes in mean blood pressure: -53 ± 3 vs. -51 ± 9 mmHg, respectively; n = 3, NS). NMDA receptor activation in the NTS also increased the level of extracellular Glu, as measured by in vivo microdialysis. This effect was also blocked by prior intracisternal injection of L-NAME, suggesting that the increase in production of Glu evoked by NMDA receptor activation in the NTS is mediated by NOS activation. Furthermore, administration of the NO donor SNAP in the NTS increased the level of Glu, and this increase was markedly attenuated when the NTS was perfused with Ca2+-free solution, suggesting that Glu is released from presynaptic terminals. Laurence and Jarrot (14), using in vivo microdialysis in anesthetized rats, also demonstrated that SNAP increases Glu and that this increase is blocked by methylene blue, suggesting that NO may increase Glu through guanylate cyclase in the NTS; however, they did not measure blood pressure and heart rate. In addition, they showed that the increase in Glu level evoked by high-K+ stimulation was abolished by removal of Ca2+ from the perfusate (14).

It has been suggested that retrograde signaling from postsynaptic cells controls presynaptic transmitter release in the hippocampus, the cerebellum, and the cerebral cortex (8, 25, 30, 33). However, because this type of retrograde signaling system has been demonstrated mainly in in vitro studies in regions of the brain other than the brain stem, it is unknown whether this system is also involved in central cardiovascular regulation. Our results are consistent with those of our previous studies and others performed in vivo, suggesting that NO within the NTS decreases blood pressure, heart rate, and sympathetic nerve activity (6, 10, 19, 32). Our results also support the hypothesis that NO is a retrograde messenger influencing blood pressure regulation in the NTS. Activation of NMDA receptors induces the synthesis of NO, which activates soluble guanylate cyclase and leads to the production of cGMP (8, 33). This phenomenon has also been confirmed by use of the specific inhibitor of soluble guanylate cyclase, 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (1, 2). Linkage between NMDA receptor activation and the NO system has thus been emphasized in previous studies (7, 24, 33). The excitatory amino acid Glu, which activates NMDA receptors, is considered a potential neurotransmitter of baroreceptor information in the rat NTS (5, 13, 27). Our results demonstrated a positive feedback system as release of Glu through NO production evoked by NMDA receptor activation. However, NOx or Glu levels returned to control levels rapidly after discontinuing infusion of NMDA. This is consistent with the results of the study examining NO release in the cerebellum by in vivo microdialysis (35). However, feedback loops are usually negative in nature. It has been suggested that attenuation of Glu effects by NO is mediated by inactivation of NMDA receptors (18, 21, 37) and also by NO-induced inhibition of protein kinase C activity (9). Thus we speculate that these mechanisms are responsible for stopping overproduction of Glu by NO. Furthermore, we did not measure levels of other amino acids in this study. It is thus possible that an inhibitory amino acid, such as GABA, also plays a role in mediating the effect of NO in the NTS. However, we were unable to detect GABA in the NTS evoked by infusion of NMDA.

Depressor and bradycardic responses evoked by NMDA receptor activation in the NTS were attenuated by prior intracisternal injection of L-NAME. These results suggest that the augmentation of Glu release by NO production evoked by NMDA receptor activation in the NTS contributes to the depressor and bradycardic effects of NMDA receptor activation. In previous studies of anesthetized animals, administration of NO in the NTS decreased blood pressure (6, 10, 19, 32), although the opposite result (22) and no effect of NO on blood pressure have also been reported (26). However, these studies were performed with the microinjection method, and physiological parameters such as blood pressure, heart rate, and/or sympathetic nerve activity were measured in the anesthetized state. In our study, to further explore the mechanisms involved in the effects of NO in the NTS on cardiovascular responses, we measured NOx or Glu simultaneously with blood pressure and heart rate monitoring. Di Paola et al. (6) showed that microinjection of Glu in the NTS decreased blood pressure; they showed that this decrease was abolished by prior injection of the selective NMDA receptor antagonist 2-amino-7-phosphonoheptanoic acid and reduced in magnitude by prior injection of methylene blue and L-NMMA in the NTS (6). Although several studies have demonstrated that NO in the NTS causes hypotension and bradycardia (6, 10, 19, 32), only one study has shown that the depressor and bradycardic responses evoked by direct application of NMDA in the NTS are attenuated after L-NMMA (19). They also found that prior administration of the NMDA receptor antagonist MK-801 attenuated the depressor and bradycardic effects of L-arginine (19). However, they did not measure either NOx or Glu in that study (19). Therefore, the mechanism(s) involved in the attenuated hypotensive and bradycardic responses evoked by NMDA is not known from the results of their study. We also confirmed attenuation of the depressor and bradycardic responses evoked by NMDA receptor activation after microinjection of L-NAME in the NTS. More importantly, we have shown that facilitatory release of Glu via NO production evoked by NMDA receptor stimulation in the NTS enhances hypotension and bradycardia in vivo by simultaneous measurement of NOx or Glu with blood pressure and heart rate. It has been shown using in vivo microdialysis that intravenous infusion of phenylephrine, which elevates blood pressure, increases Glu in the NTS (15, 28). These results suggest that Glu is released in the NTS with activation of baroreceptors and support the hypothesis that Glu is a neurotransmitter of baroreceptor afferents terminating in the NTS. Furthermore, the facilitatory effect of NO on the release of Glu in the NTS might be related to the attenuated central adaptation baroreflex control of sympathetic nerve activity that we described previously (11).

We must consider the possibility that intracisternal injection of L-NAME influenced brain stem regions other than the NTS, such as the ventrolateral medulla (12, 32), and thereby affected our results. However, this seems unlikely, since the depressor and bradycardic effects of NMDA receptor activation in the NTS were similar before and during the blood pressure elevation caused by intravenous infusion of phenylephrine. We also confirmed the attenuation of cardiovascular responses to NMDA after microinjection of L-NAME directly in the NTS. We did not perfuse the NTS with L-NAME from the dialysis probe because this procedure interfered with measurement of Glu or NOx in preliminary experiments. This difficulty was also described in another study (35).

We also must consider the possibility that differences in baseline blood pressure influenced our results. However, this also seems unlikely, since the depressor and bradycardic effects of microinjection of NMDA in the NTS did not differ before and during intravenous infusion of phenylephrine, which caused a pressor response similar to that of intracisternal L-NAME.

We used L-NAME as an NOS inhibitor in this study. L-NAME might have nonspecific effects unrelated to NOS blockade (4). This possibility is unlikely, however, since it has been shown that prior administration of NOS inhibitors L-NMMA or L-NAME significantly attenuated the cardiovascular effects of microinjection of Glu and NMDA in the NTS (19). We also demonstrated that intracisternal injection of L-NAME, but not of D-NAME, was useful for examining rapid central baroreflex control of renal sympathetic nerve activity (11). Although L-NAME is not selective for nNOS, even intraperitoneal injection of L-NAME has been shown by in vivo microdialysis to block the increase in the level of NOx in the cerebellum (35, 36).

It has recently been shown that systemic administration of lipopolysaccharide induces release of NO via activation of inducible NOS (iNOS) in the NTS in anesthetized rats (16). It is thus possible that iNOS was induced in our study. However, this seems unlikely, since in another study NO production occurred 3-4 h after administration of lipopolysaccharide (16). In contrast, in our study, we found immediate NO production in the NTS evoked by NMDA receptor activation. We therefore believe nNOS was responsible for the NO production we observed.

In summary, our results strongly suggest that NMDA receptor activation of neurons in the NTS induces Glu release from nerve terminals through NO synthesis and that the Glu released via NO synthesis enhances depressor and bradycardic responses in vivo.

Perspectives

NMDA receptors have been demonstrated in the rat NTS in autoradiographic and electrophysiological studies (27, 29). In fact, we found that NMDA has marked depressor and bradycardic effects. However, non-NMDA receptors in the NTS are also known to play important roles in baroreflex control of sympathetic nerve activity. While we recognize the importance of non-NMDA receptors in the NTS, we did not examine the effects of non-NMDA receptor activation on NO production. Further studies are needed to examine the role of NO in activation of non-NMDA receptors in the NTS.

We could not determine in our study whether the neurons activated by NMDA received input directly from arterial baroreceptors. It is possible that they received inputs from chemoreceptors and/or other visceral receptors (5, 13). It would be interesting to examine whether facilitatory Glu release by NO occurs after electrical stimulation of depressor afferent nerves.


    ACKNOWLEDGEMENTS

This study was supported by grants-in-aid for scientific research from the Ministry of Education, Science, and Culture of Japan (no. 09770489) and by a grant for research on the autonomic nervous system and hypertension from Kimura Memorial Heart Foundation/Pfizer Pharmaceuticals.


    FOOTNOTES

This study was presented in part at the 71st Scientific Sessions of the American Heart Association, Dallas, Texas, November 8-11, 1998.

Address for reprint requests and other correspondence: Y. Hirooka, Dept. of Cardiovascular Medicine, Cardiovascular Science, Graduate School of Medical Sciences, Kyushu Univ., 3-1-1 Maidashi, Higashi-ku, Fukuoka 812-8582, Japan (E-mail: hyoshi{at}cardiol.med.kyushu-u.ac.jp).

The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

Received 15 March 2000; accepted in final form 19 December 2000.


    REFERENCES
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES

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Am J Physiol Regul Integr Comp Physiol 280(5):R1285-R1291
0363-6119/01 $5.00 Copyright © 2001 the American Physiological Society


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