- Email: [email protected]
Role of nitric oxide in the nucleus of the solitary tract of rats
Brain Research 798 Ž1998. 232–238
Research report
Role of nitric oxide in the nucleus of the solitary tract of rats Kiyoshi Matsumura ) , Takuya Tsuchihashi, Shuntaro Kagiyama, Isao Abe, Masatoshi Fujishima Second Department of Internal Medicine, Faculty of Medicine, Kyushu UniÕersity, Maidashi 3-1-1, Higashi-ku, Fukuoka 812-8582, Japan Accepted 14 April 1998
Abstract We have determined the role of nitric oxide ŽNO. in the nucleus of the solitary tract ŽNTS. of normotensive Wistar rats. The unilateral microinjection of N v-nitro-L-arginine methyl ester Ž10 nmol. to block the synthesis of NO into the NTS significantly decreased the arterial pressure, heart rate ŽHR. and renal sympathetic nerve activity ŽRSNA. Žy19 " 2 mmHg, y23 " 5 beatsrmin, y30 " 2%, respectively.. The microinjection of carboxy-2-phenyl-4,4,5,5-tetramethylimidazoline-1-oxyl 3-oxide ŽCarboxy PTIO. Žtrapper of NO; 0.1 nmol. into the NTS also decreased arterial pressure and RSNA. Conversely, the microinjection of Et 2 NwNŽO.NOxNa ŽNOC 18. ŽNO donor; 10 nmol. caused increases in arterial pressure, HR and RSNA Žq14 " 2 mmHg, q11 " 2 beatsrmin, q38 " 7%, respectively., which was inhibited by the pre-microinjection of Carboxy PTIO Ž0.1 nmol.. On the other hand, not only L-arginine Ž10 nmol. but also D-arginine Ž10 nmol., which is inactive to produce NO, significantly decreased the arterial pressure and RSNA. These results suggest that Ž1. NO acts at the NTS to increase the arterial pressure and RSNA, and Ž2. the microinjection of L-arginine as well as D-arginine led to decreases in arterial pressure and RSNA that were not mediated by the formation of NO in the NTS. q 1998 Elsevier Science B.V. All rights reserved. Keywords: Central nervous system; Microinjection; Renal sympathetic nerve activity; Sympathetic nervous system
1. Introduction Nitric oxide ŽNO., which is responsible for endothelium-dependent vasodilatation, is synthesized from Larginine in the presence of its synthetic enzyme NO synthase ŽNOS. w16x. Immunocytochemical studies have shown that high concentrations of NOS are also present in specific regions of the brain, including the sites involved in the regulation of arterial pressure and the baroreceptor reflex, such as the nucleus of the solitary tract ŽNTS. and the ventrolateral medulla ŽVLM. w8,14,23x. An intramedullary NOS neural pathway that projects from the NTS to the rostral nucleus ambiguus has also been described w3x. Furthermore, various lines of evidence suggest that NO participates in cardiovascular regulation not only by a direct effect on vascular smooth muscle but also via its action on the central nervous system w5,18–21x. The NO is now considered to be a neurotransmitter or a neuromodulator in the central nervous system w4,5x. The microinjection of N G -monomethyl-L-arginine ŽL-NMMA. into the NTS to block endogenous NO caused a decrease
)
Corresponding author. Fax: q 81-92-642-5271; [email protected]
E-mail:
0006-8993r98r$19.00 q 1998 Elsevier Science B.V. All rights reserved. PII S 0 0 0 6 - 8 9 9 3 Ž 9 8 . 0 0 4 2 0 - X
in arterial pressure at a low dose and an increase in arterial pressure at a high dose w21x. It seems to be controversial whether the blockade of endogenous NO in the NTS elicits a pressor response. On the other hand, the microinjection of L-arginine, the NO precursor, into the NTS reduces the arterial pressure w21x. However, L-arginine has been reported to have a non-specific central action w13x. The intracerebroventricular injection of L-arginine causes a transient increase in arterial pressure that is not mediated by a central NO pathway w13x. Furthermore, cardiovascular effect of NO in the NTS remains controversial w6,10,15,21x. We hypothesized that L-arginine may not be an appropriate tool for the examining the central action of NO, even though NO is synthesized from it. The NO donor, Et 2 NwNŽO.NOxNa ŽNOC 18., has been used to study the central effect of NO. It has been reported that the intracerebroventricular injection of NOC 18 led to a decrease in arterial pressure in spontaneously hypertensive rats without a transient increase in arterial pressure w2x. We considered that NOC 18 might be more appropriate than L-arginine for evaluating the central action of NO. Thus, we conducted the present study to re-evaluate the role of endogenous and exogenous NO in the NTS on the cardiovascular and sympathetic responses in anesthetized rats using NOC 18, as well as L-arginine.
K. Matsumura et al.r Brain Research 798 (1998) 232–238
2. Materials and methods 2.1. Preparation of animals Experiments were done in adult male Wistar rats Ž10–12 weeks old.. These experiments were conducted according to the institutional guidelines of Kyushu University on animal experimentation. Rats were anesthetized with urethane Ž1.2 grkg, i.p... Femoral artery and vein were cannulated for the measurement of arterial pressure and the injection of drugs, respectively. The trachea was cannulated to facilitate ventilation. Arterial blood gases and pH were maintained within the physiological range ŽpH 7.39 to 7.42; PO 2 95 to 105 mmHg; PCO 2 34 to 38 mmHg.. Body temperature was maintained at 37.0 " 0.58C by an external heating source. Depth of anesthesia throughout the experiment was monitored by a variety of indexes, including corneal reflexes, as well as respiratory rhythm, arterial pressure, and heart rate ŽHR. stability.
233
An anesthetized rat was placed in a stereotaxic frame ŽDavid Kopf Instruments, Tujunga, CA. with the head fixed downward at 458. After making a midline incision through the skin, the dorsal neck muscles were retracted with sutures to visualize the foramen magnum. The medulla oblongata was exposed by incising the atlanto-occipital membrane and the dorsal surface was kept moist either by artificial cerebrospinal fluid ŽaCSF; pH 7.4. or by production of endogenous CSF. Renal sympathetic nerve activity ŽRSNA. was recorded as described previously w11,12x. Briefly, the left kidney was exposed retroperitoneally, and a branch of the renal nerve was separated from the renal plexus and the surrounding connective tissues with the aid of a dissecting microscope. The RSNA was recorded by a pair of electrodes made from Teflon-insulated seven-stranded steel wire Ž0.001 in. diameter, A-M Systems, Carlsborg, WA.. The area of the nerve and wire interface was embedded in silicone cement ŽElastosil RT 604A and B cement, Wacker Chemicals, Munchen, Germany. to prevent drying of the ¨
Fig. 1. Representative original recordings of arterial pressure, mean arterial pressure, heart rate, renal sympathetic nerve activity ŽRSNA. and integrated RSNA elicited by the microinjection of 10 nmol of N v-nitro-L-arginine methyl ester ŽL-NAME. into the nucleus of the solitary tract.
234
K. Matsumura et al.r Brain Research 798 (1998) 232–238
nerve and to minimize movement artifacts associated with respiration. 2.2. Recording procedures of RSNA The RSNA was amplified Žmodel DPA-100E, Dia Medical System, Tokyo, Japan. and filtered Ž100–3000 Hz., and the waveforms were integrated after a full-wave rectification using an integrator amplifier Žmodel 1322, NEC San-ei, Tokyo, Japan. with the sample-hold function reset to baseline by an internal timer set at 1 s. The residual integrated RSNA which existed after intravenous administration of hexamethonium bromide Ž30 mgrkg, i.v.. was taken as the noise level associated with nerve recording. This value was subtracted from absolute values of integrated RSNA before performing further data analysis. 2.3. Microinjection procedures
2.5. Experimental protocols 2.5.1. Effect of L-NAME in the NTS N v-nitro-L-arginine methyl ester ŽL-NAME; 10 nmol. ŽSigma, St. Louis, MO. was used to block the production of endogenous NO, and was microinjected into the NTS of the rats. As the vehicle control, we administered a microinjection of aCSF Ž50 nl.. 2.6. Effects of NOC 18 and Carboxy PTIO in the NTS The NOC 18 ŽDojindo Institute, Kumamoto, Japan., 1 or 10 nmol, was microinjected into the NTS. In another group of rats, carboxy-2-phenyl-4,4,5,5-tetramethylimidazoline-1-oxyl 3-oxide ŽCarboxy PTIO; 0.1 nmol. ŽDojindo Institute, Kumamoto, Japan., an NO trapper w1x, was microinjected into the NTS 2 min prior to the microinjection of 10 nmol of NOC 18. As the vehicle control, we administered a microinjection of aCSF Ž50 nl..
Microinjections were made from multibarrel micropipettes with tip diameters of 20–50 m m. The pipettes were made from calibrated microbore capillary glass tubing ŽAccu-Fill 90 w , Clay Adams, NJ.. Tips were drawn on a micropipette puller Žmodel PE-2, Narishige Scientific Instruments, Tokyo, Japan.. Injections Ž50 nl. were made over a 30-s period with hand-held syringe as described elsewhere w22x. The injection volume was measured by observing the movement of the fluid meniscus along a reticule in a microscope. The appropriate placement of the pipette tip within the medial NTS was established by microinjection of L-glutamate Ž2 nmol. and observing a depressor response of at least 25 mmHg. On this basis, injections into the medial NTS had coordinates 0.4–0.5 mm rostral and 0.5–0.6 mm lateral to calamus scriptorius and 0.4 mm below the dorsal surface of medulla. All drugs for microinjection were dissolved in aCSF Žin mmolrl: 133.3 NaCl, 3.4 KCl, 1.3 CaCl 2 , 1.2 MgCl 2 , 0.6 NaH 2 PO4 , 32.0 NaHCO 3 , and 3.4 glucose.. Alcian blue dye Ž50 nl. was injected from a separate barrel of the pipette to mark the site of injection at the end of the experiment. 2.4. Histological analysis At the completion of the experiment, the rats were prepared for transcardial perfusion and then euthanatized with a lethal dose of intravenous sodium pentobarbital Ž75 mgrkg.. Animals were then perfused transcardially with 150 ml 0.9% NaCl followed by 150 ml 10% phosphatebuffered formaldehyde solution. The brain was removed and stored in 10% phosphate-buffered formaldehyde solution. The medulla oblongata was cut into 50 m m serial coronal frozen sections that were stained with neutral red. The location of the microinjection sites in the medial NTS was confirmed microscopically according to the atlas of Paxinos and Watson w17x.
Fig. 2. Bar graphs showing the effects of the microinjection of 10 nmol of N v-nitro-L-arginine methyl ester ŽL-NAME. and vehicle Žartificial cerebrospinal fluid; 50 nl. into the nucleus of the solitary tract on changes in mean arterial pressure ŽMAP., heart rate ŽHR. and integrated renal sympathetic nerve activity ŽRSNA. in Wistar rats. Values are means"S.E. Ž)). p- 0.01 as compared with the respective responses to vehicle by Student’s t-test. Number of rats appears in parentheses.
K. Matsumura et al.r Brain Research 798 (1998) 232–238
235
the microinjection of vehicle ŽSAS Institute, Cary, NC.. A value of p - 0.05 was considered statistically significant.
3. Results 3.1. Effect of L-NAME in the NTS Baseline levels of mean arterial pressure ŽMAP. and HR were 96.0 " 1.2 mmHg and 386.9 " 14.2 beatsrmin in L-NAME-microinjected rats, and 93.7 " 3.1 mmHg and 410.7 " 17.2 beatsrmin in aCSF-microinjected rats, respectively. Fig. 1 shows the typical responses of MAP, HR and RSNA that were elicited by the microinjection of L-NAME into the NTS. The microinjection of L-NAME into the NTS caused significant decreases in MAP, HR and RSNA as compared with the effects of aCSF ŽFig. 2..
Fig. 3. Representative original recordings of arterial pressure, mean arterial pressure, heart rate, renal sympathetic nerve activity ŽRSNA. and integrated RSNA elicited by the microinjection of 10 nmol of Et 2 NwNŽO.NOxNa ŽNOC 18. into the nucleus of the solitary tract.
2.7. Effects of L-arginine and D-arginine in the NTS L-Arginine
ŽSigma. or D-arginine ŽSigma., 10 nmol, was microinjected into the NTS of rats. As the vehicle control, we administered a microinjection of aCSF Ž50 nl.. 2.8. Statistics All values are expressed as means " S.E. Student’s t-test was used to determine the effect of the microinjection of L-NAME into the NTS on cardiovascular and sympathetic responses. A one-way analysis of variance was used to evaluate the effects of the microinjection of NOC 18, Carboxy PTIO, L-arginine and D-arginine into the NTS, followed by Duncan’s multiple range test to evaluate the significance of the mean differences in the response to
Fig. 4. Bar graphs showing the effects of Et 2 NwNŽO.NOxNa ŽNOC 18; 1 and 10 nmol., NOC 18 administered following pretreatment with carboxy-2-phenyl-4,4,5,5-tetramethylimidazoline-1-oxyl 3-oxide ŽCarboxy PTIO; 0.1 nmol., and vehicle Žartificial cerebrospinal fluid; 50 nl. microinjected into the nucleus of the solitary tract on changes in mean arterial pressure ŽMAP., heart rate ŽHR. and integrated renal sympathetic nerve activity ŽRSNA. in Wistar rats. Values are means"S.E. Ž)). p- 0.01 as compared with the respective responses to vehicle by Duncan’s multiple range test. Number of rats appears in parentheses.
236
K. Matsumura et al.r Brain Research 798 (1998) 232–238
3.2. Effects of NOC 18 and Carboxy PTIO in the NTS Fig. 3 shows the typical responses of MAP, HR and RSNA that were elicited by the microinjection of NOC 18 Ž10 nmol. into the NTS. The microinjection of NOC 18 Ž1 and 10 nmol. into the NTS caused dose-related increases in the MAP, HR and RSNA ŽFig. 4.. Although the microinjection of Carboxy PTIO into the NTS caused significant decreases in MAP and RSNA Žy9.2 mmHg and y20.1%, respectively., MAP and RSNA returned to the pre-microinjection level within 2 min. Pretreatment with Carboxy PTIO prevented the cardiovascular and RSNA responses to the microinjection of 10 nmol NOC 18 into the NTS ŽFig. 4.. 3.3. Effects of L-arginine and D-arginine in the NTS Fig. 5 illustrates the effects of L-arginine and D-arginine in the NTS. D-Arginine, as well as L-arginine, caused
Fig. 5. Bar graphs showing the effects of L-arginine Ž10 nmol., D-arginine Ž10 nmol., and vehicle Žartificial cerebrospinal fluid; 50 nl. microinjected into the nucleus of the solitary tract on changes in mean arterial pressure ŽMAP., heart rate ŽHR. and integrated renal sympathetic nerve activity ŽRSNA. in Wistar rats. Values are means"S.E. Ž). p- 0.05, Ž)). p- 0.01 as compared with the respective responses to vehicle by Duncan’s multiple range test. Number of rats appears in parentheses.
significant decreases in MAP, HR and RSNA as compared with the effects of the microinjection of aCSF.
4. Discussion We made two principal findings. First, we showed that endogenous, as well as exogenous NO, modulated the sympathetic outflow and arterial pressure in the NTS. The microinjection of NOC 18, an NO donor, into the NTS caused dose-related increases in MAP, HR and RSNA that were prevented by pre-microinjection of Carboxy PTIO, trapper of NO, into the NTS. These results suggest that the cardiovascular and sympathetic responses induced by the microinjection of NOC 18 are mediated by an NO pathway in the NTS. Our second finding was that the microinjection of L-arginine into the NTS caused a decrease in sympathetic outflow, and reduced the arterial pressure and HR. Since the microinjection of D-arginine into the NTS also decreased the MAP, HR and RSNA, cardiovascular and sympathetic responses elicited by the microinjection of L-arginine were considered not to be mediated by an NO pathway. The role of NO in the central nervous system has been examined extensively. The NOS and neuronal NADPH diaphorase in the brain have been shown to be identical; many neurons with NADPH diaphorase activity are located in the NTS and the VLM w7,8,14,23x which are involved in the regulation of the cardiovascular system and the baroreceptor reflex. The NTS is important in regulating sympathetic outflow. However, only a few studies have evaluated the central effects of NO on cardiovascular and sympathetic responses with the use of direct recording of sympathetic nerve activity w6,7,21x. The precise role of the sympathetic nervous system in this respect has not been determined. Furthermore, the effects of NO in the NTS on cardiovascular and sympathetic responses have been controversial. Harada et al. w6x demonstrated that the microinjection of L-NMMA into four sites of the NTS of rabbits to block endogenous NO produced small increases in the arterial pressure and RSNA; these responses were inhibited by pretreatment with a microinjection of L-arginine Ž16 nmol per site.. However, the microinjection of L-arginine into the NTS per se did not cause any cardiovascular or RSNA responses. On the other hand, Tseng et al. w21x reported that the microinjection of L-NMMA into the NTS increased the arterial pressure while it decreased the HR in rats. However, in that study, the microinjection of a low dose of L-NMMA Ž10 nmol. led to a decrease in the arterial pressure. A pressor response was observed only when a high dose of L-NMMA Ž33 or 66 nmol. was microinjected into the NTS; the increase in MAP was less than 10 mmHg. In contrast, the HR showed a decrease at either a low or high dose of L-NMMA. Our present results are consistent with the findings of Tseng et al. w21x regard-
K. Matsumura et al.r Brain Research 798 (1998) 232–238
ing the depressor effect of a microinjection of L-arginine into the NTS, but not with those of Harada et al. w6x. Considering the results of Harada et al. w6x, Tseng et al. w21x and those of the present study, the possible explanations for the differing responses of the arterial pressure are as follows. First, the role of NO in the NTS may differ according to species Žrats vs. rabbits.. Second, the pressor response elicited by the microinjection of a high dose of L-NMMA may be due to the leakage of L-NMMA into the systemic circulation. This could constrict the peripheral arteries and lead to an increase in arterial pressure with a decrease in HR. The occurrence of a bradycardic response, in association with the increase in arterial pressure, also suggests the involvement of a peripheral pressor mechanism that decreases the HR via the baroreceptor reflex. The differing effects of NO on the central nervous system of rats and rabbits may also be supported by studies of the microinjection of L-NAME or L-NMMA into the rostral VLM. Hirooka et al. w7x reported that the microinjection of L-NAME into the rostral VLM of rabbits reduced the arterial pressure and RSNA; conversely, Kagiyama et al. w9x and Tseng et al. w21x demonstrated that microinjections of L-NAME or L-NMMA into the rostral VLM of rats increased the arterial pressure. These divergent responses might be attributable to the species studied. A differing distribution or density of NOS in the brain has been shown in rats and rabbits w7x, and may account for such responses. In contrast to the effects of the NO donor agent ŽNOC 18., the microinjection of the NO precursor, L-arginine, into the NTS decreased the arterial pressure, as well as the HR and RSNA. D-Arginine, an inactive isomer of Larginine that produces NO, also decreased the arterial pressure, HR and RSNA, suggesting that the depressor and sympathoinhibitory responses to L-arginine are not mediated by a central NO pathway. A central cardiovascular effect of L-arginine that was independent of an NO pathway was also observed by Nishimura et al. w13x and Hirooka et al. w7x. The intracerebroventricular injection of L-arginine transiently increased the arterial pressure related to the activation of the central renin–angiotensin system w13x. Moreover, microinjections of L-arginine or D-arginine into the rostral VLM of rabbits decreased the arterial pressure and RSNA, suggesting that these responses were non-specific w7x. Recently, Ohta et al. w15x reported that S-nitrosocysteine, an NO donor, acts at the NTS to reduce the arterial pressure. Its action was independent of the release of NO from nitrosothiol, but may have been mediated via stereoselective sites on target neurons. While those authors did not determine the precise mechanism of the depressor action, their findings appear to be consistent with our present observation that L-arginine acts at the NTS independent of the release of NO. How NO regulates the blood pressure and the sympathetic outflow in the brain is not known. However, NO has been proposed to act via two models w4,5x. In the first
237
model, NO is generated postsynaptically as a result of a rise in cytosolic Ca2q concentration that follows the activation of N-methyl-D-aspartic acid receptors. It then diffuses out and acts on neighbouring neuronal structures, including the presynaptic terminals and glial cells. In the second model, NO is generated presynaptically following the action potential-dependent influx of Ca2q ions. A previous study from our laboratory demonstrated that the microinjection of NOC 18 into the rostral VLM decreases the arterial pressure, whereas the microinjection of LNAME into the rostral VLM increases it w9x. Previous studies of rostral VLM, as well as the present evaluation of NTS, suggest that NO acts on medulla oblongata of rats to inhibit neuronal transmission and to modulate sympathetic outflow. The central regulations of the cardiovascular system and of the sympathetic nervous system may differ between rats and rabbits.
5. Conclusion We have demonstrated that NO acts at the NTS to increase the sympathetic outflow in normotensive rats, leading to increases in arterial pressure and HR. Conversely, the blockade of endogenous NO in the NTS suppresses this sympathetic outflow. Endogenous NO, as well as exogenous NO, modulates sympathetic outflow so as to regulate the arterial pressure in the NTS. The physiological implications await further investigation.
Acknowledgements We thank Miss Hideko Noguchi for expert technical assistance. This work was supported in part by a grant from the Ministry of Education, Japan ŽNo. 09770495. and a Grant for Research on Sympathetic Nervous System and Hypertension.
References w1x T. Akaike, M. Yoshida, Y. Miyamoto, K. Sato, M. Kohno, K. Sasamoto, K. Miyazaki, S. Ueda, H. Maeda, Antagonistic action of imidazolineoxyl N-oxides against endothelium-derived relaxing factorr Ø NO through a radical reaction, Biochemistry 32 Ž1993. 827– 832. w2x C.L. Cabrera, S.L. Bealer, D.F. Bohr, Central depressor action of nitric oxide is deficient in genetic hypertension, Am. J. Hypertens. 9 Ž1996. 237–241. w3x W.-P. Gai, J.P. Messenger, Y.-H. Yu, Z.J. Gieroba, W.W. Blessing, Nitric oxide-synthesising neurons in the central subnucleus of the nucleus tractus solitarius provide a major innervation of the rostral nucleus ambiguus in the rabbit, J. Comp. Neurol. 357 Ž1995. 348–361. w4x J. Garthwaite, Glutamate, nitric oxide and cell–cell signalling in the nervous system, Trends Neurosci. 14 Ž1991. 60–67. w5x J. Garthwaite, C.L. Boulton, Nitric oxide signalling in the central nervous system, Annu. Rev. Physiol. 57 Ž1995. 683–706.
238
K. Matsumura et al.r Brain Research 798 (1998) 232–238
w6x S. Harada, S. Tokunaga, M. Momohara, H. Masaki, T. Tagawa, T. Imaizumi, A. Takeshita, Inhibition of nitric oxide formation in the nucleus tractus solitarius increases renal sympathetic nerve activity in rabbits, Circ. Res. 72 Ž1993. 511–516. w7x Y. Hirooka, J.W. Polson, R.A.L. Dampney, Pressor and sympathoexcitatory effects of nitric oxide in the rostral ventrolateral medulla, J. Hypertens. 14 Ž1996. 1317–1324. w8x C. Iadecola, P.L. Faris, B.K. Hartman, X. Xu, Localization of NADPH diaphorase in neurons of the rostral ventral medulla: possible role of nitric oxide in central autonomic regulation and oxygen chemoreception, Brain Res. 603 Ž1993. 173–179. w9x S. Kagiyama, T. Tsuchihashi, I. Abe, M. Fujishima, Cardiovascular effects of nitric oxide in the rostral ventrolateral medulla of rats, Brain Res. 757 Ž1997. 155–158. w10x S.J. Lewis, H. Ohta, B.H. Machado, J.N. Bates, W.T. Talman, Microinjection of S-nitrosocysteine into the nucleus tractus solitarii decreases arterial pressure and heart rate via activation of soluble guanylate cyclase, Eur. J. Pharmacol. 202 Ž1991. 135–136. w11x K. Matsumura, I. Abe, T. Tsuchihashi, M. Tominaga, K. Kobayashi, M. Fujishima, Central effect of endothelin on neurohormonal responses in conscious rabbits, Hypertension 17 Ž1991. 1192–1196. w12x K. Matsumura, I. Abe, M. Fukuhara, M. Tominaga, T. Tsuchihashi, K. Kobayashi, M. Fujishima, Naloxone augments sympathetic outflow induced by centrally administered endothelin in conscious rabbits, Am. J. Physiol. 266 Ž1994. R1403–R1410. w13x M. Nishimura, H. Takahashi, A. Nanbu, M. Sakamoto, M. Yoshimura, Cardiovascular regulation by L-arginine in the brain of rats. Role of the brain renin–angiotensin system and nitric oxide, Am. J. Hypertens. 10 Ž1997. 389–396. w14x A. Ohta, H. Takagi, T. Matsui, Y. Hamai, S. Iida, H. Esumi, Localization of nitric oxide synthase-immunoreactive neurons in the
w15x
w16x w17x w18x
w19x
w20x
w21x
w22x
w23x
solitary nucleus and ventrolateral medulla oblongata of the rat: their relation to catecholaminergic neurons, Neurosci. Lett. 158 Ž1993. 33–35. H. Ohta, J.N. Bates, S.J. Lewis, W.T. Talman, Actions of Snitrosocysteine in the nucleus tractus solitarii are unrelated to release of nitric oxide, Brain Res. 746 Ž1997. 98–104. R.M.J. Palmer, D.S. Ashton, S. Moncada, Vascular endothelial cells synthesize nitric oxide from L-arginine, Nature 333 Ž1988. 664–666. G. Paxinos, C. Watson ŽEds.., The Rat Brain in Stereotaxic Coordinates, 2nd edn., Academic Press, New York, NY, 1986. I. Sakuma, H. Togashi, M. Yoshioka, H. Saito, M. Yanagida, M. Tamura, T. Kobayashi, H. Yasuda, S.S. Gross, R. Levi, N G-methylL-arginine, an inhibitor of L-arginine-derived nitric oxide synthesis, stimulates renal sympathetic nerve activity in vivo: a role for nitric oxide in the central regulation of sympathetic tone?, Circ. Res. 70 Ž1992. 607–611. L.N. Shapoval, V.F. Sagach, L.S. Pobegailo, Nitric oxide influences ventrolateral medullary mechanisms of vasomotor control in the cat, Neurosci. Lett. 132 Ž1991. 47–50. H. Togashi, I. Sakuma, M. Yoshioka, T. Kobayashi, H. Yasuda, A. Kitabatake, H. Saito, S.S. Gross, R. Levi, A central nervous system action of nitric oxide in blood pressure regulation, J. Pharmacol. Exp. Ther. 262 Ž1992. 343–347. C.-J. Tseng, H.-Y. Liu, H.-C. Lin, L.-P. Ger, C.-S. Tung, M.-H. Yen, Cardiovascular effects of nitric oxide in the brainstem nuclei of rats, Hypertension 27 Ž1996. 36–42. T. Tsuchihashi, I. Abe, M. Fujishima, Role of metabotropic glutamate receptors in ventrolateral medulla of hypertensive rats, Hypertension 24 Ž1994. 648–652. S.R. Vincent, H. Kimura, Histochemical mapping of nitric oxide synthase in the rat brain, Neuroscience 46 Ž1992. 755–784.
Research report
Role of nitric oxide in the nucleus of the solitary tract of rats Kiyoshi Matsumura ) , Takuya Tsuchihashi, Shuntaro Kagiyama, Isao Abe, Masatoshi Fujishima Second Department of Internal Medicine, Faculty of Medicine, Kyushu UniÕersity, Maidashi 3-1-1, Higashi-ku, Fukuoka 812-8582, Japan Accepted 14 April 1998
Abstract We have determined the role of nitric oxide ŽNO. in the nucleus of the solitary tract ŽNTS. of normotensive Wistar rats. The unilateral microinjection of N v-nitro-L-arginine methyl ester Ž10 nmol. to block the synthesis of NO into the NTS significantly decreased the arterial pressure, heart rate ŽHR. and renal sympathetic nerve activity ŽRSNA. Žy19 " 2 mmHg, y23 " 5 beatsrmin, y30 " 2%, respectively.. The microinjection of carboxy-2-phenyl-4,4,5,5-tetramethylimidazoline-1-oxyl 3-oxide ŽCarboxy PTIO. Žtrapper of NO; 0.1 nmol. into the NTS also decreased arterial pressure and RSNA. Conversely, the microinjection of Et 2 NwNŽO.NOxNa ŽNOC 18. ŽNO donor; 10 nmol. caused increases in arterial pressure, HR and RSNA Žq14 " 2 mmHg, q11 " 2 beatsrmin, q38 " 7%, respectively., which was inhibited by the pre-microinjection of Carboxy PTIO Ž0.1 nmol.. On the other hand, not only L-arginine Ž10 nmol. but also D-arginine Ž10 nmol., which is inactive to produce NO, significantly decreased the arterial pressure and RSNA. These results suggest that Ž1. NO acts at the NTS to increase the arterial pressure and RSNA, and Ž2. the microinjection of L-arginine as well as D-arginine led to decreases in arterial pressure and RSNA that were not mediated by the formation of NO in the NTS. q 1998 Elsevier Science B.V. All rights reserved. Keywords: Central nervous system; Microinjection; Renal sympathetic nerve activity; Sympathetic nervous system
1. Introduction Nitric oxide ŽNO., which is responsible for endothelium-dependent vasodilatation, is synthesized from Larginine in the presence of its synthetic enzyme NO synthase ŽNOS. w16x. Immunocytochemical studies have shown that high concentrations of NOS are also present in specific regions of the brain, including the sites involved in the regulation of arterial pressure and the baroreceptor reflex, such as the nucleus of the solitary tract ŽNTS. and the ventrolateral medulla ŽVLM. w8,14,23x. An intramedullary NOS neural pathway that projects from the NTS to the rostral nucleus ambiguus has also been described w3x. Furthermore, various lines of evidence suggest that NO participates in cardiovascular regulation not only by a direct effect on vascular smooth muscle but also via its action on the central nervous system w5,18–21x. The NO is now considered to be a neurotransmitter or a neuromodulator in the central nervous system w4,5x. The microinjection of N G -monomethyl-L-arginine ŽL-NMMA. into the NTS to block endogenous NO caused a decrease
)
Corresponding author. Fax: q 81-92-642-5271; [email protected]
E-mail:
0006-8993r98r$19.00 q 1998 Elsevier Science B.V. All rights reserved. PII S 0 0 0 6 - 8 9 9 3 Ž 9 8 . 0 0 4 2 0 - X
in arterial pressure at a low dose and an increase in arterial pressure at a high dose w21x. It seems to be controversial whether the blockade of endogenous NO in the NTS elicits a pressor response. On the other hand, the microinjection of L-arginine, the NO precursor, into the NTS reduces the arterial pressure w21x. However, L-arginine has been reported to have a non-specific central action w13x. The intracerebroventricular injection of L-arginine causes a transient increase in arterial pressure that is not mediated by a central NO pathway w13x. Furthermore, cardiovascular effect of NO in the NTS remains controversial w6,10,15,21x. We hypothesized that L-arginine may not be an appropriate tool for the examining the central action of NO, even though NO is synthesized from it. The NO donor, Et 2 NwNŽO.NOxNa ŽNOC 18., has been used to study the central effect of NO. It has been reported that the intracerebroventricular injection of NOC 18 led to a decrease in arterial pressure in spontaneously hypertensive rats without a transient increase in arterial pressure w2x. We considered that NOC 18 might be more appropriate than L-arginine for evaluating the central action of NO. Thus, we conducted the present study to re-evaluate the role of endogenous and exogenous NO in the NTS on the cardiovascular and sympathetic responses in anesthetized rats using NOC 18, as well as L-arginine.
K. Matsumura et al.r Brain Research 798 (1998) 232–238
2. Materials and methods 2.1. Preparation of animals Experiments were done in adult male Wistar rats Ž10–12 weeks old.. These experiments were conducted according to the institutional guidelines of Kyushu University on animal experimentation. Rats were anesthetized with urethane Ž1.2 grkg, i.p... Femoral artery and vein were cannulated for the measurement of arterial pressure and the injection of drugs, respectively. The trachea was cannulated to facilitate ventilation. Arterial blood gases and pH were maintained within the physiological range ŽpH 7.39 to 7.42; PO 2 95 to 105 mmHg; PCO 2 34 to 38 mmHg.. Body temperature was maintained at 37.0 " 0.58C by an external heating source. Depth of anesthesia throughout the experiment was monitored by a variety of indexes, including corneal reflexes, as well as respiratory rhythm, arterial pressure, and heart rate ŽHR. stability.
233
An anesthetized rat was placed in a stereotaxic frame ŽDavid Kopf Instruments, Tujunga, CA. with the head fixed downward at 458. After making a midline incision through the skin, the dorsal neck muscles were retracted with sutures to visualize the foramen magnum. The medulla oblongata was exposed by incising the atlanto-occipital membrane and the dorsal surface was kept moist either by artificial cerebrospinal fluid ŽaCSF; pH 7.4. or by production of endogenous CSF. Renal sympathetic nerve activity ŽRSNA. was recorded as described previously w11,12x. Briefly, the left kidney was exposed retroperitoneally, and a branch of the renal nerve was separated from the renal plexus and the surrounding connective tissues with the aid of a dissecting microscope. The RSNA was recorded by a pair of electrodes made from Teflon-insulated seven-stranded steel wire Ž0.001 in. diameter, A-M Systems, Carlsborg, WA.. The area of the nerve and wire interface was embedded in silicone cement ŽElastosil RT 604A and B cement, Wacker Chemicals, Munchen, Germany. to prevent drying of the ¨
Fig. 1. Representative original recordings of arterial pressure, mean arterial pressure, heart rate, renal sympathetic nerve activity ŽRSNA. and integrated RSNA elicited by the microinjection of 10 nmol of N v-nitro-L-arginine methyl ester ŽL-NAME. into the nucleus of the solitary tract.
234
K. Matsumura et al.r Brain Research 798 (1998) 232–238
nerve and to minimize movement artifacts associated with respiration. 2.2. Recording procedures of RSNA The RSNA was amplified Žmodel DPA-100E, Dia Medical System, Tokyo, Japan. and filtered Ž100–3000 Hz., and the waveforms were integrated after a full-wave rectification using an integrator amplifier Žmodel 1322, NEC San-ei, Tokyo, Japan. with the sample-hold function reset to baseline by an internal timer set at 1 s. The residual integrated RSNA which existed after intravenous administration of hexamethonium bromide Ž30 mgrkg, i.v.. was taken as the noise level associated with nerve recording. This value was subtracted from absolute values of integrated RSNA before performing further data analysis. 2.3. Microinjection procedures
2.5. Experimental protocols 2.5.1. Effect of L-NAME in the NTS N v-nitro-L-arginine methyl ester ŽL-NAME; 10 nmol. ŽSigma, St. Louis, MO. was used to block the production of endogenous NO, and was microinjected into the NTS of the rats. As the vehicle control, we administered a microinjection of aCSF Ž50 nl.. 2.6. Effects of NOC 18 and Carboxy PTIO in the NTS The NOC 18 ŽDojindo Institute, Kumamoto, Japan., 1 or 10 nmol, was microinjected into the NTS. In another group of rats, carboxy-2-phenyl-4,4,5,5-tetramethylimidazoline-1-oxyl 3-oxide ŽCarboxy PTIO; 0.1 nmol. ŽDojindo Institute, Kumamoto, Japan., an NO trapper w1x, was microinjected into the NTS 2 min prior to the microinjection of 10 nmol of NOC 18. As the vehicle control, we administered a microinjection of aCSF Ž50 nl..
Microinjections were made from multibarrel micropipettes with tip diameters of 20–50 m m. The pipettes were made from calibrated microbore capillary glass tubing ŽAccu-Fill 90 w , Clay Adams, NJ.. Tips were drawn on a micropipette puller Žmodel PE-2, Narishige Scientific Instruments, Tokyo, Japan.. Injections Ž50 nl. were made over a 30-s period with hand-held syringe as described elsewhere w22x. The injection volume was measured by observing the movement of the fluid meniscus along a reticule in a microscope. The appropriate placement of the pipette tip within the medial NTS was established by microinjection of L-glutamate Ž2 nmol. and observing a depressor response of at least 25 mmHg. On this basis, injections into the medial NTS had coordinates 0.4–0.5 mm rostral and 0.5–0.6 mm lateral to calamus scriptorius and 0.4 mm below the dorsal surface of medulla. All drugs for microinjection were dissolved in aCSF Žin mmolrl: 133.3 NaCl, 3.4 KCl, 1.3 CaCl 2 , 1.2 MgCl 2 , 0.6 NaH 2 PO4 , 32.0 NaHCO 3 , and 3.4 glucose.. Alcian blue dye Ž50 nl. was injected from a separate barrel of the pipette to mark the site of injection at the end of the experiment. 2.4. Histological analysis At the completion of the experiment, the rats were prepared for transcardial perfusion and then euthanatized with a lethal dose of intravenous sodium pentobarbital Ž75 mgrkg.. Animals were then perfused transcardially with 150 ml 0.9% NaCl followed by 150 ml 10% phosphatebuffered formaldehyde solution. The brain was removed and stored in 10% phosphate-buffered formaldehyde solution. The medulla oblongata was cut into 50 m m serial coronal frozen sections that were stained with neutral red. The location of the microinjection sites in the medial NTS was confirmed microscopically according to the atlas of Paxinos and Watson w17x.
Fig. 2. Bar graphs showing the effects of the microinjection of 10 nmol of N v-nitro-L-arginine methyl ester ŽL-NAME. and vehicle Žartificial cerebrospinal fluid; 50 nl. into the nucleus of the solitary tract on changes in mean arterial pressure ŽMAP., heart rate ŽHR. and integrated renal sympathetic nerve activity ŽRSNA. in Wistar rats. Values are means"S.E. Ž)). p- 0.01 as compared with the respective responses to vehicle by Student’s t-test. Number of rats appears in parentheses.
K. Matsumura et al.r Brain Research 798 (1998) 232–238
235
the microinjection of vehicle ŽSAS Institute, Cary, NC.. A value of p - 0.05 was considered statistically significant.
3. Results 3.1. Effect of L-NAME in the NTS Baseline levels of mean arterial pressure ŽMAP. and HR were 96.0 " 1.2 mmHg and 386.9 " 14.2 beatsrmin in L-NAME-microinjected rats, and 93.7 " 3.1 mmHg and 410.7 " 17.2 beatsrmin in aCSF-microinjected rats, respectively. Fig. 1 shows the typical responses of MAP, HR and RSNA that were elicited by the microinjection of L-NAME into the NTS. The microinjection of L-NAME into the NTS caused significant decreases in MAP, HR and RSNA as compared with the effects of aCSF ŽFig. 2..
Fig. 3. Representative original recordings of arterial pressure, mean arterial pressure, heart rate, renal sympathetic nerve activity ŽRSNA. and integrated RSNA elicited by the microinjection of 10 nmol of Et 2 NwNŽO.NOxNa ŽNOC 18. into the nucleus of the solitary tract.
2.7. Effects of L-arginine and D-arginine in the NTS L-Arginine
ŽSigma. or D-arginine ŽSigma., 10 nmol, was microinjected into the NTS of rats. As the vehicle control, we administered a microinjection of aCSF Ž50 nl.. 2.8. Statistics All values are expressed as means " S.E. Student’s t-test was used to determine the effect of the microinjection of L-NAME into the NTS on cardiovascular and sympathetic responses. A one-way analysis of variance was used to evaluate the effects of the microinjection of NOC 18, Carboxy PTIO, L-arginine and D-arginine into the NTS, followed by Duncan’s multiple range test to evaluate the significance of the mean differences in the response to
Fig. 4. Bar graphs showing the effects of Et 2 NwNŽO.NOxNa ŽNOC 18; 1 and 10 nmol., NOC 18 administered following pretreatment with carboxy-2-phenyl-4,4,5,5-tetramethylimidazoline-1-oxyl 3-oxide ŽCarboxy PTIO; 0.1 nmol., and vehicle Žartificial cerebrospinal fluid; 50 nl. microinjected into the nucleus of the solitary tract on changes in mean arterial pressure ŽMAP., heart rate ŽHR. and integrated renal sympathetic nerve activity ŽRSNA. in Wistar rats. Values are means"S.E. Ž)). p- 0.01 as compared with the respective responses to vehicle by Duncan’s multiple range test. Number of rats appears in parentheses.
236
K. Matsumura et al.r Brain Research 798 (1998) 232–238
3.2. Effects of NOC 18 and Carboxy PTIO in the NTS Fig. 3 shows the typical responses of MAP, HR and RSNA that were elicited by the microinjection of NOC 18 Ž10 nmol. into the NTS. The microinjection of NOC 18 Ž1 and 10 nmol. into the NTS caused dose-related increases in the MAP, HR and RSNA ŽFig. 4.. Although the microinjection of Carboxy PTIO into the NTS caused significant decreases in MAP and RSNA Žy9.2 mmHg and y20.1%, respectively., MAP and RSNA returned to the pre-microinjection level within 2 min. Pretreatment with Carboxy PTIO prevented the cardiovascular and RSNA responses to the microinjection of 10 nmol NOC 18 into the NTS ŽFig. 4.. 3.3. Effects of L-arginine and D-arginine in the NTS Fig. 5 illustrates the effects of L-arginine and D-arginine in the NTS. D-Arginine, as well as L-arginine, caused
Fig. 5. Bar graphs showing the effects of L-arginine Ž10 nmol., D-arginine Ž10 nmol., and vehicle Žartificial cerebrospinal fluid; 50 nl. microinjected into the nucleus of the solitary tract on changes in mean arterial pressure ŽMAP., heart rate ŽHR. and integrated renal sympathetic nerve activity ŽRSNA. in Wistar rats. Values are means"S.E. Ž). p- 0.05, Ž)). p- 0.01 as compared with the respective responses to vehicle by Duncan’s multiple range test. Number of rats appears in parentheses.
significant decreases in MAP, HR and RSNA as compared with the effects of the microinjection of aCSF.
4. Discussion We made two principal findings. First, we showed that endogenous, as well as exogenous NO, modulated the sympathetic outflow and arterial pressure in the NTS. The microinjection of NOC 18, an NO donor, into the NTS caused dose-related increases in MAP, HR and RSNA that were prevented by pre-microinjection of Carboxy PTIO, trapper of NO, into the NTS. These results suggest that the cardiovascular and sympathetic responses induced by the microinjection of NOC 18 are mediated by an NO pathway in the NTS. Our second finding was that the microinjection of L-arginine into the NTS caused a decrease in sympathetic outflow, and reduced the arterial pressure and HR. Since the microinjection of D-arginine into the NTS also decreased the MAP, HR and RSNA, cardiovascular and sympathetic responses elicited by the microinjection of L-arginine were considered not to be mediated by an NO pathway. The role of NO in the central nervous system has been examined extensively. The NOS and neuronal NADPH diaphorase in the brain have been shown to be identical; many neurons with NADPH diaphorase activity are located in the NTS and the VLM w7,8,14,23x which are involved in the regulation of the cardiovascular system and the baroreceptor reflex. The NTS is important in regulating sympathetic outflow. However, only a few studies have evaluated the central effects of NO on cardiovascular and sympathetic responses with the use of direct recording of sympathetic nerve activity w6,7,21x. The precise role of the sympathetic nervous system in this respect has not been determined. Furthermore, the effects of NO in the NTS on cardiovascular and sympathetic responses have been controversial. Harada et al. w6x demonstrated that the microinjection of L-NMMA into four sites of the NTS of rabbits to block endogenous NO produced small increases in the arterial pressure and RSNA; these responses were inhibited by pretreatment with a microinjection of L-arginine Ž16 nmol per site.. However, the microinjection of L-arginine into the NTS per se did not cause any cardiovascular or RSNA responses. On the other hand, Tseng et al. w21x reported that the microinjection of L-NMMA into the NTS increased the arterial pressure while it decreased the HR in rats. However, in that study, the microinjection of a low dose of L-NMMA Ž10 nmol. led to a decrease in the arterial pressure. A pressor response was observed only when a high dose of L-NMMA Ž33 or 66 nmol. was microinjected into the NTS; the increase in MAP was less than 10 mmHg. In contrast, the HR showed a decrease at either a low or high dose of L-NMMA. Our present results are consistent with the findings of Tseng et al. w21x regard-
K. Matsumura et al.r Brain Research 798 (1998) 232–238
ing the depressor effect of a microinjection of L-arginine into the NTS, but not with those of Harada et al. w6x. Considering the results of Harada et al. w6x, Tseng et al. w21x and those of the present study, the possible explanations for the differing responses of the arterial pressure are as follows. First, the role of NO in the NTS may differ according to species Žrats vs. rabbits.. Second, the pressor response elicited by the microinjection of a high dose of L-NMMA may be due to the leakage of L-NMMA into the systemic circulation. This could constrict the peripheral arteries and lead to an increase in arterial pressure with a decrease in HR. The occurrence of a bradycardic response, in association with the increase in arterial pressure, also suggests the involvement of a peripheral pressor mechanism that decreases the HR via the baroreceptor reflex. The differing effects of NO on the central nervous system of rats and rabbits may also be supported by studies of the microinjection of L-NAME or L-NMMA into the rostral VLM. Hirooka et al. w7x reported that the microinjection of L-NAME into the rostral VLM of rabbits reduced the arterial pressure and RSNA; conversely, Kagiyama et al. w9x and Tseng et al. w21x demonstrated that microinjections of L-NAME or L-NMMA into the rostral VLM of rats increased the arterial pressure. These divergent responses might be attributable to the species studied. A differing distribution or density of NOS in the brain has been shown in rats and rabbits w7x, and may account for such responses. In contrast to the effects of the NO donor agent ŽNOC 18., the microinjection of the NO precursor, L-arginine, into the NTS decreased the arterial pressure, as well as the HR and RSNA. D-Arginine, an inactive isomer of Larginine that produces NO, also decreased the arterial pressure, HR and RSNA, suggesting that the depressor and sympathoinhibitory responses to L-arginine are not mediated by a central NO pathway. A central cardiovascular effect of L-arginine that was independent of an NO pathway was also observed by Nishimura et al. w13x and Hirooka et al. w7x. The intracerebroventricular injection of L-arginine transiently increased the arterial pressure related to the activation of the central renin–angiotensin system w13x. Moreover, microinjections of L-arginine or D-arginine into the rostral VLM of rabbits decreased the arterial pressure and RSNA, suggesting that these responses were non-specific w7x. Recently, Ohta et al. w15x reported that S-nitrosocysteine, an NO donor, acts at the NTS to reduce the arterial pressure. Its action was independent of the release of NO from nitrosothiol, but may have been mediated via stereoselective sites on target neurons. While those authors did not determine the precise mechanism of the depressor action, their findings appear to be consistent with our present observation that L-arginine acts at the NTS independent of the release of NO. How NO regulates the blood pressure and the sympathetic outflow in the brain is not known. However, NO has been proposed to act via two models w4,5x. In the first
237
model, NO is generated postsynaptically as a result of a rise in cytosolic Ca2q concentration that follows the activation of N-methyl-D-aspartic acid receptors. It then diffuses out and acts on neighbouring neuronal structures, including the presynaptic terminals and glial cells. In the second model, NO is generated presynaptically following the action potential-dependent influx of Ca2q ions. A previous study from our laboratory demonstrated that the microinjection of NOC 18 into the rostral VLM decreases the arterial pressure, whereas the microinjection of LNAME into the rostral VLM increases it w9x. Previous studies of rostral VLM, as well as the present evaluation of NTS, suggest that NO acts on medulla oblongata of rats to inhibit neuronal transmission and to modulate sympathetic outflow. The central regulations of the cardiovascular system and of the sympathetic nervous system may differ between rats and rabbits.
5. Conclusion We have demonstrated that NO acts at the NTS to increase the sympathetic outflow in normotensive rats, leading to increases in arterial pressure and HR. Conversely, the blockade of endogenous NO in the NTS suppresses this sympathetic outflow. Endogenous NO, as well as exogenous NO, modulates sympathetic outflow so as to regulate the arterial pressure in the NTS. The physiological implications await further investigation.
Acknowledgements We thank Miss Hideko Noguchi for expert technical assistance. This work was supported in part by a grant from the Ministry of Education, Japan ŽNo. 09770495. and a Grant for Research on Sympathetic Nervous System and Hypertension.
References w1x T. Akaike, M. Yoshida, Y. Miyamoto, K. Sato, M. Kohno, K. Sasamoto, K. Miyazaki, S. Ueda, H. Maeda, Antagonistic action of imidazolineoxyl N-oxides against endothelium-derived relaxing factorr Ø NO through a radical reaction, Biochemistry 32 Ž1993. 827– 832. w2x C.L. Cabrera, S.L. Bealer, D.F. Bohr, Central depressor action of nitric oxide is deficient in genetic hypertension, Am. J. Hypertens. 9 Ž1996. 237–241. w3x W.-P. Gai, J.P. Messenger, Y.-H. Yu, Z.J. Gieroba, W.W. Blessing, Nitric oxide-synthesising neurons in the central subnucleus of the nucleus tractus solitarius provide a major innervation of the rostral nucleus ambiguus in the rabbit, J. Comp. Neurol. 357 Ž1995. 348–361. w4x J. Garthwaite, Glutamate, nitric oxide and cell–cell signalling in the nervous system, Trends Neurosci. 14 Ž1991. 60–67. w5x J. Garthwaite, C.L. Boulton, Nitric oxide signalling in the central nervous system, Annu. Rev. Physiol. 57 Ž1995. 683–706.
238
K. Matsumura et al.r Brain Research 798 (1998) 232–238
w6x S. Harada, S. Tokunaga, M. Momohara, H. Masaki, T. Tagawa, T. Imaizumi, A. Takeshita, Inhibition of nitric oxide formation in the nucleus tractus solitarius increases renal sympathetic nerve activity in rabbits, Circ. Res. 72 Ž1993. 511–516. w7x Y. Hirooka, J.W. Polson, R.A.L. Dampney, Pressor and sympathoexcitatory effects of nitric oxide in the rostral ventrolateral medulla, J. Hypertens. 14 Ž1996. 1317–1324. w8x C. Iadecola, P.L. Faris, B.K. Hartman, X. Xu, Localization of NADPH diaphorase in neurons of the rostral ventral medulla: possible role of nitric oxide in central autonomic regulation and oxygen chemoreception, Brain Res. 603 Ž1993. 173–179. w9x S. Kagiyama, T. Tsuchihashi, I. Abe, M. Fujishima, Cardiovascular effects of nitric oxide in the rostral ventrolateral medulla of rats, Brain Res. 757 Ž1997. 155–158. w10x S.J. Lewis, H. Ohta, B.H. Machado, J.N. Bates, W.T. Talman, Microinjection of S-nitrosocysteine into the nucleus tractus solitarii decreases arterial pressure and heart rate via activation of soluble guanylate cyclase, Eur. J. Pharmacol. 202 Ž1991. 135–136. w11x K. Matsumura, I. Abe, T. Tsuchihashi, M. Tominaga, K. Kobayashi, M. Fujishima, Central effect of endothelin on neurohormonal responses in conscious rabbits, Hypertension 17 Ž1991. 1192–1196. w12x K. Matsumura, I. Abe, M. Fukuhara, M. Tominaga, T. Tsuchihashi, K. Kobayashi, M. Fujishima, Naloxone augments sympathetic outflow induced by centrally administered endothelin in conscious rabbits, Am. J. Physiol. 266 Ž1994. R1403–R1410. w13x M. Nishimura, H. Takahashi, A. Nanbu, M. Sakamoto, M. Yoshimura, Cardiovascular regulation by L-arginine in the brain of rats. Role of the brain renin–angiotensin system and nitric oxide, Am. J. Hypertens. 10 Ž1997. 389–396. w14x A. Ohta, H. Takagi, T. Matsui, Y. Hamai, S. Iida, H. Esumi, Localization of nitric oxide synthase-immunoreactive neurons in the
w15x
w16x w17x w18x
w19x
w20x
w21x
w22x
w23x
solitary nucleus and ventrolateral medulla oblongata of the rat: their relation to catecholaminergic neurons, Neurosci. Lett. 158 Ž1993. 33–35. H. Ohta, J.N. Bates, S.J. Lewis, W.T. Talman, Actions of Snitrosocysteine in the nucleus tractus solitarii are unrelated to release of nitric oxide, Brain Res. 746 Ž1997. 98–104. R.M.J. Palmer, D.S. Ashton, S. Moncada, Vascular endothelial cells synthesize nitric oxide from L-arginine, Nature 333 Ž1988. 664–666. G. Paxinos, C. Watson ŽEds.., The Rat Brain in Stereotaxic Coordinates, 2nd edn., Academic Press, New York, NY, 1986. I. Sakuma, H. Togashi, M. Yoshioka, H. Saito, M. Yanagida, M. Tamura, T. Kobayashi, H. Yasuda, S.S. Gross, R. Levi, N G-methylL-arginine, an inhibitor of L-arginine-derived nitric oxide synthesis, stimulates renal sympathetic nerve activity in vivo: a role for nitric oxide in the central regulation of sympathetic tone?, Circ. Res. 70 Ž1992. 607–611. L.N. Shapoval, V.F. Sagach, L.S. Pobegailo, Nitric oxide influences ventrolateral medullary mechanisms of vasomotor control in the cat, Neurosci. Lett. 132 Ž1991. 47–50. H. Togashi, I. Sakuma, M. Yoshioka, T. Kobayashi, H. Yasuda, A. Kitabatake, H. Saito, S.S. Gross, R. Levi, A central nervous system action of nitric oxide in blood pressure regulation, J. Pharmacol. Exp. Ther. 262 Ž1992. 343–347. C.-J. Tseng, H.-Y. Liu, H.-C. Lin, L.-P. Ger, C.-S. Tung, M.-H. Yen, Cardiovascular effects of nitric oxide in the brainstem nuclei of rats, Hypertension 27 Ž1996. 36–42. T. Tsuchihashi, I. Abe, M. Fujishima, Role of metabotropic glutamate receptors in ventrolateral medulla of hypertensive rats, Hypertension 24 Ž1994. 648–652. S.R. Vincent, H. Kimura, Histochemical mapping of nitric oxide synthase in the rat brain, Neuroscience 46 Ž1992. 755–784.