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Am J Physiol Heart Circ Physiol 287: H1842-H1847, 2004. First published June 17, 2004; doi:10.1152/ajpheart.00013.2004
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Modulation of neurotransmitter release by NO is altered in mesenteric arterial bed of spontaneously hypertensive rats

Lacy L. Kolo, Thomas C. Westfall, and Heather Macarthur

Department of Pharmacological and Physiological Science, Saint Louis University School of Medicine, St. Louis, Missouri 63104

Submitted 28 January 2004 ; accepted in final form 9 June 2004


   ABSTRACT
TOP
 ABSTRACT
MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 GRANTS
 REFERENCES
 
Nitric oxide (NO) reacts with catecholamines resulting in their deactivation. In the present study with the use of the perfused mesenteric arterial bed as a model of the sympathetic neuroeffector junction, the NO synthase (NOS) inhibitor N{omega}-nitro-L-arginine methyl ester (L-NAME) resulted in the enhancement of the periarterial nerve stimulation-induced increase in perfusion pressure and norepinephrine overflow while decreasing neuropeptide Y (NPY) overflow. These changes were prevented by L-arginine, demonstrating that the effects of L-NAME were specific to the inhibition of NOS. From the fact that norepinephrine acts on prejunctional {alpha}2-adrenoceptors to inhibit the evoked release of sympathetic cotransmitters, we carried out experiments in the presence of the {alpha}2-adrenergic receptor antagonist yohimbine to investigate the possibility that the decrease in NPY observed in the presence of L-NAME was due to the increase in bioactive norepinephrine acting on its autoreceptor. Periarterial nerve stimulation in the presence of both L-NAME and yohimbine prevented the previously observed decrease in NPY, indicating that the cause of this decrease was, as predicted, due to {alpha}2-adrenoceptor activation. The periarterial nerve stimulation-induced increase of norepinephrine overflow was greater in the spontaneously hypertensive rat compared with normotensive rats. In contrast to what was observed in the isolated perfused mesenteric arterial bed obtained from normotensive animals, inhibition of NOS did not result in a further increase in the overflow of norepinephine or in a subsequent decrease in NPY. These results demonstrate that, in addition to being a direct vasodilator, NO, by deactivating norepinephrine, can modulate sympathetic neurotransmission and that this modulation is altered in the spontaneously hypertensive rat.

sympathetic neurotransmission; nitric oxide synthase; norepinephrine; neuropeptide Y; vascular tone

IT HAS BEEN ESTABLISHED that the neurotransmitters norepinephrine, neuropeptide Y (NPY), and ATP are colocalized in and coreleased from many sympathetic neurons (7, 10, 35, 41). It has also been demonstrated that the application of each transmitter mimics a phase of sympathetic nerve stimulation and that each phase can be blocked with appropriate antagonists (7, 10, 35, 41). Moreover, norepinephrine, NPY, and ATP all have prejunctional inhibitory actions on sympathetic neurotransmission and can negatively regulate their own release as well as the release of each other (36, 43).

Nonneuronal mediators such as the well-characterized endothelium-derived vasodilator nitric oxide (NO) can also modulate sympathetic neurotransmission. Studies have shown that on sympathetic nerve stimulation, inhibition of NO synthesis results in an increase in vasoconstriction in the rat tail artery (42), in the large coronary artery of anesthetized dogs (46), and in the vessels of the isolated adrenal medulla of the dog (1). We have observed that NO reacts with and deactivates catecholamines but not ATP and NPY (20, 24). For instance, in the isolated perfused mesenteric arterial bed of the rat, the ability of exogenous norepinephrine to increase perfusion pressure was attenuated by the incubation of the norepinephrine with the NO donor diethylamine NONOate (20). More importantly, the inclusion of the NO synthase (NOS) inhibitor N{omega}-nitro-L-arginine methyl ester (L-NAME) in the perfusion buffer, resulted in a concomitant increase in norepinephrine overflow and perfusion pressure in response to periarterial nerve stimulation. This finding demonstrates that through deactivation of norepinephrine, NO modulates adrenergic neurotransmission under normal conditions at the sympathetic neuroeffector junction of the isolated perfused mesenteric arterial bed of the rat (20). From the fact that norepinephrine can negatively regulate sympathetic neurotransmission, it is possible that by deactivating norepinephrine, NO may indirectly modulate release of the sympathetic cotransmitters NPY and ATP.

The ability of NO to react with catecholamines may be important for understanding disease models such as hypertension where a decrease in NO availability has been reported (25, 26, 34). There are conflicting viewpoints as to the cause of this decrease in NO availability. It has been suggested that production of NO is altered; however, expression and activity of endothelial NOS have been found to be increased (32, 39), decreased (4), or unmodified (2) in hypertension. Others (18, 19, 23, 40) have suggested that NO production may not be altered, but its bioavailability may be reduced because of oxidative inactivation by excessive production of the superoxide anion in the vascular wall.

A decrease in available NO would not only result in a decrease in direct vasodilation but also, from previous observations, cause an increase in bioactive norepinephrine and other catecholamines. Indeed, there is an increase of plasma levels of norepinephrine in the presence of NOS inhibitors in the rat chronic renal failure model of hypertension (48). In addition, prolonged blockade of NOS caused by chronic L-NAME treatment resulted in arterial hypertension and elevated plasma levels of epinephrine in rats (22). This evidence also concurs with human studies. In humans with high blood pressure, plasma catecholamine content is increased by an average of 63% compared with normotensive humans (6). There is also support of such an increase of norepinephrine in a hypertensive animal model, the spontaneously hypertensive rat (SHR) (27, 30, 33, 44, 49). SHR animals develop hypertension with age.

Norepinephrine can negatively regulate sympathetic neurotransmission; therefore, a further consequence of the deactivation of norepinephrine by NO is likely to be an indirect modulation of the sympathetic cotransmitters NPY and ATP. Thus the decrease of NO availability in hypertension may also have consequences for the release of NPY and ATP. Alterations in plasma NPY levels have frequently been reported in hypertension models (12, 27, 44), but the results have been conflicting, possibly because of contamination with platelet-derived NPY (29). Responsiveness to ATP and the ATP metabolite adenosine has also been reported to be altered in the SHR (17, 21, 31).

In this study we investigated whether the reaction between endogenous NO and norepinephrine at the vascular neuroeffector junction can modulate sympathetic cotransmission in the isolated perfused mesenteric arterial bed of the rat. Furthermore, we investigated whether there is any alteration in the modulation of sympathetic neurotransmission by NO in the model of hypertension represented by the SHR.


   MATERIALS AND METHODS
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 ABSTRACT
 MATERIALS AND METHODS
RESULTS
 DISCUSSION
 GRANTS
 REFERENCES
 
Materials. Norepinephrine (bitratrate salt); dopamine hydrochloride; 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ), L-NAME, yohimbine, S-nitroso-N-acetylpenicillamine (SNAP), trifluoroacetic acid (TFA), and L-arginine were all purchased from Sigma (St. Louis, MO). NPY was purchased from American Peptide (Sunnyvale, CA). The NPY EIA kit was purchased from Peninsula Laboratories (San Carlos, CA).

Isolated perfused mesenteric preparation of the rat: surgery. All procedures were carried out in accordance with National Institute of Health guidelines and were approved by the Institutional Animal Care and Use committee of Saint Louis University Health Sciences Center. The perfused mesenteric bed of the rat was set up as previously described (15). All experiments were performed with 10–12 wk male Sprague-Dawley (SD), Wistar-Kyoto (WKY), or SHR rats. Animals were housed two to four animals per cage in a constant temperature 12:12-h light-dark cycle room. On the day of the experiment, the rats were anesthetized with pentobarbital sodium (50 mg/kg ip). The abdomen was opened, and the mesenteric arterial bed and associated intestines were removed after ligation of the descending colon proximal to the rectum and the duodenum proximal to the stomach. The superior mesenteric artery was cannulated with polyethylene-90 tubing connected to a syringe and flushed with heparinized saline. The four main branches of the mesenteric artery were ligated. The mesenteric vascular bed was then placed in an organ bath, maintained at 37°C, and perfused and superfused with Krebs buffer using a Gilson minipump at a rate of 3 and 0.5 ml/min, respectively. The Krebs buffer was composed of (in mM) 120 NaCl, 5.0 KCl, 1.2 MgSO4, 2.4 CaCl2, 1.17 KH2PO4 0.027 EDTA, 11.1 glucose, and 25 NaHCO3. The perfusion buffer was maintained at 37°C and aerated with 95% O2-5% CO2. Perfusion pressure was continuously monitored with a pressure transducer and recorded by a Grass 79D recorder. The preparation was allowed to equilibrate for 45 min before experimentation. All drugs used were dissolved in the Krebs buffer and delivered by continuous infusion.

Mesenteric preparation: periarterial nerve stimulation. Platinum ring electrodes were placed around the artery, and the periarterial nerves were stimulated at 12 Hz continuously for 30 s (for norepinephrine) or 1.5 min (for NPY) using a Grass S-88 stimulator. The perfusion buffer was collected in 1-min fractions into 0.1 N perchloric acid with 0.1% cysteine (for norepinephrine) or 1% TFA (for NPY) from the bottom of the organ chamber with the use of a fraction collector. Six-minute samples were pooled into one, and aliquots were taken to measure norepinephrine or NPY as described below. The stimulated release of norepinephrine or NPY was calculated as stimulated transmitter overflow divided by basal transmitter overflow.

NPY measurements. For NPY measurement, the perfusate samples were purified by use of C18 Sep-Pak columns (Peninsula Labs) and measured by an enzyme immunoassay kit (Peninsula Labs). The 96-well plate was read by a Powerwave X plate reader (Biotek instruments; Winooski, VT), and the calculation of sample value was analyzed by KC Junior Software (Biotek instruments).


CATECHOLAMINE MEASUREMENTS. Catecholamines were identified and quantified by high-performance liquid chromatography coupled to electrochemical detection (HPLC-EC) as previously published (3, 8). The system consists of a Varian model 2510 solvent delivery system and a model 410 Prostar autosampler (Varian; Walnut Creek, CA) coupled to a C18 column and an ESA Coluchem II detector. Separations were performed isocratically using a filtered and degassed mobile phase consisting of 10% methanol, 0.1 M sodium phosphate, 0.2 mM sodium octyl sulfate, and 0.1 mM EDTA, adjusted to pH 2.8 with phosphoric acid. The HPLC system is coupled to a computer with which chromatograms were recorded and analyzed with Varian Star workstation software.

Statistical analysis of data. Data are expressed as means ± SE. Statistical analysis of difference was carried out by one-way analysis of variance followed by Newman-Keuls multiple-comparison tests. Statistical differences were accepted when P < 0.05.


   RESULTS
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 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
DISCUSSION
 GRANTS
 REFERENCES
 
Neurotransmitter overflow in the presence of a NOS inhibitor in the perfused mesenteric bed of the Sprague-Dawley rat. Our previous results (20, 24) show that NO reacts with and deactivates catecholamines. Furthermore, NO-induced deactivation of dopamine released from the nerve growth factor-differentiated PC12 cell was accompanied by a concomitant increase in NPY release (unpublished observations). This suggests that exogenous NO in deactivating catecholamines indirectly modulates the release of NPY. However, the question remains as to the consequence of the deactivation of catecholamines by endogenous NO on sympathetic cotransmission at the vascular neuroeffector junction. To address this we used the isolated perfused mesenteric bed of the SD rat as a model of the sympathetic neuroeffector junction.

The SD mesenteric preparation of 10- to 12-wk-old animals was stimulated at 12 Hz, and perfusion pressure, norepinephrine overflow, and NPY overflow were measured. On periarterial nerve stimulation, NPY overflow (4.3 ± 0.6 increase over basal or 266 to 1,156 ng/6 min; n = 12; Fig. 1A) and norepinephrine overflow (2.3 ± 0.4-fold increase over basal or 1.8 to 4.14 ng/6 min; n = 8; Fig. 1B) significantly increased. The inclusion of L-NAME (3 x 10–5 M) decreased NPY overflow (2.2 ± 0.4-fold increase over basal or 225 to 503 ng/6 min; n = 8; Fig. 1A), whereas norepinephrine overflow was further increased (5.7 ± 1.3-fold increase or 1.2 to 6.8 ng/6 min; n = 8; Fig. 1B) on periarterial nerve stimulation. The effect of L-NAME was prevented by L-arginine (3 x 10–4 M), demonstrating that the effects of L-NAME inclusion were indeed caused by inhibition of NOS (n = 4; Fig. 1, A–B).



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  		Fig. 1. Periarterial nerve stimulation of the 10- to 12-wk-old Sprague-Dawley (SD) mesenteric bed increased neuropeptide Y (NPY) overflow (A: S, open bars; n = 12) and norepinephrine overflow (B: S, open bars; n = 8). In the presence of N{omega}-nitro-L-arginine methyl ester (L-NAME, 3 x 10–5 M), the NPY overflow decreased (A: S + L-NAME, closed bars; n = 8), whereas the norepinephrine overflow significantly increased (B: S + L-NAME, closed bars; n = 8). Furthermore, this effect was reversible by inclusion of L-arginine (hatched bars; n = 4). C: decrease in NPY overflow in the presence of L-NAME (A: S + L-NAME) was prevented by the inclusion of yohimbine (+yohimbine, 10–6 M; n = 5). D: inclusion of yohimbine (10–6 M) increased stimulated release of norepinephrine in both the presence and absence of L-NAME (+yohimbine; n = 5). S/B is stimulated neurotransmitter divided by basal neurotransmitter overflow. *P < 0.05 compared with S. {dagger}P < 0.05 compared with S + L-NAME.

 
NOS inhibition increases perfusion pressure in the perfused mesenteric bed of SD rats. The mean basal perfusion pressure for all SD mesenteric preparation experiments was 12 ± 0.8 mmHg. Periarterial nerve stimulation at 12 Hz resulted in an increase in perfusion pressure (111 ± 12 mmHg; n = 20; Table 1). Inclusion of the NOS inhibitor L-NAME (3 x 10–5 M) did not affect the basal perfusion pressure; however, the nerve-stimulated perfusion pressure significantly increased to 174 ± 11 mmHg (n = 11; Table 1). This effect of L-NAME was prevented by the inclusion of L-arginine (3 x 10–4 M), demonstrating that the effects of L-NAME were indeed caused by inhibition of NOS (140 ± 9 mmHg; n = 4; Table 1).


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  		Table 1. Perfusion pressure of isolated mesenteric arterial beds from SD rats.

 
{alpha}2-Antagonism prevents the decrease in NPY overflow observed on NOS inhibition. The decrease in NPY observed in the presence of L-NAME is likely a result of the increase in bioactive norepinephrine in the absence of NO, and hence an increase in negative feedback by norepinephrine through the presynaptic {alpha}2-receptor. To investigate this, we included the {alpha}2-specific antagonist yohimbine in the perfusion buffer. On periarterial nerve stimulation, yohimbine alone did not significantly change NPY overflow (4.3 ± 0.6 to 4.9 ± 1.1-fold increase over basal; n = 5; Fig. 1, A and C). Conversely, yohimbine did increase norepinephrine overflow from 2.3 ± 0.2 to 5.3 ± 1.2-fold increase over basal (n = 5, respectively; Fig. 1, B and D).

In the presence of yohimbine, L-NAME (3 x 10–5 M) did not cause a decrease in NPY overflow (4.7 ± 0.8-fold release over basal; n = 5; Fig. 1C). However, L-NAME was still able to further increase norepinephrine overflow (19.3 ± 5.3-fold increase over basal; n = 5; Fig. 1D). This demonstrates that the increase in bioactive norepinephrine caused by NOS inhibition results in stimulation of the presynaptic {alpha}2-receptor resulting in negative feedback on NPY release. Yohimbine (10–6 M) did not significantly change basal perfusion pressure (16.6 ± 1 vs. 18 ± 2 mmHg), perfusion pressure on stimulation (107 ± 7 vs. 113 ± 20 mmHg), or perfusion pressure on stimulation in the presence of L-NAME (205 ± 15 vs. 193 ± 32 mmHg; Table 1).

Modulation of neurotransmission in the perfused mesenteric bed of the SHR. To ensure that the SHR animals were hypertensive compared with the WKY, the femoral artery of the animals was cannulated and the blood pressure measured by a Grass recorder. The SHR had a significantly higher blood pressure compared with the WKY (134 ± 4 vs. 86 ± 2 mmHg; n = 4; data not shown). As with the SD, the mesenteric preparations from the WKY had a corresponding decrease in stimulated release of NPY (5.8 ± 0.8 to 3.6 ± 0.4-fold increase over basal; n = 7; Fig. 2A) and an increase in the release of norepinephrine (2.3 ± 0.7 to 6.5 ± 1.0-fold increase over basal; n = 7; Fig. 2B) in the presence of L-NAME (3 x 10–5 M). There was no further increase in norepinephrine overflow in the mesenteric arterial preparation from the SHR in the presence of L-NAME (6.5 ± 1.3 to 6.3 ± 2.1-fold increase; n = 7; Fig. 2B). In addition, the SHR did not have a significant decrease in NPY overflow in the presence of L-NAME (3.8 ± 0.4 to 3.0 ± 0.6-fold increase over basal; n = 7; Fig. 2A). All changes observed in the presence of L-NAME were reversible by inclusion of L-arginine (data not shown).



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  		Fig. 2. A: on stimulation (12 Hz, 1.5 min), L-NAME (S + L-NAME, closed bars; n = 7) decreased NPY overflow compared with control stimulation in the normotensive 10- to 12-wk-old Wistar-Kyoto (WKY) rats (S, open bars; n = 7) where S/B is stimulated neurotransmitter divided by basal neurotransmitter overflow. However, in the hypertensive spontaneously hypertensive rat (SHR), L-NAME inclusion did not alter NPY overflow. B: on stimulation (12 Hz, 30 s), L-NAME (S + L-NAME, closed bars; n = 6) increased norepinephrine overflow compared with control stimulation in the normotensive 10- to 12-wk-old WKY rats (S, open bars; n = 6). However, in the 10- to 12-wk-old hypertensive animals (SHR), L-NAME did not further increase norepinephrine overflow. *P < 0.05.

 
The mean basal perfusion pressure for both the WKY and SHR mesenteric preparations was 26 ± 3 and 22 ± 1 mmHg, respectively (n = 16; Fig. 3). Periarterial nerve stimulation at 12 Hz resulted in an increase in perfusion pressure of both the WKY (94 ± 9 mmHg; n = 14; Fig. 3) and SHR animals (149 ± 11 mmHg; n = 16; Fig. 3). Upon inclusion of L-NAME (3 x 10–5 M) for 45 min, the perfusion response of the WKY significantly increased to 143 ± 10 mmHg (n = 14; Fig. 3), and the response of the SHR increased to 202 ± 13 mmHg (n = 16; Fig. 3). All changes observed in the presence of L-NAME were reversible by inclusion of L-arginine (data not shown).



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  		Fig. 3. Periarterial nerve stimulation of the 10- to 12-wk-old WKY mesenteric bed increases perfusion pressure (S, hatched bars; n = 14) over basal pressure (B, open bars; n = 16). In the presence of L-NAME (3 x 10–5 M), the perfusion significantly increased (S + L-NAME, closed bars; n = 14). In the SHR beds, periarterial stimulation increased perfusion pressure over basal pressure (S, hatched bars; n = 16), and the inclusion of L-NAME further increased the perfusion pressure (S + L-NAME, closed bars; n = 16). *P < 0.05.

 

   DISCUSSION
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 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
GRANTS
 REFERENCES
 
It is well accepted that neurotransmission at the sympathetic neuroeffector junction is a finely tuned process controlled by the activation of both auto- and heteroreceptors located on the sympathetic nerve terminal (7, 10, 35, 41). The findings obtained in this study indicate that this fine-tuning also extends to factors released from other locations. First, we have confirmed that the inhibition of endogenous NO synthesis results in a simultaneous increase in nerve-stimulated perfusion pressure in, and norepinephrine overflow from, the isolated perfused mesenteric bed of the rat. These findings further support our previous work showing that endogenous NO modulates sympathetic neurotransmission via deactivation of catecholamines. Furthermore, NOS inhibition results in a concomitant decrease in NPY overflow. This decrease in NPY overflow was prevented in the presence of the {alpha}2-adrenergic antagonist yohimbine. This new finding demonstrates that a consequence of the increase in norepinephrine upon NOS inhibition is an increased stimulation of the prejunctional {alpha}2-receptors, thus resulting in a negative feedback of NPY release. In the hypertensive model of the SHR, norepinephrine overflow and perfusion pressure response are greater than the normotensive animals. Furthermore, the modulation of sympathetic neurotransmission by NO is compromised in that while NOS inhibition does cause a further increase in perfusion pressure, the neurotransmitter overflow is not altered.

Our previous results show that NO reacts with and deactivates catecholamines but not ATP or NPY, as found by the inability of catecholamines to increase the perfusion pressure of an isolated mesenteric arterial bed after incubation with a short-life NO donor (20, 24). Furthermore, our previous studies demonstrate that inhibition of endogenous NOS increases stimulated norepinephrine overflow. However, it was not determined whether NOS inhibition would affect NPY overflow. The studies in this current paper demonstrate that in addition to increasing norepinephrine overflow, NOS inhibition results in a decrease in stimulated NPY overflow. Experiments with the {alpha}2-receptor antagonist yohimbine demonstrated that this decrease in NPY overflow was caused by the actions of norepinephrine. We can surmise from these results that under normal conditions there is a negative modulation of the bioactivity of norpepinephrine by NO and that this in turn alters the cotransmission of NPY, and presumably ATP as well.

Before NOS inhibition, inclusion of yohimbine alone increased nerve-stimulated norepinephrine, but not NPY, overflow from the mesenteric bed, which is expected because antagonism of this receptor prevents norepinephrine from inhibiting neurotransmitter release per se (13). The ability of yohimbine to alter overflow of norepinephrine, but not NPY, may be due to differential modulation of sympathetic cotransmitters. There is emerging evidence that sympathetic cotransmitters can be differentially released and modulated. For instance, clonidine inhibits the release of norepinephrine but not ATP from the myenteric plexus of the guinea pig ileum (14). In addition, yohimbine produces a greater increase in the overflow of norepinephrine than ATP, suggesting that endogenously released norepinephrine has a greater influence on its own release than that of ATP (37). It is likely that this is also the case with the effect of norepinephrine on NPY release.

The ability of NO to react with catecholamines may be important for understanding disease models such as hypertension where a decrease in NO availability has been reported (25, 26, 34). Many types of hypertension are associated with an inadequate availability of NO either through decreased NO synthesis or increased NO metabolism (5, 9, 11). This loss of NO will lead to an increase in vasoconstriction both directly (by removal of a direct vasodilator) and indirectly (by increasing the availability of catecholamines). The increase in catecholamines should not only stimulate the vasculature but also alter the release of the sympathetic cotransmitters ATP and NPY through an increase in negative feedback mechanisms.

To examine how the modulation of sympathetic neurotransmission by NO may be altered in hypertension, the model of the SHR and their genetic controls the WKY was chosen. SHR have elevated sympathoadrenomedullary activity, resulting in an increased efferent sympathetic outflow coupled with the development of hypertension (16, 38). All animals were age matched as the SHR blood pressure changes with age. From birth to 6 wk, the blood pressure of the SHR is comparable to the WKY (28, 45). The blood pressure of the SHR then begins to rise until the rats are 10–12 wk old. After this age, the blood pressure then stabilizes (47).

In the 10- to 12-wk-old WKY, the basal perfusion pressure was significantly higher than the SD. This may be due to differences in genetic strain. For this reason, all experiments were repeated in the WKY animal as a control for SHR. The perfusion pressure response and the level of norepinephrine overflow from the SHR mesenteric preparation upon periarterial nerve stimulation was significantly greater than that of the age-matched normotensive SD and WKY animals. These data are consistent with studies examining catecholamine overflow from the perfused mesenteric arterial bed and the isolated caudal artery (44) and also in the plasma (49) of SHR animals. More importantly, the neurotransmitter release from the SHR mesenteric preparation in the absence of L-NAME closely resembled the release from SD and WKY in the presence of L-NAME, suggesting that NO availability is compromised in the SHR preparation. Conversely, in the same SHR preparation there is an increased perfusion pressure response on nerve stimulation in the presence of L-NAME, suggesting that NO availability is not compromised, at least at the level of the vascular smooth muscle. From the fact that in the vasculature, endothelium-derived NO must first diffuse through the vascular smooth muscle layer to react with norepinephrine at the vascular neuroeffector junction, it is possible that NO (in addition to relaxing the smooth muscle) may react with substances released from this cell layer. Several studies have found that in hypertensive models, there is an excessive production of superoxide anion within the vascular smooth muscle (18, 19, 23, 40). Superoxide anion reacts swiftly with NO, thereby decreasing the bioavailability of NO. Therefore, a possible explanation for the fact that NOS inhibition had no effect on nerve-stimulated norepinephrine overflow from mesenteric preparations taken from the SHR but still increased nerve-stimulated perfusion pressure is that the NO released from the endothelial cells diffuses to the vascular smooth muscle and causes relaxation but is deactivated before it can reach the neuroeffector junction.

In conclusion, our results demonstrate that NO not only modulates vascular reactivity directly by vasorelaxation and indirectly by deactivating norepinephrine, but that the deactivation of catecholamines further results in modulation of the release of the sympathetic cotransmitter NPY. Our results also show that in the 10- to 12-wk-old SHR, the modulation of sympathetic neurotransmission by NO is compromised, although the ability of NO to cause direct vasorelaxation is not.


   GRANTS
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 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 GRANTS
REFERENCES
 
This study was supported by National Heart, Lung, and Blood Institute Grants HL-60260 (to T. C. Westfall) and HL-61836 (to H. Macarthur) and National Institute of General Medical Sciences Grant GM-008306. L. L. Kolo is a recipient of an American Heart Association Predoctoral Fellowship.


   FOOTNOTES
 

Address for reprint requests and other correspondence: H. Macarthur, Dept. of Pharmacological and Physiological Science, Saint Louis Univ., 1402 South Grand Blvd., St. Louis, MO 63104 (E-mail macarthu{at}slu.edu)

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.


   REFERENCES
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 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 GRANTS
 REFERENCES
 

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