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Cardiac sympathetic nerve activity is an important short-term controller of cardiac function and arterial pressure. Studies also suggest that long-term increases in cardiac sympathetic nerve activity may contribute to hypertension, coronary artery disease, and cardiac remodeling in heart failure. However, our understanding of the role of cardiac sympathetic nerves in chronic models of cardiovascular disease has been limited by inadequate experimental approaches. The present study was conducted to develop a surgical method to surgically denervate the sympathetic nerves of the rat heart for long-term cardiovascular studies. We characterized the effect of cardiac sympathetic denervation on basal levels of mean arterial pressure (MAP) and heart rate (HR) and the responses to a chronic administration of atenolol, a [[beta].sub.1]-adrenoceptor antagonist. Rats were instrumented with telemetry transmitters for continuous recording of MAP and HR. After a 4-day baseline period, the rats were subjected to bilateral stellate ganglionectomy (SGX; n = 9) or sham surgery (Sham; n = 8). Seven days following SGX or Sham, the rats were administered atenolol for 5 days, followed by a 7-day recovery period. Following a transient decrease, SGX had no effect on basal MAP but decreased HR compared with baseline and Sham rats. Five days of atenolol treatment decreased MAP similarly in SGX and Sham rats. Atenolol resulted in a marked bradycardia in Sham rats but had a neglible effects on HR in SGX rats. The measurement of the content of cardiac catecholamines in all cardiac chambers at the end of the study verified a successful sympathetic denervation. This study confirms that bilateral SGX is a useful method to study the contribution of cardiac sympathetic nerves on the regulation of cardiac function. Moreover, these results suggest that cardiac sympathetic nerves are relatively unimportant in maintaining the basal level of MAP or the depressor response to atenolol in conscious, unrestrained rats. cardiac sympathetic nerve activity; heart rate

The cardiac neuronal norepinephrine (NE) transporter (NET) in sympathetic neurons is responsible for uptake of released NE from the neuroeffector junction. The purpose of this study was to assess the chamber distribution of cardiac NET protein measured using [[sup.3]H]nisoxetine binding in rat heart membranes and to correlate NE content to NET amount. In whole mounts of atria, NET was colocalized in nerve fibers with tyrosine hydroxylase (TH) immunoreactivity. NE content expressed as micrograms NE per gram tissue was lowest in the ventricles; however, NET binding was significantly higher in the left ventricle than the right ventricle and atria (P < 0.05), resulting in a significant negative correlation ([r.sup.2] = 0.922; P < 0.05) of NET to NE content. The neurotoxin 6-hydroxydopamine, an NET substrate, reduced NE content more in the ventricles than the atria, demonstrating functional significance of high ventricular NET binding. In summary, there is a ventricular predominance of NET binding that corresponds to a high NE reuptake capacity in the ventricles, yet negatively correlates to tissue NE content. reuptake; 6-hydroxydopamine; stellate ganglia; noradrenaline

Statins have recently been shown to exert neuronal protection in ischemic stroke. Reactive oxygen species, specifically superoxide formed during the early phase of reperfusion, augment neuronal injury. NADPH oxidase is a key enzyme for superoxide production. The present study tested the hypothesis that atorvastatin protects against cerebral infarction via inhibition of NADPH oxidase-derived superoxide in transient focal ischemia. Transient focal ischemia was created in halothane-anesthetized adult male Sprague-Dawley rats (250-300 g) by middle cerebral artery occlusion (MCAO). Atorvastatin (Lipitor, 10 mg/kg sc) was administered three times before MCAO. Infarct volume was measured by triphenyltetrazolium chloride staining. NADPH oxidase enzymatic activity and superoxide levels were quantified in the ischemic core and penumbral regions by lucigenin (5 [micro]M)-enhanced chemiluminescence. Expression of NADPH oxidase membrane subunit [gp91.sup.phox] and membrane-translocated subunit [p47.sup.phox] and small GTPase Rac-1 was analyzed by Western blot. NADPH oxidase activity and superoxide levels increased after reperfusion and peaked within 2 h of reperfusion in the penumbra, but not in the ischemic core, in MCAO rats. Atorvastatin pretreatment prevented these increases, blunted expression of membrane subunit [gp91.sup.phox], and prevented translocation of cytoplasmic subunit [p47.sup.phox] to the membrane in the penumbra 2 h after repeffusion. Consequently, cerebral infarct volume was significantly reduced in atorvastatin-treated compared with nontreated MCAO rats 24 h after reperfusion. These results indicate that atorvastatin protects against cerebral infarction via inhibition of NADPH oxidase-derived superoxide in transient focal ischemia. transient focal ischemia; neuronal injury; inducible nitric oxide synthase; oxidative stress; statins

Superoxide anion ([O.sup.-.sub.2]*) production is elevated in sympathetic ganglion neurons and in the vasculature of hypertensive animals; however, it is not known what enzymatic pathway(s) are responsible for [O.sup.-.sub.2]* production. To determine the pathway(s) of [O.sup.-.sub.2]* production in sympathetic neurons, we examined the presence of mRNA of NADPH oxidase subunits in sympathetic ganglionic neurons and differentiated PC-12 cells. The mRNAs for NADPH oxidase subunits [p47.sup.phox], [p22.sup.phox], [gp91.sup.phox], and NOX1 were present in sympathetic neurons and PC-12 cells, whereas the NOX4 homologue was present in sympathetic neurons but not PC-12 cells. Freshly dissociated celiac ganglion neurons from normal rats and PC-12 cells produced [O.sup.-.sub.2]* when treated with the PKC activator PMA; [O.sup.-.sub.2]* production increased by 317% and 254%, respectively. The PMA-evoked increases were reduced by pretreatment with the NADPH oxidase inhibitor apocynin. These findings indicate that NADPH oxidase is the primary source of [O.sup.-.sub.2]* in sympathetic ganglion neurons. When celiac ganglia from hypertensive rats were incubated with apocynin, [O.sup.-.sub.2]* levels were reduced to the same levels as normotensive animals, indicating that NADPH oxidase activity accounted for the elevated [O.sup.-.sub.2]* levels in hypertensive animals. To test this latter finding, we compared NADPH oxidase activity in extracts of prevertebral sympathetic ganglia of DOCA-salt hypertensive rats and sham-operated rats. NADPH oxidase activities were 49.9% and 78.6% higher in sympathetic ganglia of DOCA rats compared with normotensive controls when using [[beta]-NADH and [[beta]-NADPH as substrates, respectively. Thus elevated [O.sup.-.sub.2]* levels in hypertension may be a result of the increased activity of NADPH oxidase in postganglionic sympathetic neurons. sympathetic ganglia; phorbol ester; reduced nicotinamide adenine dinucleotide phosphate oxidase; hypertension; deoxycorticosterone acetate

Dai and colleagues (Dai X, Galligan JJ, Watts SW, Fink GD, and Kreulen DL. Hypertension 43: 1048-1054, 2004) found that endothelin (ET) stimulated [O.sup.-.sub.2] production in sympathetic ganglion neurons in vitro by activating [ET.sub.B] receptors. The objective of the present study was to determine whether activation of [ET.sub.B] receptors in vivo elevates Of levels in sympathetic ganglia. Because [ET.sub.B] receptor activation increases blood pressure, we also sought to determine whether alteration in [O.sup.-.sub.2] levels was a direct effect of [ET.sub.B] receptor activation on sympathetic ganglia or an indirect consequence of hypertension. Male Sprague-Dawley rats received intravenous infusions of either the specific [ET.sub.B] receptor agonist sarafotoxin 6c (S6c; 5 pmol x [kg.sup.-1] x [min.sup.-1]) or isotonic saline at 0.01 ml/min (control) for 120 min. To measure [O.sup.-.sub.2] levels, we removed the inferior mesenteric ganglion immediately after infusion and stained it with dihydroethidine (DHE). Mean arterial pressure increased 26.6 [+ or -] 1.7 mmHg in the S6c-treated rats and 3.65 [+ or -] 6 mmHg in control rats. Measurements of average pixel intensity revealed that the DHE fluorescence in ganglionic neurons and surrounding glial cells were 96.7% and 160% greater in S6c-treated than in control rats, respectively. To evaluate the effect of elevated blood pressure on [O.sup.-.sub.2] production, a separate group of rats received phenylephrine (PE; 10 [micro]g x [kg.sup.-1] x [min.sup.-1] iv) for 2 h. MAP increased 31 [+ or -] 1.2 mmHg in PE-infused rats. The DHE fluorescence intensity in ganglia of PE-infused rats was significantly greater than that of control rats, 137.7% in neurons and 104.6% in glia but significantly lower than in ganglia from S6c rats. We conclude that ETB receptor activation in vivo significantly enhances [O.sup.-.sub.2] levels in sympathetic ganglia, due to both pressor effects and direct stimulation of [ET.sub.B] receptors in ganglion cells. hypertension; sarafotoxin 6c; sympathetic nervous activity; oxidative stress; reactive oxygen species

Considering the coexistence of neuropeptide Y (NPY) and norepinephrine in perivascular sympathetic nerves and the known vasoconstrictor cooperation of NPY with norepinephrine, we investigated the involvement of NPY in long-term control of cardiovascular functions using NPY transgenic (NPY-tg) rats. These rats were developed by injection of the rat (Sprague-Dawley) pronuclei with a 14.5-kb clone of the rat structural NPY gene. When compared with nontransgenic littermates, NPY concentrations were significantly increased in a number of cardiovascular tissues of NPY-tg hemizygotes. Direct basal mean arterial pressure and heart rate were not changed, but calculated total vascular resistance was significantly increased in NPY-tg subjects. Arterial pressure increases, in response to norepinephrine injection, were greater in the NPY-tg rats. Also, the hypotension and bradycardia in response to hemorrhage were significantly reduced in NPY-tg subjects. These results indicate that NPY, when expressed in increased amounts, potentiates the pressor effects of norepinephrine and contributes to maintaining blood pressure during hemorrhage, but it does not alter resting blood pressure. These transgenic rats will facilitate studies of the role of NPY signaling in cardiovascular regulation, particularly regarding its functional cooperation with norepinephrine. vasoconstriction; cardiac output; adrenoceptors; hemorrhage; hypersensitivity

Inferior mesenteric ganglion (IMG) neurons projecting to mesenteric arteries are different from neurons projecting from mesenteric veins. These two groups of sympathetic neurons of the IMG projecting to mesenteric arteries were distinguished by their localization, neurochemical phenotype and electrophysiological properties from neurons projecting to mesenteric veins. Their distinction supports the concept of differential sympathetic control of arteries and veins.

Cardiac sympathetic nerve activity is an important short-term controller of cardiac function and arterial pressure. Studies also suggest that long-term increases in cardiac sympathetic nerve activity may contribute to hypertension, coronary artery disease, and cardiac remodeling in heart failure. However, our understanding of the role of cardiac sympathetic nerves in chronic models of cardiovascular disease has been limited by inadequate experimental approaches. The present study was conducted to develop a surgical method to surgically denervate the sympathetic nerves of the rat heart for long-term cardiovascular studies. We characterized the effect of cardiac sympathetic denervation on basal levels of mean arterial pressure (MAP) and heart rate (HR) and the responses to a chronic administration of atenolol, a 31-adrenoceptor antagonist. Rats were instrumented with telemetry transmitters for continuous recording of MAP and HR. After a 4-day baseline period, the rats were subjected to bilateral stellate ganglionectomy (SGX; n = 9) or sham surgery (Sham; n = 8). Seven days following SGX or Sham, the rats were administered atenolol for 5 days, followed by a 7-day recovery period. Following a transient decrease, SGX had no effect on basal MAP but decreased HR compared with baseline and Sham rats. Five days of atenolol treatment decreased MAP similarly in SGX and Sham rats. Atenolol resulted in a marked bradycardia in Sham rats but had a neglible effects on HR in SGX rats. The measurement of the content of cardiac catecholamines in all cardiac chambers at the end of the study verified a successful sympathetic denervation. This study confirms that bilateral SGX is a useful method to study the contribution of cardiac sympathetic nerves on the regulation of cardiac function. Moreover, these results suggest that cardiac sympathetic nerves are relatively unimportant in maintaining the basal level of MAP or the depressor response to atenolol in conscious, unrestrained rats. [ABSTRACT FROM AUTHOR]

Statins have recently been shown to exert neuronal protection in ischemic stroke. Reactive oxygen species, specifically superoxide formed during the early phase of reperfusion, augment neuronal injury. NADPH oxidase is a key enzyme for superoxide production. The present study tested the hypothesis that atorvastatin protects against cerebral infarction via inhibition of NADPH oxidase-derived superoxide in transient focal ischemia. Transient focal ischemia was created in halothane-anesthetized adult male Sprague-Dawley rats (250-300 g) by middle cerebral artery occlusion (MCAO). Atorvastatin (Lipitor, 10 mg/kg Sc) was administered three times before MCAO. Infarct volume was measured by triphenyltetrazohum chloride staining. NADPH oxidase enzymatic activity arid superoxide levels were quantified in the ischemic core and penumbral regions by lucigenin (5 µM)-enhanced chemiluminescence. Expression of NADPH oxidase membrane subunit gp91phox and membrane-translocated subunit p47phox and small GTPase Rac-1 was analyzed by Western blot. NADPH oxidase activity and superoxide levels increased after reperfusion and peaked within 2 h of reperfusion in the penumbra, but not in the ischemic core, in MCAO rats. Atorvastatin pretreatment prevented these increases, blunted expression of membrane subunit gp91phox, and prevented translocation of cytoplasmic subunit p47phox to the membrane in the penumbra 2 h after reperfusion. Consequently, cerebral infarct volume was significantly reduced in atorvastatin-treated compared with nontreated MCAO rats 24 h after reperfusion. These results indicate that atorvastatin protects against cerebral infarction via inhibition of NADPH oxidase-derived superoxide in transient focal ischemia. [ABSTRACT FROM AUTHOR]

Superoxide anion (O-2·) production is elevated in sympathetic ganglion neurons and in the vasculature of hypertensive animals; however, it is not known what enzymatic pathway(s) are responsible for O-2· production. To determine the pathway(s) of O-2· production in sympathetic neurons, we examined the presence of mRNA of NADPH oxidase subunits in sympathetic ganglionic neurons and differentiated PC-12 cells. The mRNAs for NADPH oxidase subunits p47phox, p22phox, gp91phox, and NOX1 were present in sympathetic neurons and PC-12 cells, whereas the NOX4 homologue was present in sympathetic neurons but not PC-12 cells. Freshly dissociated celiac ganglion neurons from normal rats and PC-12 cells produced O-2· when treated with the PKC activator PMA; O-2· production increased by 317% and 254%, respectively. The PMA-evoked increases were reduced by pretreatment with the NADPH oxidase inhibitor apocynin. These findings indicate that NADPH oxidase is the primary source of O-2· in sympathetic ganglion neurons. When celiac ganglia from hypertensive rats were incubated with apocynin, O-2· levels were reduced to the same levels as normotensive animals, indicating that NADPH oxidase activity accounted for the elevated O-2· levels in hypertensive animals. To test this latter finding, we compared NADPH oxidase activity in extracts of prevertebral sympathetic ganglia of DOCA-salt hypertensive rats and shamoperated rats. NADPH oxidase activities were 49.9% and 78.6% higher in sympathetic ganglia of DOCA rats compared with normotensive controls when using β-NADH and β-NADPH as substrates, respectively. Thus elevated O-2· levels in hypertension may be a result of the increased activity of NADPH oxidase in postganglionic sympathetic neurons. [ABSTRACT FROM AUTHOR]

Dai and colleagues (Dai X, Galligan JJ, Watts SW, Fink GD, and Kreulen DL. Hypertension 43: 1048–1054, 2004) found that endothelin (ET) stimulated O2- production in sympathetic ganglion neurons in vitro by activating ETB receptors. The objective of the present study was to determine whether activation of ETB receptors in vivo elevates O2- levels in sympathetic ganglia. Because ETB receptor activation increases blood pressure, we also sought to determine whether alteration in O2- levels was a direct effect of ETH receptor activation on sympathetic ganglia or an indirect consequence of hypertension. Male Sprague-Dawley rats received intravenous infusions of either the specific ETB receptor agonist sarafotoxin 6c (S6c; 5 pmol·kg-1·min-1) or isotonic saline at 0.01 ml/min (control) for 120 min. To measure O2- levels, we removed the inferior mesenteric ganglion immediately after infusion and stained it with dihydroethidine (DHE). Mean arterial pressure increased 26.6 ± 1.7 mmHg in the S6c-treated rats and 3.65 ± 6 mmHg in control rats. Measurements of average pixel intensity revealed that the DHE fluorescence in ganglionic neurons and surrounding glial cells were 96.7% and 160% greater in S6c-treated than in control rats, respectively. To evaluate the effect of elevated blood pressure on O2- production, a separate group of rats received phenylephrine (PE; 10 μg·kg-1·min-1 iv) for 2 h. MAP increased 31 ± 1.2 mmHg in PE-infused rats. The DHE fluorescence intensity in ganglia of PE-infused rats was significantly greater than that of control rats, 137.7% in neurons and 104.6% in glia but significantly lower than in ganglia from $6c rats. We conclude that ETB receptor activation in vivo significantly enhances O2- levels in sympathetic ganglia, due to both pressor effects and direct stimulation of ETB receptors in ganglion cells.N [ABSTRACT FROM AUTHOR]