Modulation of intrinsic cardiac neurons by spinal cord stimulation: implications for its therapeutic use in angina pectoris.

OBJECTIVE: Electrical stimulation of the dorsal aspect of the upper thoracic spinal cord is used increasingly to treat patients with severe angina pectoris refractory to conventional therapeutic strategies. Clinical studies show that spinal cord stimulation (SCS) is a safe adjunct therapy for cardiac patients, producing anti-anginal as well as anti-ischemic effects. However, little information is yet available about the underlying mechanisms involved. METHODS: In order to determine its mechanism of action, the effects of SCS on the final common integrator of cardiac function, the intrinsic cardiac nervous system, was studied during basal states as well as during transient (2 min) myocardial ischemia. Activity generated by intrinsic cardiac neurons was recorded in 9 anesthetized dogs in the absence and presence of myocardial ischemia before, during and after stimulating the dorsal T1-T2 segments of the spinal cord at 66 and 90% of motor threshold using epidural bipolar electrodes (50 Hz; 0.2 ms; parameters within the therapeutic range used in humans). RESULTS: The SCS suppressed activity generated by intrinsic cardiac neurons. No concomitant change in monitored cardiovascular indices was detected. Neuronal activity increased during transient ventricular ischemia (46%), as well as during the early reperfusion period (68% compared to control). Despite that, activity was suppressed during both states by SCS. CONCLUSIONS: SCS modifies the capacity of intrinsic cardiac neurons to generate activity. SCS also acts to suppress the excitatory effects that local myocardial ischemia exerts on such neurons. Since no significant changes in monitored cardiovascular indices were observed during SCS, it is concluded that modulation of the intrinsic cardiac nervous system might contribute to the therapeutic effects of SCS in patients with angina pectoris.

Selective antagonism of humoral versus neural vasoconstrictor responses by nisoldipine.

The effects of nisoldipine administration on vascular reactivity to humoral and neural vasoconstrictor stimuli were examined in the intact rat. For these experiments, rats were instrumented with miniaturized pulsed Doppler flow probes to allow continuous measurement of renal, mesenteric, and hindquarters blood flow. In conscious and anesthetized rats, intravenous doses of angiotensin II (75 and 150 ng/kg), norepinephrine (0.6 and 1.2 microgram/kg), and epinephrine (0.6 and 1.2 microgram/kg) caused dose-dependent increases in arterial pressure and renal and mesenteric vascular resistance. Nisoldipine (0.7 microgram/min) administration significantly attenuated (p less than 0.05) the pressor and regional vasoconstrictor actions of all three circulating pressor agents; however, nisoldipine infusion had little effect on neurally mediated regional vasoconstrictor responses elicited by electrical stimulation of the posterior hypothalamus or greater splanchnic nerve. These data indicate that nisoldipine depressed vascular responsiveness to humoral vasoconstrictor agents, while neural vasoconstrictor responses were unaffected. Thus nisoldipine appears to exert preferential antagonistic effects on humoral rather than on neural vasoconstrictor stimuli.

Vasopressin-central nervous system interactions in the development of DOCA hypertension.

DOCA-salt hypertension does not develop in rats with hereditary lack of vasopressin (DI rats) nor in rats with lesion of the anteroventral region of the third ventricle (AV3V), an area controlling vasopressin (VP) release. We examined, therefore, the effect of VP treatment on the development of DOCA salt hypertension in AV3V-lesioned (AV3V-L) normal Sprague-Dawley rats and in Brattleboro rats homozygous for diabetes insipidus (DI rats). We also examined changes in vascular reactivity in isolated, perfused kidneys in the experimental groups. Whereas sham-lesioned (SL) rats showed hypertension at 5 weeks, AV3V-L rats showed no change in arterial pressure (AP) after DOCA. AV3V-L rats given VP exhibited only an intermediate rise in AP in spite of the fact that plasma VP levels were comparable in DOCA-treated SL rats and AV3V-L rats. SL and AV3V-L rats given VP showed enhanced renal vascular activity whereas no vascular changes occurred in AV3V-L rats. At 5 weeks post DOCA, intact DI rats given VP were hypertensive and exhibited enhanced renal vascular reactivity. AV3V lesion in DI rats completely prevented VP-induced DOCA/salt hypertension and enhanced vascular responsiveness. These data suggest that VP plays a primary role in DOCA-salt hypertension through an induction of enhanced vascular reactivity and through central mechanisms requiring the integrity of the AV3V region.

Regional hemodynamic effects of antihypertensive renomedullary lipids in conscious rats.

Renomedullary tissue has been proposed to exert an antihypertensive endocrine-like action. The antihypertensive polar renomedullary lipids (APRL) and neutral renomedullary lipids (ANRL) are potential mediators of this action. We evaluated the blood pressure and regional hemodynamic responses to APRL administered peripherally (i.v.) and to the central nervous system (CNS) in normal rats and rats with sinoaortic deafferentation (SAD) to remove baroreflex buffering. Rats were chronically instrumented with Doppler flow probes for measurement of mesenteric, renal, and hind-quarter vascular resistance, with arterial pressure and intravenous catheters, and with lateral cerebroventricular cannuli for intracerebroventricular (i.c.v.) administration. Intravenous APRL (0.01 to 1.0 micrograms) produced a dose-dependent decrease in blood pressure, tachycardia, and dilation of all vascular beds studied. The dose-response relationships were shifted to the left in SAD animals. APRL administered i.c.v. had no effect on intact or SAD rats. Pressor and regional vasoconstrictor responses to norepinephrine, angiotensin, and vasopressin were markedly reduced in SAD animals during constant infusion of APRL. In a second group of conscious SAD animals instrumented for blood pressure and heart rate measurements, intravenous ANRL (500 micrograms) decreased both arterial pressure (-45 +/- 16 mm Hg) and heart rate (-50 +/- 16 bpm). When given i.c.v., however, ANRL (10-100 micrograms) had no significant effect on resting blood pressure or heart rate. These studies suggest that APRL and ANRL produce no significant cardiovascular effects that are mediated through the CNS. However, both lipids are potent depressor agents, and APRL exhibits a strong peripheral vasodilator action and nonspecifically reduces reactivity to vasoconstrictor agents.

Cardiovascular effects of L-glutamate and tetrodotoxin microinjected into the rostral and caudal ventrolateral medulla in normotensive and spontaneously hypertensive rats.

The purpose of this study was to compare the responsiveness of the rostral and caudal ventrolateral medulla in spontaneously hypertensive (SH) and normotensive Wistar-Kyoto (WKY) rats to microinjection of L-glutamate, and to estimate tonic output of these areas by microinjecting the neurotoxin tetrodotoxin. Rats were anesthetized with 1.25 g/kg urethane s.c., implanted with arterial (femoral) and venous (femoral) catheters, artificially ventilated and paralyzed with gallamine triethiodide (10 mg/kg). Using a ventral approach to the brainstem, the mean arterial pressure and heart rate responses to microinjection (30 nl) of L-glutamate (1, 10 and 100 mM) and tetrodotoxin (10 microM) into the rostral and caudal ventrolateral medulla were compared in SH (n = 7) and WKY (n = 7) groups. Microinjection of L-glutamate into the rostral ventrolateral medulla produced equivalent increases in mean arterial pressure (maximum +33 +/- 3 and +36 +/- 6 mm Hg, SH and WKY groups respectively) and minimal changes in heart rate. Similar administration of L-glutamate into the caudal ventrolateral medulla caused decreases in mean arterial pressure and heart rate; changes in mean arterial pressure were significantly greater in the SH group than in the WKY group (-52.3 +/- 2.9 mm Hg for SH, -22.6 +/- 2.6 mm Hg for WKY). Bilateral microinjection of tetrodotoxin into the caudal ventrolateral medulla produced significantly larger increases of mean arterial pressure in WKY rats (+8 +/- 4 vs +46 +/- 8 mm Hg for SH vs WKY). These data indicate that SH rats may have a lower tonic activity of neurons in the caudal ventrolateral medulla, resulting in a lower restraining influence on sympathetic outflow in the SH rat.

The rostral and caudal ventrolateral medulla in young spontaneously hypertensive rats.

This study was designed to compare the cardiovascular influences of the rostral ventrolateral medulla (RVLM) and the caudal ventrolateral medulla (CVLM) in young (5-6 weeks) spontaneously hypertensive (SH) and normotensive Wistar-Kyoto (WKY) rats. SH and WKY groups had similar pressor and depressor responses to microinjection of L-glutamate into the RVLM and the CVLM, respectively. In addition the results of this study indicate a reduced tonic sympathoinhibitory function of the CVLM in young SH rats, which may contribute to the development of hypertension in the spontaneously hypertensive rat.

Diazepam-sensitive GABA(A) receptors in the NTS participate in cardiovascular control.

The present study employed neuropharmacological and receptor binding protocols to determine if diazepam-sensitive (DS) gamma-aminobutyric acid-A (GABA(A)) receptors in the nucleus tractus solitarius (NTS) participate in autonomic regulation of cardiovascular function. The first set of protocols was designed to determine if GABA(A) receptors in the NTS were functionally modulated by the benzodiazepine agonist, diazepam. Mean arterial pressure and heart rate responses to microinjection of GABAergic substances into the NTS were examined in urethane-anesthetized rats. Microinjection of the GABA(A) agonist isoguvacine into the NTS increased mean arterial pressure and heart rate, and these effects were blocked by the GABA(A) receptor antagonist, bicuculline. Preadministration of diazepam into the NTS potentiated the pressor actions of isoguvacine and had variable effects on heart rate changes. Flumazenil, a benzodiazepine antagonist, blocked the diazepam-induced potentiation of the pressor response to isoguvacine. The second protocol employed receptor autoradiography to examine the presence of DS and diazepam-insensitive (DI) GABA(A) receptors in the NTS. Autoradiography confirmed that DS GABA(A) receptors were present in the NTS; however, no measurable levels of DI GABA(A) receptors were detected. We conclude that GABA(A)-mediated integration of central autonomic control in the NTS is mediated solely by DS GABA(A) receptors.

Corticosterone delivery to the amygdala increases corticotropin-releasing factor mRNA in the central amygdaloid nucleus and anxiety-like behavior.

The present study examined the effects of stereotaxic delivery of corticosterone to the amygdala on anxiety-like behavior and corticotropin-releasing factor (CRF) mRNA level in the central nucleus of the amygdala (CeA). Micropellets (30 microg) of crystalline corticosterone or cholesterol (control) were implanted bilaterally at the dorsal margin of the CeA in Wistar rats. Seven days post-implantation, anxiety-like behavior was accessed using an elevated plus-maze. CRF mRNA level in the CeA was determined by in situ hybridization 4 h after being tested on the elevated plus-maze. Corticosterone implants increased indices of anxiety on the elevated plus-maze and produced a concomitant increase in both basal level of CRF mRNA per neuron and the number of neurons with CRF hybridization signal in the CeA. The plus-maze increased CRF mRNA levels in the CeA of cholesterol implanted rats to the elevated basal levels observed in corticosterone treated animals. Exposure to the plus-maze did not increase CRF mRNA level in the CeA of corticosterone implanted rats beyond elevated basal levels. Taken together, these findings support the involvement of the amygdala in anxiety-like behaviors in response to chronically elevated corticosterone and suggests that elevated glucocorticoids may increase anxiety by inducing CRF expression in the CeA.

Low intensity spinal cord stimulation may induce cutaneous vasodilation via CGRP release.

This study examined whether spinal cord stimulation (SCS) at intensities below motor threshold (MT) produces cutaneous vasodilation through sympathetic inhibition and/or antidromic activation of sensory fibers. SCS was applied to anesthetized rats with stimulus parameters used clinically, i.e. 50 Hz, 0.2 ms and stimulus intensities at 30, 60 or 90% of MT. SCS-induced vasodilation was not attenuated by hexamethonium, an autonomic ganglion blocking agent, but was abolished by CGRP-(8-37), an antagonist of the calcitonin gene-related peptide (CGRP) receptor. We concluded that SCS-induced vasodilation under the conditions of this study was mediated by peripheral release of CGRP via antidromic activation of sensory fibers.

Cutaneous blood flow increases in the rat hindpaw during dorsal column stimulation.

The purpose of this study was to determine the optimal stimulation site and parameters that result in the greatest changes in cutaneous blood flow during dorsal column stimulation (DCS). Laser Doppler flowmetry was used to assess cutaneous blood flow changes in both rat hindpaws during DCS with a unipolar ball electrode. We found that stimulating the dorsal column at the L2 spinal segment at 0.6 mA at either 25 or 50 Hz with a pulse duration of 0.2 ms resulted in the largest cutaneous blood flow increases in the rat hindpaw. In addition, the DCS response appeared to be limited primarily to the hindpaw ipsilateral to the site of DCS.

Neurosteroid modulation of [3H]flunitrazepam binding in the medulla: an autoradiographic study.

Neurosteroids bind to unique sites on the GABA(A) receptor complex and modulate receptor function. The effects of neurosteroids on GABA(A) receptors have been well characterized in forebrain regions. However, little is known about their effects on GABA(A) receptors in the medulla, especially those areas involved in autonomic reflex pathways. Stimulation of [3H]flunitrazepam binding to the GABA(A) receptor by two progesterone metabolites, 3alpha-hydroxy-5alpha-pregnan-20-one (3alpha-OH-DHP) and 3beta-hydroxy-5alpha-pregnan-20-one (3beta-OH-DHP), was studied using autoradiographic methods in the medulla and cerebellum of female rats at estrus. [3H]Flunitrazepam binding was enhanced by 3alpha-OH-DHP in every nucleus examined in the medulla and cerebellum. This effect was stereoselective since 3beta-OH-DHP had no effect on binding in any region. No differences were observed in the degree of stimulation of [3H]flunitrazepam binding by 3alpha-OH-DHP among medullary brain regions. However, in the cerebellum, the stimulation of binding was significantly greater in the granular layer than in the molecular layer. Stimulation of [3H]flunitrazepam binding by 3alpha-OH-DHP in nuclei involved in the baroreflex pathways supports previous studies which report that neurosteroids modulate autonomic regulation of blood pressure. These actions may also underlie alterations in autonomic function during pregnancy.

Tonic GABA(A) receptor-mediated neurotransmission in the dorsal vagal complex regulates intestinal motility in rats.

Vagal motor outflow from the dorsal vagal complex is important in the regulation of intestinal motility. The aim of our study was to test the hypothesis that within the dorsal vagal complex, tonic GABA(A)-receptor mediated neurotransmission modulates intestinal motility. The GABA(A) receptor antagonist, bicuculline (methiodide), was microinjected into the dorsal vagal complex, and the effects on small intestinal and colonic motility were investigated. Rats were anesthetized and the mean arterial pressure and heart rate were monitored. Jejunal and colonic motility were measured manometrically, and motility indices were calculated manually. Bicuculline at concentrations of 0.25 or 0.5 mM in 30 nl was microinjected bilaterally into the dorsal vagal complex through stereotaxically placed micropipettes. The injection sites were confirmed histologically using the dye Alcian Blue. Bicuculline (0.5 mM) inhibited spontaneous jejunal motility by 76.3%, colonic motility by 51.7%, mean arterial pressure by 23.3% and heart rate by 27.6%. The lower concentration of bicuculline (0.25 mM) showed no inhibitory effects on intestinal motility but decreased mean arterial blood pressure by 24.1% and heart rate by 13.6%. Bilateral cervical vagotomy attenuated the bicuculline (0.5 mM)-induced inhibition of spontaneous jejunal motility, whereas the bicuculline effect on colonic motility was unaffected. The results of this study show that GABA(A) receptor-mediated neurotransmission in the dorsal vagal complex is involved in autonomic integration of motility of the small intestine and colon. Furthermore, our results indicate that the dorsal vagal complex regulation of jejunal motility involves vagal outflow, whereas vagal pathways do not participate in the bicuculline-induced inhibition of colonic motility.

Alpha 1-adrenoceptor subtypes and the regulation of peripheral hemodynamics in the conscious rat.

The peripheral hemodynamic effects of SZL-49, a prazosin analog capable of selectively inactivating the alpha 1a-adrenoceptor subtype, was evaluated in the conscious rat. One hour after SZL-49 administration, total peripheral vascular resistance and arterial blood pressure significantly decreased and cardiac output and heart rate increased. Twenty-four hours after SZL-49, blood pressure returned to control preinjection levels while peripheral resistance remained decreased and cardiac output and heart rate were elevated. The phenylephrine dose-response curves for mean arterial blood pressure and total peripheral vascular resistance were shifted to the right but the maximal responses were not decreased. These data show that the alpha 1a receptor plays a role in the tonic maintenance of arterial blood pressure. The alpha 1b receptor appears to participate in the response to exogenously administered agonists.

Effects of chronic smoke exposure on the cardiovascular responses to acute nicotine infusion in the rat.

Rats were exposed daily to cigarette smoke for 17-22 weeks in order to characterize mean arterial pressure and regional hemodynamic effects of chronic smoke exposure and to determine if cardiovascular reactivity to acute nicotine infusions is altered by chronic smoke exposure. Urethane-anesthetized animals were instrumented with miniaturized pulsed-Doppler flow probes on the iliac and mesenteric vascular beds. Under resting conditions sham-smoked and smoke-exposed animals had similar levels of mean arterial pressure and mesenteric blood flow; however, resting heart rate was lower in the smoke-exposed group, while iliac blood flow was elevated in the smoke-exposed group. Acute nicotine infusion (6.25, 12.5 and 25 micrograms/kg per min) produced equivalent, dose-dependent pressor effects as well as increases in iliac and mesenteric resistance in sham and smoke-exposed groups. Thus, chronic cigarette smoke-exposure in rats may exert significant cardiovascular effects other than on arterial pressure such as lowered heart rate and elevated blood flow to skeletal muscle beds, while cardiovascular responses to nicotine are not altered by chronic smoke-exposure.

Spinal integration of antidromic mediated cutaneous vasodilation during dorsal spinal cord stimulation in the rat.

The purpose of this study was to determine the involvement of supraspinal centers and spinal synaptic integration in cutaneous vasodilation mediated by dorsal spinal cord stimulation (DCS). Laser Doppler flowmetry was used to assess cutaneous blood flow changes in the rat hindpaw during DCS with a unipolar ball electrode placed at the L2-L3 spinal level. Results demonstrated that transecting the spinal cord at the T10 spinal segment did not alter the DCS response while T13 spinal transection abolished the DCS-induced vasodilation. Inhibition of synaptic activity with topical application of muscimol (0.2 mM) on the dorsal surface of the spinal cord markedly attenuated the DCS response. In conclusion DCS-induced vasodilation involved synaptic integration but did not require input from rostral spinal sites or supraspinal areas.

Role of nitric oxide in cutaneous blood flow increases in the rat hindpaw during dorsal column stimulation.

OBJECTIVE: Dorsal column stimulation (DCS) increases blood flow to the extremities and may produce a limb-saving effect in addition to treatment of refractory chronic pain in patients with peripheral vascular disease. The purpose of this study was to examine the importance of nitric oxide in cutaneous vasodilation caused by DCS. METHODS: Male Sprague-Dawley rats were anesthetized with pentobarbital (60 mg/kg, intraperitoneally). A unipolar ball electrode was placed on the left-side of the exposed spinal cord at approximately L1-L2. Blood flow was concurrently recorded from both hindpaw foot pads with laser doppler flowmeters. Blood flow responses were assessed during 1 minute of DCS (0.6 mA at 50 Hz, 0.2-ms pulse) at 10-minute intervals. Nitric oxide synthase was inhibited with NG-nitro-L-arginine methyl ester (L-NAME). Four groups of animals were examined. The first and second groups involved examination of the effects of DCS after 2 and 10 mg/kg L-NAME, respectively. In the third group, the effect of another nitric oxide synthase inhibitor, NG-monomethyl-L-arginine (10 mg/kg), was examined on the responses to DCS. The fourth group of animals entailed comparison of the effects of DCS under control conditions, after the nicotinic receptor antagonist, hexamethonium (10 mg/kg), and during the combined presence of hexamethonium and L-NAME (10 mg/kg). RESULTS: L-NAME markedly attenuated the cutaneous blood flow increases caused by DCS at doses of 2 or 10 mg/kg. Similarly, NG-monomethyl-L-arginine also attenuated the DCS response. Hexamethonium did not affect the cutaneous vasodilation caused by DCS. After hexamethonium, L-NAME no longer attenuated the DCS response. CONCLUSION: Our results demonstrated that nitric oxide played a significant role in producing the DCS-induced increase in rat cutaneous hindpaw blood flow. The involvement of nitric oxide does not require the presence of autonomic efferent function; however, ganglionic blockade may unmask a mechanism for vasodilation during DCS that is independent of nitric oxide release.

Effect of motor neuromuscular electrical stimulation on microvascular perfusion of stimulated rat skeletal muscle.

The purpose of this study was to determine the effect of neuromuscular electrical stimulation (NMES) (2,500-pps sine wave interrupted at 50 bps) on the degree of microvascular perfusion in stimulated skeletal muscle. The tibialis anterior (TA) and extensor digitorum longus (EDL) muscles of 36 male rats were treated with NMES for 30 minutes at current amplitudes sufficient to produce a sustained muscle contraction (motor NMES). Muscle tissue was removed at 0, 5, 10, 15, and 30 minutes after NMES. The perfused vessel/muscle fiber ratio (PV/F) of the stimulated animals at time 0 minutes was greater than that of the unstimulated control animals. A gradual decrease in the magnitude of the PV/F increase was noted over time. Depending on the muscle's fiber-type composition, the PV/F values returned to control levels by 10 to 30 minutes after motor NMES. The results indicate (1) that motor NMES significantly increases the degree of microvascular perfusion in stimulated rat skeletal muscle and (2) that the increased degree of perfusion persists for various lengths of time, depending on the fiber-type composition of the muscle. Thus, if responses in an animal model can be used as indicators of similar human responses, then the results of this study suggest that NMES can be used to increase the degree of microvascular perfusion in human skeletal muscle.

The influence of muscle contraction on the degree of microvascular perfusion in rat skeletal muscle following transcutaneous neuromuscular electrical stimulation.

Electrotherapy is used clinically according to a variety of protocols and at various intensities with the intent of effecting any number of physiological changes. The purpose of this study was to determine if the increased degree of microvascular perfusion observed following 2,500 Hz transcutaneous neuromuscular electrical stimulation (TNMES) is dependent on evoked muscle contractions. The tibialis anterior (TA) and extensor digitorum longus (EDL) muscles from 30 male rats were analyzed. Six animals were untreated and served as controls, while the TA and EDL muscles of six animals were treated with TNMES at current intensities three times that needed to evoke a minimum visible contraction in the TA (M-TNMES). The remaining animals were treated with gallamine, which effectively blocked neurally mediated muscle contraction. The TA and EDL muscles of six gallamine-treated rats received no TNMES and served as shams (G-Sham), six received M-TNMES (GM-TNMES), and six received TNMES at intensities sufficient to produce sustained muscle contraction with a neuromuscular blockade in place (G-HIS). Perfused microvessels were labeled with fluorescein isothiocyanate-bovine serum albumin. The degree of microvascular perfusion was determined by calculating perfused microvessel/muscle fiber ratios (PV/F). The mean PV/F ratios of all groups were compared using Fisher's LSD (alpha = 0.05). When compared to controls, the PV/F ratios of the TA and EDL muscles in M-TNMES and G-HIS groups showed a significant (p < or = 0.05) increase while the G-Sham and GM-TNMES groups were similar to controls.(ABSTRACT TRUNCATED AT 250 WORDS)