Modulation of Neurally Mediated Vasodepression and Bradycardia by Electroacupuncture through Opioids in Nucleus Tractus Solitarius.

Stimulation of vagal afferent endings with intravenous phenylbiguanide (PBG) causes both bradycardia and vasodepression, simulating neurally mediated syncope. Activation of µ-opioid receptors in the nucleus tractus solitarius (NTS) increases blood pressure. Electroacupuncture (EA) stimulation of somatosensory nerves underneath acupoints P5-6, ST36-37, LI6-7 or G37-39 selectively but differentially modulates sympathoexcitatory responses. We therefore hypothesized that EA-stimulation at P5-6 or ST36-37, but not LI6-7 or G37-39 acupoints, inhibits the bradycardia and vasodepression through a µ-opioid receptor mechanism in the NTS. We observed that stimulation at acupoints P5-6 and ST36-37 overlying the deep somatosensory nerves and LI6-7 and G37-39 overlying cutaneous nerves differentially evoked NTS neural activity in anesthetized and ventilated animals. Thirty-min of EA-stimulation at P5-6 or ST36-37 reduced the depressor and bradycardia responses to PBG while EA at LI6-7 or G37-39 did not. Congruent with the hemodynamic responses, EA at P5-6 and ST36-37, but not at LI6-7 and G37-39, reduced vagally evoked activity of cardiovascular NTS cells. Finally, opioid receptor blockade in the NTS with naloxone or a specific µ-receptor antagonist reversed P5-6 EA-inhibition of the depressor, bradycardia and vagally evoked NTS activity. These data suggest that point specific EA stimulation inhibits PBG-induced vasodepression and bradycardia responses through a µ-opioid mechanism in the NTS.

Role of TRPV1 in acupuncture modulation of reflex excitatory cardiovascular responses.

We have shown that acupuncture, including manual and electroacupuncture (MA and EA), at the P5-6 acupoints stimulates afferent fibers in the median nerve (MN) to modulate sympathoexcitatory cardiovascular reflexes through central regulation of autonomic function. However, the mechanisms underlying acupuncture activation of these sensory afferent nerves and their cell bodies in the dorsal root ganglia (DRG) are unclear. Transient receptor potential vanilloid type 1 (TRPV1) is present in sensory nerve fibers distributed in the general region of acupoints like ST36 and BL 40 located in the hindlimb. However, the contribution of TRPV1 to activation of sensory nerves by acupuncture, leading to modulation of pressor responses, has not been studied. We hypothesized that TRPV1 participates in acupuncture's activation of sensory afferents and their associated cell bodies in the DRG to modulate pressor reflexes. Local injection of iodoresiniferatoxin (Iodo-RTX; a selective TRPV1 antagonist), but not 5% DMSO (vehicle), into the P6 acupoint on the forelimb reversed the MA's inhibition of pressor reflexes induced by gastric distension (GD). Conversely, inhibition of GD-induced sympathoexcitatory responses by EA at P5-6 was unchanged after administration of Iodo-RTX into P5-6. Single-unit activity of Group III or IV bimodal afferents sensitive to both mechanical and capsaicin stimuli responded to MA stimulation at P6. MA-evoked activity was attenuated significantly ( P < 0.05) by local administration of Iodo-RTX ( n = 12) but not by 5% DMSO ( n = 12) into the region of the P6 acupoint in rats. Administration of Iodo-RTX into P5-6 did not reduce bimodal afferent activity evoked by EA stimulation ( n = 8). Finally, MA at P6 and EA at P5-6 induced phosphorylation of extracellular signal-regulated kinases (ERK; an intracellular signaling messenger involved in cellular excitation) in DRG neurons located at C7-8 spinal levels receiving MN inputs. After TRPV1 was knocked down in the DRG at these spinal levels with intrathecal injection of TRPV1-siRNA, expression of phosphorylated ERK in the DRG neuron was reduced in MA-treated, but not EA-treated animals. These data suggest that TRPV1 in Group III and IV bimodal sensory afferent nerves contributes to acupuncture inhibition of reflex increases in blood pressure and specifically plays an important role during MA but not EA.

Evidence-based blood pressure reducing actions of electroacupuncture: mechanisms and clinical application.

Hypertension is a serious world-wide health problem as it increases cardiovascular atherosclerotic risk, stroke and attending morbidity and mortality. Both systolic and diastolic blood pressures and particularly systolic pressure increase with aging. The downsides from pharmacological therapy have led to consideration of additional treatments, including acupuncture, which evokes endogenous neural-hormonal systems to lower blood pressure. Using basic science studies to guide clinical approaches to research, it is apparent that low frequency, low intensity electroacupuncture reduces sympathetic outflow in approximately 70% of patients with mild to moderate hypertension who are off antihypertensive drugs. Systolic and, to a lesser extent, diastolic arterial blood pressures can be lowered over two to four weeks for prolonged periods, lasting as long as one month, after cessation of an eight weeks of once weekly stimulation. Many questions about long-term therapy, treatment of resistant patients and efficacy in patients on medication remain to be studied. Current data, however, suggest that there may be a role of acupuncture in treatment of hypertension.

Repetitive Electroacupuncture Attenuates Cold-Induced Hypertension through Enkephalin in the Rostral Ventral Lateral Medulla.

Acupuncture lowers blood pressure (BP) in hypertension, but mechanisms underlying its action are unclear. To simulate clinical studies, we performed electroacupuncture (EA) in unanesthetized rats with cold-induced hypertension (CIH) induced by six weeks of cold exposure (6 °C). EA (0.1 - 0.4 mA, 2 Hz) was applied at ST36-37 acupoints overlying the deep peroneal nerve for 30 min twice weekly for five weeks while sham-EA was conducted with the same procedures as EA except for no electrical stimulation. Elevated BP was reduced after six sessions of EA treatment and remained low 72 hrs after EA in 18 CIH rats, but not in sham-EA (n = 12) and untreated (n = 6) CIH ones. The mRNA level of preproenkephalin in the rostral ventrolateral medulla (rVLM) 72 hr after EA was increased (n = 9), compared to the sham-EA (n = 6), untreated CIH rats (n = 6) and normotensive control animals (n = 6). Microinjection of ICI 174,864, a δ-opioid receptor antagonist, into the rVLM of EA-treated CIH rats partially reversed EA's effect on elevated BP (n = 4). Stimulation of rVLM of CIH rats treated with sham-EA using a δ-opioid agonist, DADLE, decreased BP (n = 6). These data suggest that increased enkephalin in the rVLM induced by repetitive EA contributes to BP lowering action of EA.

elPBN neurons regulate rVLM activity through elPBN-rVLM projections during activation of cardiac sympathetic afferent nerves.

The external lateral parabrachial nucleus (elPBN) within the pons and rostral ventrolateral medulla (rVLM) contributes to central processing of excitatory cardiovascular reflexes during stimulation of cardiac sympathetic afferent nerves (CSAN). However, the importance of elPBN cardiovascular neurons in regulation of rVLM activity during CSAN activation remains unclear. We hypothesized that CSAN stimulation excites the elPBN cardiovascular neurons and, in turn, increases rVLM activity through elPBN-rVLM projections. Compared with controls, in rats subjected to microinjection of retrograde tracer into the rVLM, the numbers of elPBN neurons double-labeled with c-Fos (an immediate early gene) and the tracer were increased after CSAN stimulation (P < 0.05). The majority of these elPBN neurons contain vesicular glutamate transporter 3. In cats, epicardial bradykinin and electrical stimulation of CSAN increased the activity of elPBN cardiovascular neurons, which was attenuated (n = 6, P < 0.05) after blockade of glutamate receptors with iontophoresis of kynurenic acid (Kyn, 25 mM). In separate cats, microinjection of Kyn (1.25 nmol/50 nl) into the elPBN reduced rVLM activity evoked by both bradykinin and electrical stimulation (n = 5, P < 0.05). Excitation of the elPBN with microinjection of dl-homocysteic acid (2 nmol/50 nl) significantly increased basal and CSAN-evoked rVLM activity. However, the enhanced rVLM activity induced by dl-homocysteic acid injected into the elPBN was reversed following iontophoresis of Kyn into the rVLM (n = 7, P < 0.05). These data suggest that cardiac sympathetic afferent stimulation activates cardiovascular neurons in the elPBN and rVLM sequentially through a monosynaptic (glutamatergic) excitatory elPBN-rVLM pathway.

Paraventricular Nucleus Modulates Excitatory Cardiovascular Reflexes during Electroacupuncture.

The paraventricular nucleus (PVN) regulates sympathetic outflow and blood pressure. Somatic afferent stimulation activates neurons in the hypothalamic PVN. Parvocellular PVN neurons project to sympathoexcitatory cardiovascular regions of the rostral ventrolateral medulla (rVLM). Electroacupuncture (EA) stimulates the median nerve (P5-P6) to modulate sympathoexcitatory responses. We hypothesized that the PVN and its projections to the rVLM participate in the EA-modulation of sympathoexcitatory cardiovascular responses. Cats were anesthetized and ventilated. Heart rate and mean blood pressure were monitored. Application of bradykinin every 10-min on the gallbladder induced consistent pressor reflex responses. Thirty-min of bilateral EA stimulation at acupoints P5-P6 reduced the pressor responses for at least 60-min. Inhibition of the PVN with naloxone reversed the EA-inhibition. Responses of cardiovascular barosensitive rVLM neurons evoked by splanchnic nerve stimulation were reduced by EA and then restored with opioid receptor blockade in the PVN. EA at P5-P6 decreased splanchnic evoked activity of cardiovascular barosensitive PVN neurons that also project directly to the rVLM. PVN neurons labeled with retrograde tracer from rVLM were co-labeled with µ-opioid receptors and juxtaposed to endorphinergic fibers. Thus, the PVN and its projection to rVLM are important in processing acupuncture modulation of elevated blood pressure responses through a PVN opioid mechanism.

Long-Lasting Reduction of Blood Pressure by Electroacupuncture in Patients with Hypertension: Randomized Controlled Trial.

Background: Acupuncture at specific acupoints has experimentally been found to reduce chronically elevated blood pressure. Objective: To examine effectiveness of electroacupuncture (EA) at select acupoints to reduce systolic blood pressure (SBP) and diastolic blood pressures (DBP) in hypertensive patients. Design: Two-arm parallel study. Patients: Sixty-five hypertensive patients not receiving medication were assigned randomly to one of the two acupuncture intervention (33 versus 32 patients). Intervention: Patients were assessed with 24-hour ambulatory blood pressure monitoring. They were treated with 30-minutes of EA at PC 5-6+ST 36-37 or LI 6-7+GB 37-39 once weekly for 8 weeks. Four acupuncturists provided single-blinded treatment. Main outcome measures: Primary outcomes measuring effectiveness of EA were peak and average SBP and DBP. Secondary outcomes examined underlying mechanisms of acupuncture with plasma norepinephrine, renin, and aldosterone before and after 8 weeks of treatment. Outcomes were obtained by double-blinded evaluation. Results: After 8 weeks, 33 patients treated with EA at PC 5-6+ST 36-37 had decreased peak and average SBP and DBP, compared with 32 patients treated with EA at LI 6-7+GB 37-39 control acupoints. Changes in blood pressures significantly differed between the two patient groups. In 14 patients, a long-lasting blood pressure-lowering acupuncture effect was observed for an additional 4 weeks of EA at PC 5-6+ST 36-37. After treatment, the plasma concentration of norepinephrine, which was initially elevated, was decreased by 41%; likewise, renin was decreased by 67% and aldosterone by 22%. Conclusions: EA at select acupoints reduces blood pressure. Sympathetic and renin-aldosterone systems were likely related to the long-lasting EA actions.

GABA in nucleus tractus solitarius participates in electroacupuncture modulation of cardiopulmonary bradycardia reflex.

Phenylbiguanide (PBG) stimulates cardiopulmonary receptors and cardiovascular reflex responses, including decreases in blood pressure and heart rate mediated by the brain stem parasympathetic cardiac neurons in the nucleus ambiguus and nucleus tractus solitarius (NTS). Electroacupuncture (EA) at P5-6 stimulates sensory fibers in the median nerve and modulates these reflex responses. Stimulation of median nerves reverses bradycardia through action of γ-aminobutyric acid (GABA) in the nucleus ambiguus, important in the regulation of heart rate. We do not know whether the NTS or the neurotransmitter mechanisms in this nucleus participate in these modulatory actions by acupuncture. We hypothesized that somatic nerve stimulation during EA (P5-6) modulates cardiopulmonary inhibitory responses through a GABAergic mechanism in the NTS. Anesthetized and ventilated cats were examined during either PBG or direct vagal afferent stimulation while 30 min of EA was applied at P5-6. Reflex heart rate and blood pressure responses and NTS-evoked discharge were recorded. EA reduced the PBG-induced depressor and bradycardia reflexes by 67% and 60%, respectively. Blockade of GABAA receptors in the NTS reversed EA modulation of bradycardia but not the depressor response. During EA, gabazine reversed the vagally evoked discharge activity of cardiovascular NTS neurons. EA modulated the vagal-evoked cardiovascular NTS cellular activity for 60 min. Immunohistochemistry using triple labeling showed GABA immunoreactive fibers juxtaposed to glutamatergic nucleus ambiguus-projecting NTS neurons in rats. These glutamatergic neurons expressed GABAA receptors. These findings suggest that EA inhibits PBG-evoked bradycardia and vagally evoked NTS activity through a GABAergic mechanism, likely involving glutamatergic nucleus ambiguus-projecting NTS neurons.

Acupuncture regulation of blood pressure: two decades of research.

Although mechanisms underlying acupuncture regulation of pain have been studied by a number of laboratories in many countries, much less is known about its ability to modulate cardiovascular function. In the last two decades, our laboratory has systematically investigated the peripheral and central neural mechanisms underlying acupuncture regulation of blood pressure. These observations account for acupuncture's distant actions and, to some extent, its local actions, with respect to the site of needling. Four fundamental findings have advanced our knowledge. First, point-specific effects of acupuncture underlie its cardiovascular actions. Second, variable regions in the supraspinal and spinal central nervous system that receive input from somatic afferent stimulation account for acupuncture's ability to modulate blood pressure. Thus, depending on the underlying situation, for example, high or low blood pressure, acupuncture modifies autonomic outflow by reducing activity in brain stem nuclei that participate in the primary response. Third, repetitive acupuncture through a molecular mechanism can cause prolonged cardiovascular effects that far outlast acupuncture stimulation. Fourth, there is a range of cardiovascular responsiveness to acupuncture that depends, at least in part, on interactions between neural modulators that synaptically regulate autonomic function in the brain stem. Thus, acupuncture has the capability of profoundly regulating cardiovascular function in patients with disease, for example, hypertension, and the experimental laboratory is directing best approaches to study its actions in humans.

Electroacupuncture modulation of reflex hypertension in rats: role of cholecystokinin octapeptide.

Acupuncture or electroacupuncture (EA) potentially offers a nonpharmacological approach to reduce high blood pressure (BP). However, ~70% of the patients and animal subjects respond to EA, while 30% do not. EA acts, in part, through an opioid mechanism in the rostral ventrolateral medulla (rVLM) to inhibit sympathoexcitatory reflexes induced by gastric distention. CCK-8 opposes the action of opioids during analgesia. Therefore, we hypothesized that CCK-8 in the rVLM antagonizes EA modulation of sympathoexcitatory cardiovascular reflex responses. Male rats anesthetized with ketamine and α-chloralose subjected to repeated gastric distension every 10 min were examined for their responsiveness to EA (2 Hz, 0.5 ms, 1-4 mA) at P5-P6 acupoints overlying median nerve. Repeated gastric distension every 10 min evoked consistent sympathoexcitatory responses. EA at P5-P6 modulated gastric distension-induced responses. Microinjection of CCK-8 in the rVLM reversed the EA effect in seven responders. The CCK1 receptor antagonist devazepide microinjected into the rVLM converted six nonresponders to responders by lowering the reflex response from 21 ± 2.2 to 10 ± 2.9 mmHg (first vs. second application of EA). The EA modulatory action in rats converted to responders with devazepide was reversed with rVLM microinjection of naloxone (n = 6). Microinjection of devazepide in the absence of a second application of EA did not influence the primary pressor reflexes of nonresponders. These data suggest that CCK-8 antagonizes EA modulation of sympathoexcitatory cardiovascular responses through an opioid mechanism and that inhibition of CCK-8 can convert animals that initially are unresponsive to EA to become responsive.

Functional role of peripheral opioid receptors in the regulation of cardiac spinal afferent nerve activity during myocardial ischemia.

Thinly myelinated Aδ-fiber and unmyelinated C-fiber cardiac sympathetic (spinal) sensory nerve fibers are activated during myocardial ischemia to transmit the sensation of angina pectoris. Although recent observations showed that myocardial ischemia increases the concentrations of opioid peptides and that the stimulation of peripheral opioid receptors inhibits chemically induced visceral and somatic nociception, the role of opioids in cardiac spinal afferent signaling during myocardial ischemia has not been studied. The present study tested the hypothesis that peripheral opioid receptors modulate cardiac spinal afferent nerve activity during myocardial ischemia by suppressing the responses of cardiac afferent nerve to ischemic mediators like bradykinin and extracellular ATP. The nerve activity of single unit cardiac afferents was recorded from the left sympathetic chain (T2-T5) in anesthetized cats. Forty-three ischemically sensitive afferent nerves (conduction velocity: 0.32-3.90 m/s) with receptive fields in the left and right ventricles were identified. The responses of these afferent nerves to repeat ischemia or ischemic mediators were further studied in the following protocols. First, epicardial administration of naloxone (8 µmol), a nonselective opioid receptor antagonist, enhanced the responses of eight cardiac afferent nerves to recurrent myocardial ischemia by 62%, whereas epicardial application of vehicle (PBS) did not alter the responses of seven other cardiac afferent nerves to ischemia. Second, naloxone applied to the epicardial surface facilitated the responses of seven cardiac afferent nerves to epicardial ATP by 76%. Third, administration of naloxone enhanced the responses of seven other afferent nerves to bradykinin by 85%. In contrast, in the absence of naloxone, cardiac afferent nerves consistently responded to repeated application of ATP (n = 7) or bradykinin (n = 7). These data suggest that peripheral opioid peptides suppress the responses of cardiac sympathetic afferent nerves to myocardial ischemia and ischemic mediators like ATP and bradykinin.

Medullary GABAergic mechanisms contribute to electroacupuncture modulation of cardiovascular depressor responses during gastric distention in rats.

Electroacupuncture (EA) at P5-P6 acupoints overlying the median nerves typically reduces sympathoexcitatory blood pressure (BP) reflex responses in eucapnic rats. Gastric distention in hypercapnic acidotic rats, by activating both vagal and sympathetic afferents, decreases heart rate (HR) and BP through actions in the rostral ventrolateral medulla (rVLM) and nucleus ambiguus (NAmb), leading to sympathetic withdrawal and parasympathetic activation, respectively. A GABAA mechanism in the rVLM mediates the decreased sympathetic outflow. The present study investigated the hypothesis that EA modulates gastric distention-induced hemodynamic depressor and bradycardia responses through nuclei that process parasympathetic and sympathetic outflow. Anesthetized hypercapnic acidotic rats manifested repeatable decreases in BP and HR with gastric distention every 10 min. Bilateral EA at P5-P6 for 30 min reversed the hypotensive response from -26 ± 3 to -6 ± 1 mmHg and the bradycardia from -35 ± 11 to -10 ± 3 beats/min for a period that lasted more than 70 min. Immunohistochemistry and in situ hybridization to detect c-Fos protein and GAD 67 mRNA expression showed that GABAergic caudal ventral lateral medulla (cVLM) neurons were activated by EA. Glutamatergic antagonism of cVLM neurons with kynurenic acid reversed the actions of EA. Gabazine used to block GABAA receptors microinjected into the rVLM or cVLM reversed EA's action on both the reflex depressor and bradycardia responses. EA modulation of the decreased HR was inhibited by microinjection of gabazine into the NAmb. Thus, EA through GABAA receptor mechanisms in the rVLM, cVLM, and NAmb modulates gastric distention-induced reflex sympathoinhibition and vagal excitation.

Repetitive electroacupuncture causes prolonged increased met-enkephalin expression in the rVLM of conscious rats.

Enkephalinergic neurons in the rostral ventrolateral medulla (rVLM), an important presympathetic region in the brainstem, are activated by 30 min of low frequency (2 Hz) electroacupuncture (EA) at acupoints P5-P6, which overlie the median nerves. To more closely model the clinical application of acupuncture, we administered EA for 30 min twice over a 72 h period to unsedated conscious rats to examine its prolonged action. We hypothesized that repetitive EA would increase preproenkephalin mRNA and met-enkephalin in the rVLM of unsedated conscious rats. Rats received either EA (1-4 mA, 0.5 ms, 2 Hz) or sham stimulation (needle placement without electrical stimulation) twice at P5-P6 acupoints bilaterally. Preproenkephalin mRNA and its peptide met-enkephalin in the rVLM were measured 24 or 48 h after the final EA or sham procedure. Relative ratios of preproenkephalin mRNA levels (normalized with the 18S housekeeping gene) were almost doubled at 24h compared to sham (6.1 ± 0.79 vs. 3.1 ± 0.47). Met-enkephalin measured in rVLM tissue pooled from several rats exposed to the same treatment was increased by repeated EA by 68% after 24h and 51% after 48h, relative to sham. These findings suggest that repeated application of EA in the conscious rats enhances transcription and translation of enkephalin in rVLM for days. Since opioids in the rVLM contribute importantly to the action of EA on sympathetic outflow, this mechanism may contribute to the prolonged action of acupuncture on elevated blood pressure in patients.

Absence of actions of commonly used Chinese herbal medicines and electroacupuncture on myocardial infarct size.

BACKGROUND: Some studies have suggested that certain Chinese herbal remedies and acupuncture could limit ischemia/reperfusion damage. We sought to determine whether the commonly used single herb Danshen (DS), either alone or in combination with Jiang Xiang (JX), or electroacupuncture (EA) reduces myocardial infarct size. METHODS: An anesthetized rat model of proximal left coronary artery occlusion (30 minutes) and reperfusion (180 minutes) was used to measure infarct size (triphenyltetrazolium chloride) and ischemic risk zone (blue dye technique). Rats were either untreated (saline) or received an infusion of DS or DS + JX, starting 30 minutes prior to coronary occlusion. In a separate protocol, rats were untreated, received static needle (ND) placement without stimulation or EA at P5-P6 acupuncture points in the rat forearm starting 5 minutes before occlusion and lasting for 40 minutes, or starting 30 minutes before occlusion and lasting for 90 minutes. RESULTS: In the herbal experiments, myocardial infarct size expressed as a fraction of the ischemic risk zone was 0.43 ± 0.06 in controls, 0.39 ± 0.05 in the DS group, and 0.42 ± 0.04 in the Danshen + JX groups (P = not significant [NS]). In the acupuncture study, there was no significant difference in infarct size as a fraction of the risk zone among the control group (0.38 ± 0.04), the ND group (0.47 ± 0.04), or the EA group (0.32 ± 0.05). When EA was started 30 minutes prior to coronary occlusion and continued for 30 minutes into reperfusion, infarct size was 0.41 ± 0.07 in controls and 0.38 ± 0.10 in EA (P = NS). Neither herbs nor EA altered heart rate or blood pressure. In a separate study of 5 minutes of coronary occlusion plus reperfusion, EA failed to reduce ventricular arrhythmias. CONCLUSION: Our studies do not suggest a cardioprotective effect of DS or DS + JX or EA in an experimental model of myocardial ischemia/reperfusion.

Point specificity in acupuncture.

The existence of point specificity in acupuncture is controversial, because many acupuncture studies using this principle to select control points have found that sham acupoints have similar effects to those of verum acupoints. Furthermore, the results of pain-related studies based on visual analogue scales have not supported the concept of point specificity. In contrast, hemodynamic, functional magnetic resonance imaging and neurophysiological studies evaluating the responses to stimulation of multiple points on the body surface have shown that point-specific actions are present. This review article focuses on clinical and laboratory studies supporting the existence of point specificity in acupuncture and also addresses studies that do not support this concept. Further research is needed to elucidate the point-specific actions of acupuncture.

Nucleus ambiguus cholinergic neurons activated by acupuncture: relation to enkephalin.

Acupuncture regulates autonomic function. Our previous studies have shown that electroacupuncture (EA) at the Jianshi-Neiguan acupoints (P5-P6, underlying the median nerve) inhibits central sympathetic outflow and attenuates excitatory cardiovascular reflexes, in part, through an opioid mechanism. It is unknown if EA at these acupoints influences the parasympathetic system. Thus, using c-Fos expression, we examined activation of nucleus ambiguus (NAmb) neurons by EA, their relation to cholinergic (preganglionic parasympathetic) neurons and those containing enkephalin. To enhance detection of cell bodies containing enkephalin, colchicine (90-100 µg/kg) was administered into the subarachnoid space of cats 30 h prior to EA or sham-operated controls for EA. Following bilateral barodenervation and cervical vagotomy, either EA for 30 min at P5-P6 acupoints or control stimulation (needle placement at P5-P6 without stimulation) was applied. While perikarya containing enkephalin were observed in some medullary nuclei (e.g., raphé), only enkephalin-containing neuronal processes were found in the NAmb. Compared to controls (n=4), more c-Fos immunoreactivity, located principally in close proximity to fibers containing enkephalin was noted in the NAmb of EA-treated cats (n=5; P<0.01). Moreover, neurons double-labeled with c-Fos and choline acetyltransferase in the NAmb were identified in EA-treated, but not control animals. These data demonstrate for the first time that EA activates preganglionic parasympathetic neurons in the NAmb. Because of their close proximity, these EA-activated neurons likely interact with nerve fibers containing enkephalin. These results suggest that EA at the P5-P6 acupoints has the potential to influence parasympathetic outflow and cardiovascular function, likely through an enkephalinergic mechanism.

Neuroendocrine mechanisms of acupuncture in the treatment of hypertension.

Hypertension affects approximately 1 billion individuals worldwide. Pharmacological therapy has not been perfected and often is associated with adverse side effects. Acupuncture is used as an adjunctive treatment for a number of cardiovascular diseases like hypertension. It has long been established that the two major contributors to systemic hypertension are the intrarenal renin-angiotensin system and chronic activation of the sympathetic nervous system. Recent evidence indicates that in some models of cardiovascular disease, blockade of AT1 receptors in the rostral ventrolateral medulla (rVLM) reduces sympathetic nerve activity and blood pressure, suggesting that overactivity of the angiotensin system in this nucleus may play a role in the maintenance of hypertension. Our experimental studies have shown that electroacupuncture stimulation activates neurons in the arcuate nucleus, ventrolateral gray, and nucleus raphe to inhibit the neural activity in the rVLM in a model of visceral reflex stimulation-induced hypertension. This paper will discuss current knowledge of the effects of acupuncture on central nervous system and how they contribute to regulation of acupuncture on the endocrine system to provide a perspective on the future of treatment of hypertension with this ancient technique.

Ionotropic glutamate receptors in the external lateral parabrachial nucleus participate in processing cardiac sympathoexcitatory reflexes.

Stimulation of cardiac sympathetic afferents during myocardial ischemia with metabolites such as bradykinin (BK) evokes sympathoexcitatory reflex responses and activates neurons in the external lateral parabrachial nucleus (elPBN). The present study tested the hypothesis that this region in the pons processes sympathoexcitatory cardiac reflexes through an ionotropic glutamate receptor mechanism. The ischemic metabolite BK (0.1-1 µg) was injected into the pericardial space of anesthetized and bilaterally vagotomized or intact cats. Hemodynamic and renal sympathetic nerve activity (RSNA) responses to repeated administration of BK before and after unilateral 50-nl microinjections of kynurenic acid (Kyn; 25 mM), 2-amino-5-phosphonopentanoic acid (AP5; 25 mM), and 2,3-dihydroxy-6-nitro-7-sulfamoyl-benzol(F)quinoxaline (NBQX; 10 mM) into the elPBN were recorded. Intrapericardial BK evoked significant increases in mean arterial pressure (MAP) and RSNA in seven vagotomized cats. After blockade of glutamate receptors with the nonselective glutamate receptor antagonist Kyn, the BK-evoked reflex increases in MAP (50 ± 6 vs. 29 ± 2 mmHg) and RSNA (59 ± 8.6 vs. 29 ± 4.7%, before vs. after) were significantly attenuated. The BK-evoked responses returned to pre-Kyn levels 85 min after the application of Kyn. Similarly, BK-evoked reflex responses were reversibly attenuated by blockade of glutamate N-methyl-d-aspartate (NMDA) receptors with AP5 (n = 5) and α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) receptors with NBQX (n = 5). In contrast, we observed that the repetitive administration of BK evoked consistent reflex responses including MAP and RSNA before and after microinjection of 50 nl of the artificial cerebrospinal fluid vehicle into the elPBN in five animals. Microinjection of glutamate receptor antagonists into regions outside the elPBN did not alter BK-induced reflex responses. Microinjection of Kyn into the elPBN reversibly attenuated BK-induced reflex responses in four vagus intact animals. These data are the first to show that NMDA and AMPA ionotropic glutamate receptors in the elPBN play an important role in processing cardiac excitatory reflex responses.

Modulation of cardiopulmonary depressor reflex in nucleus ambiguus by electroacupuncture: roles of opioids and γ-aminobutyric acid.

Stimulation of cardiopulmonary receptors with phenylbiguanide (PBG) elicits depressor cardiovascular reflex responses, including decreases in blood pressure and heart rate mediated in part by the brain stem parasympathetic cardiac neurons in the nucleus ambiguus (NAmb). The present study examined NAmb neurotransmitter mechanisms underlying the influence of electroacupuncture (EA) on the PBG-induced hypotension and bradycardia. We hypothesized that somatic stimulation during EA modulates PBG responses through opioid and γ-aminobutyric acid (GABA) modulation in the NAmb. Anesthetized and ventilated cats were studied during repeated stimulation with PBG or cardiac vagal afferents while low-frequency EA (2 Hz) was applied at P5-6 acupoints overlying the median nerve for 30 min and NAmb neuronal activity, heart rate, and blood pressure were recorded. Microinjection of kainic acid into the NAmb attenuated the PBG-induced bradycardia from -60 ± 11 to -36 ± 11 beats/min. Likewise, EA reduced the PBG-induced depressor and bradycardia reflex by 52 and 61%, respectively. Cardiac vagal afferent evoked preganglionic cellular activity in the NAmb was reduced by EA for about 60 min. Blockade of opioid or GABA(A) receptors using naloxone and gabazine reversed the EA-related modulation of the evoked cardiac vagal activity by 73 and 53%, respectively. Similarly, naloxone and gabazine reversed EA modulation of the negative chronotropic responses from -11 ± 5 to -23 ± 6 and -13 ± 4 to -24 ± 3 beats/min, respectively. Thus EA at P5-6 decreases PBG evoked hypotension and bradycardia as well as the NAmb PBG-sensitive preganglionic cardiac vagal outflow through opioid and GABA neurotransmitter systems.

Central and peripheral mechanisms underlying gastric distention inhibitory reflex responses in hypercapnic-acidotic rats.

We have observed that in chloralose-anesthetized animals, gastric distension (GD) typically increases blood pressure (BP) under normoxic normocapnic conditions. However, we recently noted repeatable decreases in BP and heart rate (HR) in hypercapnic-acidotic rats in response to GD. The neural pathways, central processing, and autonomic effector mechanisms involved in this cardiovascular reflex response are unknown. We hypothesized that GD-induced decrease in BP and HR reflex responses are mediated during both withdrawal of sympathetic tone and increased parasympathetic activity, involving the rostral (rVLM) and caudal ventrolateral medulla (cVLM) and the nucleus ambiguus (NA). Rats anesthetized with ketamine and xylazine or α-chloralose were ventilated and monitored for HR and BP changes. The extent of cardiovascular inhibition was related to the extent of hypercapnia and acidosis. Repeated GD with both anesthetics induced consistent falls in BP and HR. The hemodynamic inhibitory response was reduced after blockade of the celiac ganglia or the intraabdominal vagal nerves with lidocaine, suggesting that the decreased BP and HR responses were mediated by both sympathetic and parasympathetic afferents. Blockade of the NA decreased the bradycardia response. Microinjection of kainic acid into the cVLM reduced the inhibitory BP response, whereas depolarization blockade of the rVLM decreased both BP and HR inhibitory responses. Blockade of GABA(A) receptors in the rVLM also reduced the BP and HR reflex responses. Atropine methyl bromide completely blocked the reflex bradycardia, and atenolol blocked the negative chronotropic response. Finally, α(1)-adrenergic blockade with prazosin reversed the depressor. Thus, in the setting of hypercapnic-acidosis, a sympathoinhibitory cardiovascular response is mediated, in part, by splanchnic nerves and is processed through the rVLM and cVLM. Additionally, a vagal excitatory reflex, which involves the NA, facilitates the GD-induced decreases in BP and HR responses. Efferent chronotropic responses involve both increased parasympathetic and reduced sympathetic activity, whereas the decrease in BP is mediated by reduced α-adrenergic tone.