American Journal  of Physiology. 2013 Aug; 305(4):R404-13.

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

Min Li, Stephanie C. Tjen-A-Looi, Zhi-Ling Guo, John C. Longhurst

Department of Medicine and Susan Samueli Center of Integrative Medicine, University of California, Irvine, CA 92697

 

Abstract

Acupuncture or electroacupuncture (EA) potentially offers a non-pharmacological approach to reduce high blood pressure (BP).  However, approximately 70% of the patients and animal subjects respond to EA, while 30% do not.  EA acts, in part, through an opioid mechanism in rostral ventrolateral medulla (rVLM) to inhibit sympathoexcitatory reflexes induced by gastric distention.  CCK-8 opposes the action of opioids during analgesia.  We therefore 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 non-responders 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 non-responders.  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.

KEYWORDS: acupuncture, nonresponders, rostral ventrolateral medulla, sympathoexcitation

PMID: 23785073

 

Supplement:

The lifetime risk of developing hypertension, an important underlying cause of stroke and heart attacks, increases with age for middle-aged adults (2013 AHA Statistical Fact Sheet).  The 7th Joint National Committee on Prevention, Detection, Evaluation and Treatment of High Blood Pressure report recommends lifestyle modification (i.e., non-traditional approaches) with or without pharmacological intervention.  Acupuncture or electroacupuncture (EA) potentially offers a non-pharmacological approach to reduce high blood pressure (BP).  Recently, we and others have shown that in mildly hypertensive patients, EA applied once weekly over an eight-week course of therapy lowers BP by 10-12 mmHg, a  response that persists for an additional four weeks after termination of EA stimulation (1,2).  Notably, the hypotensive action of acupuncture, including manual and electroacupuncture (EA), is absent in a subgroup (non-responders) comprising ~ 30% of clinical and experimental subjects undergoing treatment.  The mechanisms of the cardiovascular non-responsiveness to EA are unknown.

Endogenous opioids, including enkephalins, β-endorphin, and dynorphins are chemicals that are produced naturally in the body and function as morphine, a potent analgesic and psychoactive drug.  Opioids act through binding to opioid receptors. There are three kinds of opioid receptors, delta (δ), mu(µ), and kappa (κ) respectively. In addition to addiction and pain modulation, opioid receptors are widely involved in various physiological and pathophysiological activities, including the modulation of cardiovascular responses.

The rostral ventrolateral medulla (rVLM) is located in brain stem. It is an important site for blood pressure regulation.  Our laboratory has shown that enkephalins, endorphin as well as their related receptors in the rVLM importantly participate in cardiovascular modulation by EA.

Cholecystokinin (CCK) is originally found to be located in the gastrointestinal tract and more recently has been identified in the central nervous system as a neurotransmitter that is involved in many important functions, including satiety, pain, cognition and emotion.  The sulfated C-terminal octapeptide of cholecystokinin (CCK-8) is the predominant form of CCK in the central nervous system.  Neurons containing CCK and its receptors are localized in several regions of the brain, including the brain stem.  CCK-8 in the rVLM modulates sympathetic outflow.  This peptide antagonizes the opioid regulation of pain, mood disorders as well as acupuncture’s action through stimulation of CCK1 and CCK2 receptors.  In the last two decades the CCK system has been shown to reduce responsiveness to the analgesic effect of EA.  The specific nuclear location of this action and the mechanism by which CCK improved EA analgesia have not been assessed.  Furthermore, CCK’s role in EA modulation of cardiovascular function has not been evaluated nor the mechanism underlying the complete absence of responsiveness to EA.  Thus, in the present study, we hypothesized that CCK-8 in the rVLM antagonizes the ability of EA modulation of blood pressure increases.

Studies were performed on adult Sprague-Dawley male rats. A femoral vein was cannulated for the administration of fluids, and a femoral artery was cannulated and connected to a pressure transducer to monitor blood pressure.

An unstressed tiny latex balloon was attached to a polyurethane tube and inserted into the stomach through the mouth and esophagus.  A syringe was attached to the cannula to inflate and deflate the balloon with air.  To induce increases in blood pressure, the balloon was inflated inside the stomach.  Distention pressures were selected to fall within the range that a rat normally experiences during ingestion of food and fluids in a single meal.  Increases in blood pressure were observed within 30 s of inflation.  The balloon was deflated within 30 s after reaching the maximal increase in blood pressure.  This sudden increase of BP can be repeated consistently to study how EA can influence reflex hypertension.

Acupuncture needles were placed bilaterally at P5-P6 acupoints at a depth of ~3 mm.  This region of the pericardial meridian located just above the flexor crease in the paw overlies the median nerve.  Stimulation of these acupoints has been shown to evoke strong input into the rVLM.  The needles were connected to a constant current stimulator with stimulus isolation unit and stimulator.  Each set of electrodes was stimulated separately so that current did not flow from one location to the contralateral forelimb.  Correct placement of the needles at the P5-P6 acupoints was confirmed by observing slight repetitive paw twitches at or near motor threshold during EA.  Application of EA lasted 30 min while gastric distention during median nerves afferent stimulation was repeated every 10 min.

Rats not responsive to a single application of EA received microinjection of devazepide, CCK receptor 1 antagonist, into the rVLM.  The CCK antagonist converted all non-responders into responders during a second application of 30 min EA (Fig. 1).

Min Li-1

The importance of this study  Approximately 77.9 million (1 out of every 3) adults have high blood pressure in the US.  Approximately 69% of people with their first heart attack, 77% who experience their first stroke, and 74% with congestive heart failure have blood pressures above 140/90 mmHg.  High blood pressure was either the primary or a contributing cause of death in about 348,102 of the 2.4 million U.S. deaths in 2009 (2013 AHA Statistical Fact Sheet).  It is clear that hypertension and its consequences represent an enormous public health problem.  Acupuncture represents a viable non-pharmacological treatment option.  However, its hypotensive action is absent in ~30% of clinical and experimental subjects undergoing treatment.  The current study demonstrates that non-responders can be converted to responders following blockade CCK1 receptor in rVLM.  This study thus provides not only a better understanding of the complex mechanisms underlying EA’s brainstem modulatory action on reflex elevations in blood pressure but also suggests a potential method for converting EA non-responders into responders to enhance EA’s efficacy in treating hypertension.

 

References

1. Flachskampf FA, Gallasch J, Gefeller O, Gan J, Mao J, Pfahlberg AB, Wortmann A, Klinghammer L, Pflederer W and Daniel WG. Randomized trial of acupuncture to lower blood pressure. Circulation 115: 3121-3129, 2007.

2. Li P and Longhurst JC. Long-lasting inhibitory effect of EA on blood pressure in patients with mild to moderate hypertension. Soc Neurosci 37: 2007.

Acknowledgments: This study was supported by NIH grants HL-63313 and HL-072125 and AHA 10POST4190125.

 

Contact:

Min Li, Ph.D.

Medical Science, C240

School of Medicine

University of California

Irvine CA 92697-4075

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