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Participation of Chloride Channel for Cardioprotective Effect Induced by Isoflurane in Guinea Pig Single Cardiac Myocyte
Noboru Hatakeyama, M.D., Ph.D.; Masana Yamada, M.D.; Nobuko Shibuya, M.D., Ph.D.; Yasunori Momose, Ph.D.; Mitsuaki Yamazaki, M.D., Ph.D.
Anesthesiology, Toyama Medical and Pharmaceutical University, Toyama, Toyama, Japan
Introduction: Protection of the myocardium against ischemia-reperfusion damage has been reported with drugs such as K+ channel openers, β-blockers and Ca2+ channel antagonists. These drugs have a common feature that they exert their cardioprotective effect by modifying cation homeostasis and reducing oxygen requirement. It is also reported that general anesthetics such as halothane, isoflurane and sevoflurane seem to have cardioprotective effect through the similar mechanisms mainly via the activation of ATP sensitive K+ channel. Recently, chloride channel blockers such as anthracene-9-carboxylic acid (9-AC), 4-acetamide-4'-isothiocyanato-stilbene-2,2'-disulfonic acid and methyl 2-(N-benzyl-N-methylamino) ethyl-2,6-dimethyl-4-(2-isopropyl-pyrazolo[1,5-a]pyridine-3-yl)-1,4-dihydro-pyridine-3,5-dicarboxylate enhanced the recovery of developed tension after reperfusion1,2. Because the intracellular Cl- activity was reported to be increased during myocardial ischemia3, these chloride channel blockers might attenuate this activity and show cardioprotective effect. Thus, anions can play a role for the protection of myocardium against ischemia- reperfusion damage. We examined the effect of isoflurane on chloride channel current (ICl) and studied whether chloride channel was involved in cardioprotective mechanism induced by isoflurane.

Methods: With institutional approval, guinea pig hearts were excised under sodium pentobarbital anesthesia. Single cardiac myocytes were obtained by circulating enzymes (collagenase and trypsin) through coronary arteries. Whole-cell voltage clamp technique was occupied for the measurement of ICl. Tyrode solution including 0.1 mM CdCl2, 1mM BaCl2 and 10-6 M isoproterenol was used for bathing solution to characterize ICl. To minimize the other K+ currents, pipette solution contained (in mM) Cs+ aspartate 70, CsCl 30, HEPES 5, MgCl2 1, ATP 5, EGTA 5 and tetraethylammonium chloride 20. ICl was elicited by applying test pulses (300 msec, from -80 mV to +60 mV by 10 mV step, every 5 sec.) from a holding potential of -40 mV. In the presence of 9-AC (2 x 10-4 M), ICl was blocked. Isoflurane was applied by bubbling and dissolving into bathing solution after vaporized by the oxygen. The concentration of isoflurane in the vaporized gas was monitored by the anesthetic gas monitor. Further to characterize the effect of isoflurane on ICl, use-dependent block (0.2 Hz) of ICl was examined. All of the experiments were performed in room temperature. Data were analyzed by ANOVA and P<0.05 was considered as significant.

Results: Isoflurane (4%) decreased ICl to 66.2 ± 5.5 % of control (mean ± SEM, n=11, p<0.05) at +60 mV. This inhibitory effect was reversed after 10 min washout. Shift of reversal potential on ICl was not observed in the presence of isoflurane. Inhibition of ICl induced by isoflurane did not show use-dependent block.

Conclusions: Isoflurane (4%) inhibited isoproterenol-induced ICl. The mechanism of inhibition seemed like a direct effect of isoflurane to chloride channels. This mechanism can play a role, at least in part for the myocardial protection induced by isoflurane against ischemia-reperfusion damage.

References: 1. Naunyn-Schmiedeberg's Arch Pharmacol 356: 853-855, 1997. 2. J Pharmacol Exp Ther 278: 854-861, 1996. 3. Jpn J Pharmacol 72: 161-174, 1996.

Anesthesiology 2001; 95:A604