Hulme, J. T., and C. H. Orchard Effect of acidosis on transient outward potassium current in isolated rat ventricular myocytes. Am. J. Physiol. Heart Circ. Physiol. 278: H50-H59, 2000.-The effect of acidosis on the transient outward K ϩ current (I to ) of rat ventricular myocytes has been investigated using the perforated patch-clamp technique. When the holding potential was Ϫ80 mV, depolarizing pulses to potentials positive to Ϫ20 mV activated I to in subepicardial cells but activated little I

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... to in subendocardial cells. Exposure to an acid solution (pH 6.5) had no significant effect on I to activated from this holding potential in either subepicardial or subendocardial cells. When the holding potential was Ϫ40 mV, acidosis significantly increased I to at potentials positive to Ϫ20 mV in subepicardial cells but had little effect on I to in subendocardial cells. The increase in I to in subepicardial cells was inhibited by 10 mM 4-aminopyridine. In subepicardial cells, acidosis caused a ϩ8.57-mV shift in the steady-state inactivation curve. It is concluded that in subepicardial rat ventricular myocytes acidosis increases the amplitude of I to as a consequence of a depolarizing shift in the voltage dependence of inactivation. heart; cardiac electrophysiology; subepicardium; subendocardium THE TRANSIENT OUTWARD K ϩ current (I to ) is present throughout the heart of many species; it is found in atrial and ventricular muscle, the sinoatrial node, atrioventricular node, and Purkinje fibers (see Ref. 8 for review). This current is an important determinant of the early repolarization phase of the cardiac action potential, and previous studies have shown a marked heterogeneity in action potential configuration within the ventricular wall that has been ascribed to regional differences in the density of I to . Thus the action potential has a characteristic "spike and dome" morphology in the subepicardium, where I to is prominent, that is less marked in the subendocardium, where I to is small or absent (3, 21). Given its important role in determining the configuration of the cardiac action potential, and the heterogeneity in the density of I to in different regions of the heart, interventions that modulate I to may have important consequences on action potential dispersion and hence on the spread of the action potential through the heart. This may be important in a number of pathologi-cal conditions that have been reported to alter I to , for example, metabolic inhibition (25) and heart failure (5). Antzelevitch et al. (3) reported that simulated ischemia caused a marked depression of the subepicardial action potential, which could be reversed by 4-aminopyridine (4-AP), an inhibitor of I to , but had little effect on the subendocardial action potential. These data suggest that an increase in I to may, at least in part, underlie the changes in action potential configuration recorded during simulated ischemia so that the regional differences in the response to such "ischemia" may reflect regional differences in the density of I to . Because acidosis is known to have marked effects on several membrane currents (for reviews see Refs. 23, 24) and is a major component of both the simulated ischemia described above and of true ischemia, then if acidosis increases I to , this could explain the changes in action potential configuration observed by Antzelevitch et al. (3) and could play a role in altering the electrical activity of the heart during true ischemia. We have, therefore, investigated the effects of acidosis on I to in cells isolated from the subepicardial and subendocardial regions of the rat left ventricle. MATERIALS AND METHODS Cell Isolation Rat ventricular myocytes were isolated as described previously (13). Briefly, adult Wistar rats of either sex were stunned and then killed by cervical dislocation. The heart was rapidly removed and transferred to a beaker containing oxygenated physiological salt solution (PSS; for composition see below) where it was gently massaged to remove excess blood. The aorta was cannulated, and the heart was retrogradely perfused at a constant flow (8 ml · min Ϫ1 · g wet wt Ϫ1 ) with PSS containing Ca 2ϩ (0.75 mM) at 37°C. Once the preparation appeared stable and the coronary vessels had cleared of blood, the perfusion was switched to nominally Ca 2ϩ -free PSS for 4 min. The perfusate was then switched to PSS containing 1 mg/ml collagenase (type II, Worthington), 0.1 mg/ml protease (type XIV, Sigma), and 50 µM Ca 2ϩ . This solution was recirculated to give a total exposure to the enzyme of 10 min. At the end of the perfusion, the heart was cut down and the ventricles dissected free. The ventricles were then cut open, and thin slices of tissue were cut from the subepicardium (Ͻ2 mm from the epicardial surface) and the subendocardium (Ͻ2 mm from the endocardial surface) of the left ventricle using a fine pair of scissors. The slices were cut parallel to the surface of the ventricular wall and along the apex-base axis. The slices from each region were placed in separate conical flasks and gently agitated in enzyme-containing PSS supplemented The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.