Experimental Studies Of Effects Of Hyposmotic Swelling On Action Potential And Delayed Rectifier Potassium Current (I_k) In Guinea Pig Ventricular Myocytes

Objectives: Swelling of cardiac myocytes occurs during acute myocardial ischemia episode and this is exacerbated after reperfusion. The electro-physiological properties of myocytes alter as a result of the membrane mechanical stretching during the cell swelling. Cell swelling can modulate the function of a number of membrane channels and ion transporters, which may contribute to action potential changes. The effects will lead to changes of the membrane electrical activity and may contribute to arrhythmogenesis under pathological conditions. Single acute isolated guinea pig ventricular myocytes were superfused with hyposmotic solution to simulate this situation. The aims of the present study were as follows : 1. Observing the changes of the action potential of single ventricular myocytes in hyposmotic solution, and discussing the arrhythmogenesis mechanisms underlying hyposmotic swelling. 2. Investigating channel kinetic properties of the delayed rectifier potassium current (IK) and its two subtypes, which is important for determining the action potential depolarization configuration. 3. Comparing the different effects of hyposmotic swelling on two subtypes (1, and 1) of IK, in order to partly explain the ionic mechanisms underlying ischemia- and reperfusion-induced arrhythmogenesis.Methods: Single guinea pig ventricular myocytes were obtained by retrogradely perfusing with collagenase through the aorta. Ventricular myocytes with integrated membrane and clear cross striations were selected for further experiments under the inverted microscope. Cells were continuously superfused with isosmotic Tyrode's solution at a flow-rate of Iml/min. The whole-cell recording configuration of the patch clamp technique was formed after giga-sealing, membrane rupturing and capacitance and series resistance compensation. Action potential underlying the basic cycle length (BCL) of 600ms, 1000ms and 3000ms were recorded with a pulse stimulus of 3ms duration and twofold threshold. Measurements were made of the following: action potential amplitude (APA), resting membrane potential (RMP) and action potential duration measured at 30%, 50%, and 90% of repolarization (APD30, APD50 and APD90). The above parameters were measured again after the superfusate was switched to hyposmotic solution. IK of ventricular myocytes was measured by step depolarizing to +60mV from a holding potential of-40mV, which the pulseduration was 100ms (sampling frequency 2.5kHz) and 3000ms (sampling frequency 0.5kHz), respectively. IK was recorded again while cells were exposed to hyposmotic superfusate. The amplitude of IK and Iuil were measured. The I-V relationships of the current was measured over the range-40 to +60mV, before and during cell swelling.Results: 1. The hyposmotic cell swelling had no effect on APA, RMP and the threshold of the ventricular myocytes. 2. APD underlying three BCL was significantly shortened under hyposmotic swelling condition, where shortening of APD50 and APD90 was especially notable, furthermore physiological frequency dependency of APD disappeared. 3. Hyposmotic swelling caused a parallel increase in peak amplitude and tail current amplitude of 1 , thus the ratio of IiCi,uiAci ePt constant; though it caused a decrease in current amplitude of 1 and the ratio of l increased. 4. The I-V relationships of IK and Itail shifted to upper left under hyposmotic swelling condition. Cell swelling caused an increase in outward current flowing at all membrane potentials in the range +10 to +60mV, indicative of an increase in IK. 5. Hyposmotic swelling caused a decrease of deactivation time constant ( T ) of IK, suggesting that swelling may accelerate the activation and deactivation of IK.Conclusion: 1. Hyposmotic swelling did not affect the excitability and conductivity of ventricular myocytes. 2. The shortening of APD especially APD50 and APD90 in the state of hyposmotic swelling, suggested that hyposmotic swelling mainly affected the plateau and 3 phase depolarization of action potential, which may induce the occurrence of...