Effect Of Actin Microfilament On Potassium Current In Guinea-pig Gastric Antral Myocytes

The cytoskeleton is an intracellular superstructure that consists of microfilaments of actin and associated proteins, microtubules, and intermediate filaments. The actin microfilament, in particular, is involved in structural support and a functional role in cell motility. Recent evidence indicated that the actin-based cytosleleton was involved in the control of ion channel activity across the plasma membrane of different cell type. In neuron cells, cardisc mucle cells and renal tuble epithelial cells, the effect of cytoskeleton on potassium current and the underlyling mechanism have been found. However, such experiments have not been done in gastrointestinal smooth muscle cells.The aim of this study was to explain the effect of actin microfilament on calcium-activated potassium current(IK(Ca)) and delayed rectifier potassium current(IK(V)) in guinea-pig gastric antral myocytes.In this study, we applied 0.1% type II collagenase to dissociate gastric antral circular smooth muscle cells in guinea-pig. The majority of the freshly dissociated mucle cells possessed an intact structure. By the whole-cell configuration of patch-clamp techniques, in this study, the relationship between actin microfilament and outward potassium current under normal and membrane stretch condition had been investigated. All values were expressed as x s, statistical significance was evaluated by t-test. Results are as follows:1. Using the conventional whole cell mode, the membrane potential was clamped at -60mV, and command pulse from -40mV to +100mV for 440ms with a 20mV increment at 10 second intervals. Ca2+-activated potassium current( IK(Ca)) was elicitedby the step depolarization with the pipette solution containing egtazic acid(EGTA) 0.1 mM. Under the conventional whole cell, the mean amplitude of /K(Ca) at +60mV was 826.6 72.6pA(n=15).2. When the membrane potential was held at -60m V, 20umol/L cytochalasin-B (Cyt-B, an actin microfilament disruptor) increased /K(Ca)to 133.4% 7.8% at +60mV. There is significant difference between control group and Cyt-B group (p<0.01, n=15).3. When the membrane potential was held at ~60mV, 20umol/L phalloidin(an actrin microfilament stabilizer) inhibited /K(Ca) to 71.8% 3.3% at +60mV. There is significant difference between control group and phalloidin group (/?<0.01, n=15).4. The membrane potential was clamped at -60mV. and command pulse from -40mV to +80mV for 440ms with a 20mV increment at 10 second intervals. Using the conventional whole cell mode, delayed rectifier potassium current(/K(V)) was elicited by the step depolarization with the pipette solution containing egtazic acid(EGTA) lOmM. Under the conventional whole cell mode, the mean amplitude of /K0.05, n=15).10. In the condition of 20umol/L phalloidin in pipette solution, hyposmotic membrane stretch increased /K(ca) by 50.6% 9.7% at +60mV. There was no significant difference betwee...