DEPRIVATION OF MEMBRANE CHOLESTEROL DEPRESSESCAMP-DEPENDENT ENHANCEMENT OF L-TYPE CACURRENT IN RABBIT VENTRICULAR MYOCYTES

摘要

Cholesterol is a primary structural component in caveolae that are involved in various signal transductions. We applied methyl-p-cyclodetrin (MbCD) that deprives membrane cholesterol to isolated rabbit ventricular myocytes intracellularly by whole-cell pipettes to study the role of cholesterol in cAMP-dependent increase of L-type Ca~(2+) current (I_(Ca,L))-^sWhole-cell I_(Ca,L)s were recorded by applying double pulses in the presence of Na~+, Cs~+ and 1.8 mM Ca~(2+) in test solutions and CsCl and BAPTA in pipette solutions. Isoproterenol (luM, ISO), forskolin (lOuM, FSK) and dibutyl-cAMP (3 mM, cAMP) together with IBMX were applied 10 min after dialysis of pipette solutions. ISO changed the maximal amplitude of I_(Ca,L) to 2.9+-0.1 (n=21, normalized to the control amplitude) in control and to 1.0+-0.2 (n=12) in the presence of 30 mM MbCD in the pipette solution, thereby the density of I_(Ca,L) before the drug application was nearly the same with control; FSK to 3.0+-0.4 (n=8) in control and 1.3+-0.2 (n=8) in MbCD; cAMP + IBMX to 3.1+-0.4 (n=9) in control and to 1.4+-0.2 (n=9) in MbCD. These results suggest that L-type Ca channels densely localize in caveolae in ventricular myocytes and that cholesterol in the caveolar membrane is indispensable for their functional modulation by protein kinase A-mediated phosphorylation.