Decreased inward rectifier current in adult rabbit ventricular myocytes maintained in primary culture: a single-channel study

Objective: Regulation of ion channel function in heart has been shown to be affected by changes in the cellular environment. Recently it was shown tha t rabbit ventricular myocytes kept in primary culture, show a strong reduct ion in inward rectifier current (I-KI). The aim of the present study was to elucidate the mechanism underlying this decrease in I-KI, using single-cha nnel measurements. In addition, we studied the effects of primary culture o n the ATP-regulated K+ (K.ATP) channel, also a member of the inwardly recti fying K+ channel family. Methods: Adult rabbit ventricular myocytes were cu ltured for up to 3 days in Ham's F-10 medium complemented with 18 rabbit se rum-and 5% glutamine. I-KI and K.ATP channel activity was studied in the in side-out patch configuration of the patch-clamp technique with equimolar K concentrations (140 mM K+) on the intra- and extracellular side. Single ch annel characteristics were determined at various times during culture and c ompared to those present in freshly isolated myocytes. Results: I-KI channe ls in freshly isolated myocytes (day 0) had a single-channel conductance of 56.1+/-2.5 pS (mean+/-SEM) and an open probability of 0.64+/-0.05 (mean+/- SEM). Neither the single-channel conductance nor the open probability (P-o) underwent significant changes during culture. The mean number of channels per patch, however, was drastically reduced from 1.2+/-0.3 (mean+/-SEM) at day 0 to 0.17+/-0.06 at day three. K.ATP channel density and open probabili ty, on the other hand, were both increased with an optimum at day two. P-o increased from 0.27+/-0.06 at day 0 to 0.63+/-0.06 at day three. The mean n umber of channels per patch was 2.29+/-0.57 and 3.25+/-0.48 at days 0 and 3 respectively. The unitary current amplitude at -50 mV remained unchanged, suggesting no change in the K.ATP single-channel conductance. Conclusions: The decrease in I-KI in rabbit ventricular myocytes as has been observed du ring primary culture is the result of a reduction in the number of active c hannels and not of altered kinetic or conductive channel properties. The in crease in K.ATP channel activity under the same conditions suggests that ge ne expression of both channel types is differently regulated. (C) 1999 Else vier Science B.V. All rights reserved.