Ca++ distribution after Na+ pump inhibition in cultured neonatal rat myocardial cells

The influence of inhibition of the Na+ pump, with secondary stimulation of Na+-Ca++ exchange, on cellular Ca++ distribution is examined using the on-line scintillation disk technique and cultured neonatal rat myocardial cells. Under control conditions, La+++ displaced 78.1 +/- 1.13% (SEM) of the total cell associated 45Ca. Application of 1 mm ouabain or reduction of [K+]o to 0.5 mM resulted in a net increase of 10.6 +/- 1.3% and 13.8 +/- 2%, respectively, in total cell-associated Ca++. Of this added 45Ca, 75.9 +/- 2.7% and 78.4 +/- 2.1%, respectively remained La+++-displaceable. The 45Ca-binding characteristics of isolated sarcolemma, prepared from the cultured neonatal rat myocardial cells using the gas dissection technique, were examined. When treated with either ouabain or low [K+]o solutions, sarcolemmal 45Ca binding did not change. This result indicates that functional, intact tissue is necessary to observe the Ca++ increase. Treatment of the cells with verapamil before and during ouabain exposure failed to inhibit the ouabain-induced increase in cell-associated 45Ca. The evidence indicates that inhibition of the Na+-pump, and secondary stimulation of Na+-Ca++ exchange, result in a net increase of 11-15% in cell-associated Ca++, 78% of which remains La+++-displaceable and is, therefore, localized to the sarcolemma-glycocalyx complex.