Comparison of Na +/Ca 2+ exchanger current and of its response to isoproterenol between acutely isolated and short-term cultured adult ventricular myocytes

Abstract

The Na +/Ca 2+ exchanger protein is present in the cell membrane of many tissue types and plays key roles in Ca 2+ homeostasis, excitation–contraction coupling, and generation of electrical activity in the heart. The use of adult ventricular myocyte cell culture is important to molecular biological approaches to study the roles and modulation of the cardiac Na +/Ca 2+ exchanger. Therefore, we characterised the functional expression of the exchanger in adult guinea-pig ventricular myocytes maintained in short-term culture (for 4 days) and compared the response of ionic current ( I NaCa) carried by the exchanger from acutely isolated and Day 4 cells to β-adrenoceptor activation with isoproterenol (ISO). Functional activity of the exchanger was assessed by measuring I NaCa using whole cell patch clamp, under selective recording conditions. I NaCa amplitude measured at both +60 and −100 mV declined significantly by Day 1 of cell culture, showing a further small decline by Day 4. However, cell surface area (assessed by measuring membrane capacitance) also declined over this time-frame. I NaCa normalised to membrane capacitance ( I NaCa density) did not differ significantly between acutely isolated and cells cultured for 4 days. However, although ISO (1 μM) increased I NaCa in acutely isolated myocytes, it exerted no significant effect on I NaCa from Day 4 cells. This was not due to an inherent inability of these cells to respond to ISO, as L-type calcium current amplitude from Day 4 cells was increased by ISO to a similar extent as that from acutely isolated cells. Our data suggest that the functional expression of the Na/Ca exchanger is well maintained during short-term culture of adult ventricular myocytes. The lack of response to ISO of I NaCa from Day 4 cells suggests: (a) that, despite a well-maintained I NaCa density, cultured adult myocytes may not necessarily be suitable for studies of exchanger modulation by some agonists and (b) that there may exist subtle differences between β-adrenergic regulation of the exchanger protein and of L-type Ca channels.