H+-extrusion by Na+-H+ exchange is localised at the surface sarcolemma in rat ventricular myocytes.

Na+-H+ exchange (NHE) is the principal H+-extrusion mechanism from cardiac cells. We have investigated the spatial location of functionally active NHE in rat isolated ventricular myocytes. Cells were subjected to transient hypertonic shock using 1.5M formamide (Brette et al, Am J Physiol 283; 2002) to disrupt the t-system, and then assayed for NHE-mediated H+-extrusion (using SNARF to monitor pHi recovery following an NH4Cl-prepulse; Hepes-buffered superfusates). Successful detubulation was checked in some cells by membrane staining with di-8-Anneps, and in others by monitoring the electrically evoked Ca2+i-transient (confocal linescan using fluo-3; a sub-sarcolemmal Ca2+i-increase in detubulated myocytes). Both intrinsic buffer capacity and H+-extrusion at a given pHi (7.2-6.2) were unaffected by detubulation, suggesting NHE activity is principally at the sarcolemmal surface. Immunofluorescent staining with NHE1 mouse monoclonal antibody 611775, clone 54 (BD Transduction Laboratories) revealed limited expression at the surface sarcolemma, widespread punctate expression at intracellular sites, but no appearance at t-system sites. We conclude that H+extrusion from the ventricular myocyte is achieved functionally at surface rather than t-system membranes. [ABSTRACT FROM AUTHOR]