Studies on the mechanism of swelling-induced lysosomal alkalinization in vascular smooth muscle cells.

Abstract

Previous studies in renal cells and hepatocytes have shown that cell swelling leads to a rapid and reversible increase in pH in acidic cellular compartments, including lysosomes. Among the consequences are an inhibition of proteolysis. The present study shows that a similar lysosomal alkalinization occurs upon osmotic swelling of vascular smooth muscle cells, as evidenced by acridine orange and fluorescein isothiocyanate fluorescence. Furthermore, we have studied the mechanism underlying lysosomal alkalinization, which had remained unclear. The lysosomal alkalinization was not abolished by inhibition of vacuolar H+-ATPases (100 nM bafilomycin), Cl- channels [100 microM] 5-nitro-2-(3-phenylpropylamino) benzoic acid (NPPB), carbonic anhydrase (100 microM acetazolamide) or Na+/H+ exchange (10 microM HOE 694). The Ca2+ ionophore A23187 (10 microM) led to a slight increase in lysosomal pH, but removal of extracellular Ca2+ and depletion of cellular Ca2+ stores (100 nM thapsigargin) did not appreciably blunt the swelling-induced lysosomal alkalinization. In the presence of bafilomycin the alkalinizing effect of osmotic cell swelling was not reversible, in contrast to that of NH4Cl. In conclusion, osmotic swelling of vascular smooth muscle cells leads to lysosomal alkalinization, presumably in large part through activation of a hydrogen ion leak.