Vesicular pH Is Sensitive to Changes in Cell Volume

Abstract

As shown previously, osmotic swelling of proximal renal tubules, vascular smooth muscle cells and of hepatocytes leads to an increase of the pH in acidic cellular compartments. This vesicular alkalinization is reflected by an increase of acridine orange fluorescence intensity and a corresponding alteration of the FITC-dextran fluorescence ratio. The present study has been performed to test whether this phenomenon can be observed in other cell types. Osmotic cell swelling (–30 mOsm) increased acridine orange fluorescence intensity in cardiac myocytes as demonstrated using fluorescence imaging. Microspectrophotometry was used to demonstrate a similar increase in acridine orange fluorescence intensity following treatment with hypotonic solution in MDCK cells, macrophages, brown fat cells, dendritic cells, pancreatic β-cells, alveolar cells and normal and ras oncogene expressing fibroblasts. Swelling of MDCK and L2 cells, induced by a block of K+ channels with Ba2+ or the elevation of extracellular K+, also led to vesicular alkalinization. The uptake of various amino acids in MDCK cells, resulting in cell swelling, similarly increased the acridine orange fluorescence intensity. Further, the acridine orange results were confirmed and quantified with FITC-dextran in brown fat cells, where treatment with hypotonic solution increased the vesicular pH by as much as 0.53 ± 0.04 (n = 4). It is concluded that osmotic swelling of a wide variety of cells increases the pH of acidic cellular compartments, which in turn is expected to modify the respective activity of lysosomal proteases and trafficking of vesicles and receptors.