Volume-sensitive Tyrosine Kinases Regulate Liver Cell Volume through Effects on Vesicular Trafficking and Membrane Na⁺ Permeability

In liver cells, the influx of Na⁺ mediated by nonselective cation (NSC) channels in the plasma membrane contributes importantly to regulation of cell volume. Under basal conditions, channels are closed; but both physiologic (e.g. insulin) and pathologic (e.g. oxidative stress) stimuli that are known to stimulate tyrosine kinases are associated with large increases in membrane Na⁺ permeability to ∼80 pA/pF or more. Consequently, the purpose of these studies was to evaluate whether volume-sensitive tyrosine kinases mediate cell volume increases through effects on the activity or distribution of NSC channel proteins. In HTC hepatoma cells, decreases in cell volume evoked by hypertonic exposure increased total cellular tyrosine kinase activity ∼20-fold. Moreover, hypertonic exposure (320-400 mosM) was followed after a delay by NSC channel activation and partial recovery of cell volume toward basal values (regulatory volume increase (RVI)). The tyrosine kinase inhibitors genistein and erbstatin prevented both NSC channel activation and RVI. Similarly, hypertonic exposure resulted in an increase in p60^c-src activity, and intracellular dialysis with recombinant p60^c-src led to activation of NSC currents in the absence of an osmolar gradient. Utilizing FM1-43 fluorescence, exposure to hypertonic media caused a rapid increase in the rate of exocytosis of ∼40% (p < 0.01), and genistein inhibited both exocytosis and channel activation. These findings indicate that volume-sensitive increases in p60^c-src and/or related tyrosine kinases play a key role in the regulation of membrane Na⁺ permeability, suggesting that increases in the NSC conductance may be mediated in part through rapid recruitment of a distinct pool of channel-containing vesicles.