FIGURE 6.

Effect of P2X4 dominant-negative mutant overexpression on 11% hypotonicity-induced VSOR Cl⁻ currents and Ca_o²⁺ and ATP on RVD. A, representative experiments showing the time course of VSOR Cl⁻ currents evoked by 11% hypotonicity in cells overexpressing the P2X4 K313A dominant-negative construct in the presence of 2 mm external Ca²⁺ with (●) or without (○) 10 μm ATP. Currents were measured at 80 mV every 7 s and normalized by cell capacitance. B, summary of the data presented in A for normalized maximal currents at 80 mV (left axis) and half-time for current onset (right axis) obtained from n = 5–6 independent experiments. C, cell volume was monitored in single HTC cells loaded with calcein. Representative experiment in which cell swelling was induced by exposure to 33% hypotonicity during the time depicted by the bar is shown. At the time indicated by the arrow, the solution was switched to a NMDG-Cl (replacing all NaCl) containing 10 μm gramicidin solution with (●) or without Ca_o²⁺ (○). D, summary of the rate constants obtained by adjusting the fall in cell volume in each case to a single decreasing exponential from experiments depicted in C. Results shown are averages of 15–20 cells measured in single coverslips, n = 3–4 independent experiments. E, cell volume was monitored in single HTC cells loaded with calcein. Representative experiment in which cell swelling was induced by exposure to 11% hypotonicity during the time depicted by the bar is shown. At the time indicated by the arrow, the solution was switched to a NMDG-Cl (replacing all NaCl) containing 10 μm gramicidin solution with (○) or without 10 mm ATP (●) in the presence of 2 mm Ca_o²⁺. F, summary of the rate constants obtained by adjusting the fall in cell volume in each case to a single decreasing exponential from experiments depicted in E. Results shown are averages of 15–20 cells measured in single coverslips, n = 3–4 independent experiments. *, p < 0.05 compared with control conditions.