Volume-sensitive chloride current activated by hyposmotic swelling in antral gastric myocytes of the guinea-pig

Abstract

 The characteristics of volume-sensitive chloride current (I _Cl) induced by osmotic cell swelling were studied using the whole-cell patch-clamp technique and cell diameters of antral circular guinea-pig myocytes were simultaneously measured under isosmotic and hyposmotic conditions by using a video image analysis system. At –60 mV, osmotic cell swelling (200 mosmol/l) activated a sustained inward current. Instantaneous current/voltage (I-V) relations obtained by step voltage pulses showed an outward rectification. At potentials above +40 mV, the current exhibited time-dependent decay. The outward current amplitude was decreased and the reversal potential was shifted to more positive potentials by replacement of external Cl^– with gluconate^–, while the current amplitude and the I/V relation were not affected by replacing extracellular Na⁺ with N-methyl-D-glucamine. The anion permeability sequence of the swelling-induced current was I^– (1.80) > Br^– (1.31) > Cl^– (1) > F^– (0.85) > gluconate^– (0.18). The I _Cl was effectively inhibited by the Cl^– channel blockers, 4,4′-diisothiocyanatostilbene-2,2’-disulphonic acid (DIDS, 100 μM), and niflumic acid (10 μM). DIDS suppressed outward current more effectively than inward current. Also, the I _Cl was dose-dependently inhibited by arachidonic acid, an unsaturated fatty acid and also inhibited by other unsaturated fatty acids (linoleic acid and oleic acid) but not by stearic acid, a saturated fatty acid. The inhibitory effect of arachidonic acid on I _Cl was not prevented by indomethacin, a cyclo-oxygenase inhibitor and chelerythrine, a protein kinase C inhibitor. Under whole-cell patch-clamp conditions, the cell diameter was continuously measured using video image analysis, which reflects the change in cell volume. A hyposmotic-stimulation-induced increase of cell diameter was followed by I _Cl activation. In intact single gastric myocytes, relatively severe hyposmotic (176 mosmol/l) superfusing solution increased the cell diameter and the pretreatment with DIDS or with niflumic acid significantly potentiated the above effect of hyposmotic superfusion. These results suggest that volume-sensitive outwardly rectifying chloride current (I _Cl) is present in guinea-pig gastric myocyte and the I _Cl may play a role in smooth muscle cell volume regulation.