DPC BLOCKADE OF TRANSEPITHELIAL CHLORIDE ABSORPTION AND SINGLE ANION CHANNELS IN TELEOST URINARY-BLADDER

The columnar cell epithelium of the euryhaline goby (Gillichthys mirab ilis) urinary bladder actively absorbs NaCl from the lumen, thereby dr iving water transport and reducing water loss to the hypertonic extern al environment. Transcellular transport of Cl- involves apical membran e entry via Na+-coupled cotransport driven by the Na+ electrochemical gradient and subsequent basolateral membrane exit. An anion channel in the basolateral cell membrane of columnar epithelial cells was identi fied using patch-clamp technique. This channel may be one avenue for b asolateral Cl- exit from the urinary bladder columnar cell. Single-cha nnel conductance (G(c)) of channels in excised, inside-out membrane pa tches was approximately 75 pS in symmetrical solutions containing 140 mM Cl-. The channel was selective to Cl- over other anions [Cl- > 2-(N -morpholino)ethanesulfonic acid (MES) > F- almost-equal-to Br- almost- equal-to I- > NO3- almost-equal-to SO42-). Channel activity, expressed as the open probability (P(o)), was voltage dependent in the physiolo gical range of membrane potential, with membrane depolarization increa sing P(o). Decreasing the pH of the solution bathing the cytoplasmic f ace of the membrane patch over the range 8.4-6.0 reduced P(o). There w as no effect of pH on either G(c) or ionic selectivity. Radiochloride flux technique was also applied to intact columnar epithelial cell she ets to relate anion channel activity to macroscopic transcellular tran sport. Serosal exposure to the anion channel blocker diphenylamine-2-c arboxylic acid (DPC, 30 muM to 3 mM) reduced and abolished transcellul ar radiochloride fluxes and net Cl- absorption across short-circuited tissues in a dose-dependent fashion. DPC addition (10 muM to 1 mM) to the solution bathing the cytoplasmic face of excised, inside-out membr ane patches reduced P(o) in a dose-dependent manner and had no effect on G(c). These parallel findings of DPC blockade on intact epithelia a nd on single anion channels support the notion that this anion channel is a basolateral membrane component of the pathway for Cl- movement i n transcellular Cl- absorption.