| 8661992 |
Tauc M, Bidet M, Poujeol P: Chloride currents activated by calcitonin and cAMP in primary cultures of rabbit distal convoluted tubule. J Membr Biol. 1996 Apr;150(3):255-73.
Chloride (Cl-) conductances were studied in primary cultures of the bright part of rabbit distal convoluted tubule (DCTb) by the whole cell patch clamp technique. The bath solution (33 degrees C) contained (in mM): 140 NaCl, 1 CaCl2, 10 N-2-hydroxy-ethylpiperazine-N'-2-ethanesulfonic acid (HEPES), pH 7.4 and the pipette solution 140 N-methyl-D-glucamine (NMDG)-Cl, 5 MgATP, 1 ethylene-glycol-bis (b-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA), 10 HEPES, pH 7.4. We identified a Cl- current activated by 10 (-5) M forskolin, 10 (-3) M 8-bromo adenosine 3',5'-cyclic monophophosphate (8 Br-cAMP), 10 (-6) M phorbol 12-myristate 13-acetate (PMA), 10 (-3) M intracellular adenosine 3',5'-cyclic monophophosphate (cAMP) and 10 (-7) M calcitonin. The current-voltage relationship was linear and the relative ion selectivity was Br- > Cl- > > I- > glutamate. This current was inhibited by 10 (-3) M diphenylamine-2-carboxylate (DPC) and 10 (-4) M 5-nitro-2-(3-phenylpropylamino)-benzoate (NPPB) and was insensitive to 10 (-3) M 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS). These characteristics are similar to those described for the cystic fibrosis transmembrane conductance regulator (CFTR) Cl- conductance. In a few cases, forskolin and calcitonin induced an outwardly rectifying Cl- current blocked by DIDS. To determine the exact location of the Cl- conductance 6-methoxy-1-(3-sulfonatopropyl) quinolinium (SPQ) fluorescence experiments were carried out. Cultures seeded on collagen-coated permeable filters were loaded overnight with 5 mM SPQ and the emitted fluorescence analyzed by laser-scan cytometry. Cl- removal from the apical solution induced a Cl- efflux which was stimulated by 10 (-5) M forskolin, 10 (-7) calcitonin and inhibited by 10 (-5) M NPPB. In 140 mM NaBr, forskolin stimulated an apical Br- influx through the Cl- pathway. Forskolin and calcitonin had no effect on the basolateral Cl- permeability. Thus in DCTb cultured cells, exposure to calcitonin activates a Cl- conductance in the apical membrane through a cAMP-dependent mechanism. |
82(1,1,1,2) |