Volume-activated chloride channels in mice Leydig cells.
Production and secretion of testosterone in Leydig cells are mainly controlled by the luteinizing hormone (LH). Biochemical evidences suggest that the activity of Cl(-) ions can modulate the steroidogenic process, but the specific ion channels involved are not known. Here, we extend the characterization of Cl(-) channels in mice Leydig cells (50-60 days old) by describing volume-activated Cl(-) currents (I(Cl,swell)). The amplitude of I(Cl,swell) is dependent on the osmotic gradient across the cell membrane, with an apparent EC(50) of approximately 75 mOsm. These currents display the typical biophysical signature of volume-activated anion channels (VRAC): dependence on intracellular ATP, outward rectification, inactivation at positive potentials, and selectivity sequence (I(- )> Cl(- )> F(-)). Staurosporine (200 nM) did not block the activation of I(Cl,swell). The block induced by 5-nitro-2-(3-phenylpropylamino) benzoic acid (NPPB; 128 microM), SITS (200 microM), ATP (500 microM), pyridoxal-phosphate-6-azophenyl-2',4'-disulfonate (PPADS; 100 miccroM), and Suramin (10 microM) were described by the permeant blocker model with apparent dissociation constant at 0 mV K(do) and fractional distance of the binding site (delta) of 334 microM and 47 %, 880 microM and 35 %, 2,100 microM and 49%, 188 microM and 27%, and 66.5 microM and 49%, respectively. These numbers were derived from the peak value of the currents. We conclude that I(Cl,swell) in Leydig cells are activated independently of purinergic stimulation, that Suramin and PPADS block these currents by a direct interaction with VRAC and that ATP is able to permeate this channel.
Department of Physiology, University of São Paulo, Av. Bandeirantes 3900, 14049-900, Ribeirão Preto/SP, Brazil.
Luiz Artur
LA
Poletto Chaves
Wamberto Antonio
WA
Varanda
D012856:4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D002463:Cell Membrane Permeability; D048429:Cell Size; D018118:Chloride Channels / Q000037:antagonists & inhibitors / Q000378:metabolism*; D002712:Chlorides / Q000378:metabolism*; D000242:Cyclic AMP / Q000378:metabolism; D066298:In Vitro Techniques; D015640:Ion Channel Gating / Q000187:drug effects; D007700:Kinetics; D007985:Leydig Cells / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D008297:Male; D008564:Membrane Potentials; D051379:Mice; D009579:Nitrobenzoates / Q000494:pharmacology; D009997:Osmotic Pressure; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D011494:Protein Kinases / Q000378:metabolism; D011732:Pyridoxal Phosphate / Q000031:analogs & derivatives / Q000494:pharmacology; D019311:Staurosporine / Q000494:pharmacology; D013498:Suramin / Q000494:pharmacology
D018118:Chloride Channels; D002712:Chlorides; D009579:Nitrobenzoates; D047428:Protein Kinase Inhibitors; C077792:pyridoxal phosphate-6-azophenyl-2',4'-disulfonic acid; D012856:4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid; C058176:5-nitro-2-(3-phenylpropylamino)benzoic acid; D011732:Pyridoxal Phosphate; D013498:Suramin; D000255:Adenosine Triphosphate; D000242:Cyclic AMP; D011494:Protein Kinases; D019311:Staurosporine