A GTP-binding protein activates chloride channels in a renal epithelium.
Although G proteins have been shown to regulate cation channels, regulation of Cl- channels by G proteins has not been demonstrated directly. Accordingly, the objective of this study was to examine whether a G protein regulates Cl- channels in the apical membrane of rabbit kidney CCD cells grown in culture. Previous studies showed that this channel is activated by adenosine and protein kinase C and has a single channel conductance of 305 picosiemens. The PCl-:PNa+ is 9:1 and the PCl-:PHCO3- is 2:1 (Schwiebert, E.M., Light, D.B., Dietl, P., Fejes-Toth, G., Naray-Fejes-Toth, A., and Stanton, B. (1990) Kidney Int. 37,216). In the present study, Cl- channels in the apical membrane of CCD cells were studied by the patch clamp technique. GTP and guanosine 5'-O(3-thiophosphate) (GTP gamma S), a nonhydrolyzable analog of GTP, increased the single channel open probability (Po). In contrast, guanosine 5'-O-(2-thiophosphate), a nonhydrolyzable analog of GDP, and pertussis toxin (PTX) decreased the Po. GTP gamma S, but not GTP, reversed PTX inhibition of the channel. The alpha i-3-subunit of Gi increased the Po in both untreated and PTX-treated membrane patches. Because GTP gamma S activated the Cl- channel in the presence of H8, a protein kinase inhibitor, we conclude that the G protein does not activate the channel by stimulating a protein kinase. Thus, a PTX-sensitive G protein activates a Cl- channel in the apical membrane of renal CCD cells.
Schwiebert
E M
EM
Department of Physiology, Dartmouth Medical School, Hanover, New Hampshire 03755.
Light
D B
DB
Fejes-Toth
G
G
Naray-Fejes-Toth
A
A
Stanton
B A
BA
D000241:Adenosine / Q000494:pharmacology; D000818:Animals; D002478:Cells, Cultured; D018118:Chloride Channels; D002712:Chlorides / Q000502:physiology*; D004553:Electric Conductivity; D004848:Epithelium / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000502:physiology*; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D006153:Guanosine Diphosphate / Q000031:analogs & derivatives / Q000494:pharmacology; D006160:Guanosine Triphosphate / Q000031:analogs & derivatives / Q000494:pharmacology; D007684:Kidney Tubules / Q000502:physiology*; D007685:Kidney Tubules, Collecting / Q000502:physiology*; D008565:Membrane Proteins / Q000502:physiology*; D037342:Pertussis Toxin; D011493:Protein Kinase C / Q000378:metabolism; D011817:Rabbits; D013873:Thionucleotides / Q000494:pharmacology; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
D018118:Chloride Channels; D002712:Chlorides; D008565:Membrane Proteins; D013873:Thionucleotides; D010566:Virulence Factors, Bordetella; D006153:Guanosine Diphosphate; D016244:Guanosine 5'-O-(3-Thiotriphosphate); C023427:guanosine 5'-O-(2-thiodiphosphate); D006160:Guanosine Triphosphate; D037342:Pertussis Toxin; D011493:Protein Kinase C; D019204:GTP-Binding Proteins; D000241:Adenosine