Role of cell volume and protein kinase C in regulation of a Cl- conductance in single proximal tubule cells of Rana temporaria.
1. The whole-cell patch clamp technique was used to investigate Cl- currents in single proximal tubule cells isolated from kidneys of Rana temporaria. 2. Immediately following establishment of the whole-cell clamp, the Cl- conductance (gCl) of the cells was low. However, with 2 mM ATP in the pipette there was a time-dependent activation of gCl. Such activation was inhibited when the bath contained a hypertonic Ringer solution. 3. The Cl- conductance was not directly dependent on cell volume; gCl increased with hypotonic shock and decreased with hypertonic shock, but only in the presence of ATP. 4. Activation of gCl by ATP was dependent on extracellular Ca2+; however, the conductance was not directly Ca2+ sensitive. Activation was inhibited by Gd3+, which also had a direct inhibitory action on gCl. 5. Inhibition of protein kinase C (PKC), by 10 microM PKC pseudo-substrate (PKC-ps), completely abolished the ATP-dependent activation of gCl, while stimulation of PKC, by the PKC activator 4 beta-phorbol 12-myristate, 13-acetate (PMA), increased the degree of activation typically observed with ATP. 6. We propose that gCl is activated by PKC-mediated phosphorylation and plays a role in volume regulation of the cells.
Robson
L
L
Department of Physiology, Medical School, Leeds University, UK.
Hunter
M
M
D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000494:pharmacology; D048429:Cell Size / Q000502:physiology; D018118:Chloride Channels / Q000187:drug effects / Q000378:metabolism*; D004789:Enzyme Activation / Q000187:drug effects; D005682:Gadolinium / Q000494:pharmacology; D066298:In Vitro Techniques; D015759:Ionomycin / Q000494:pharmacology; D007687:Kidney Tubules, Proximal / Q000166:cytology / Q000201:enzymology / Q000378:metabolism*; D009994:Osmolar Concentration; D018408:Patch-Clamp Techniques; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000502:physiology*; D011896:Rana temporaria; D015290:Second Messenger Systems / Q000502:physiology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D018118:Chloride Channels; D015759:Ionomycin; D000255:Adenosine Triphosphate; D005682:Gadolinium; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate; D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid; D002118:Calcium