Regulation of cation channels in liver cells by intracellular calcium and protein kinase C.
The regulation of Ca(2+)-permeant cation channels in HTC hepatoma cells was investigated using patch clamp and fluorescence techniques. In intact cells, exposure to nucleotide analogues ATP, uridine 5'-triphosphate (UTP), and adenosine 5'-O-(3-thiotriphosphate) (ATP gamma S) caused transient opening of channels with linear conductances of approximately 18 and approximately 28 pS. Channels were permeable to Na+, K+, and Ca2+ and carried inward (depolarizing) current at the resting potential. Exposure to thapsigargin to increase cytosolic Ca2+ concentration ([Ca2+]i) opened similar channels, suggesting that opening is stimulated by a rise in [Ca2+]i. In subconfluent monolayers, ATP increased [Ca2+]i with half-maximal effects at approximately 7.4 microM; at 10(-4) M, the peak increase in [Ca2+]i was ATP > UTP > ATP gamma S >> 2-methylthioadenosine 5'-triphosphate, alpha,beta-methyleneadenosine 5'-triphosphate, and adenosine. The relative potency suggests that the effects are mediated by 5'-nucleotide receptors. In excised inside-out patches, channels were not activated by myo-inositol 1,4,5-trisphosphate (50-100 microM) or myo-inositol 1,3,4,5-trisphosphate (20 microM) but opened after increases in Ca2+ to greater than approximately 250 nM, consistent with a direct role for Ca2+ in channel opening. In intact cells, channel opening was followed by a prolonged refractory period. Protein kinase C appears to contribute by inhibition of the ATP-stimulated [Ca2+]i response and by direct inhibitory effects on the channel. These findings indicate that extracellular ATP leads to modulation of liver cell cation channels through activation of 5'-nucleotide receptors and are consistent with a model in which transient opening of channels is stimulated by a rise in [Ca2+]i and subsequent closure is mediated by protein kinase C-dependent pathways.
Fitz
J G
JG
Department of Medicine, Duke University Medical Center, Durham, North Carolina 27710.
Sostman
A H
AH
Middleton
J P
JP
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002412:Cations / Q000378:metabolism*; D005110:Extracellular Space / Q000378:metabolism; D007425:Intracellular Membranes / Q000378:metabolism*; D007473:Ion Channels / Q000037:antagonists & inhibitors / Q000187:drug effects / Q000378:metabolism*; D008099:Liver / Q000166:cytology / Q000378:metabolism*; D009711:Nucleotides / Q000378:metabolism / Q000494:pharmacology; D009994:Osmolar Concentration; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D051381:Rats; D014407:Tumor Cells, Cultured
D002412:Cations; D007473:Ion Channels; D009711:Nucleotides; D011493:Protein Kinase C; D002118:Calcium