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, JG; Sostman, AH; Middleton, JP
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