Mechanisms of receptor-mediated Ca2+ signaling in rat hepatocytes.
The Ca2+ signal observed in individual fura-2-loaded hepatocytes stimulated with the alpha 1-adrenergic agonist phenylephrine consisted of a variable latency period, a rapid biphasic increase in the cytosolic free Ca2+, followed by a period of maintained elevated cytosolic Ca2+ (plateau phase) that depended on the continued presence of both agonist and external Ca2+. Microinjection of guanosine-5'-O-(3-thiophosphate) elicited a Ca2+ transient with the same basic features. The Ca2+ transient resulting from microinjecting inositol 1,4,5-trisphosphate (Ins-1,4,5-P3) occurred with essentially no latency period and consisted of a rapid spike that decayed back to preinjection levels within 15 s. Microinjection of inositol 1,4,5-trisphosphorothioate (thio-IP3), a nonmetabolizable analog of Ins-1,4,5-P3, elicited a Ca2+ transient that was initially identical to that observed with Ins-1,4,5-P3, except that the cytosolic Ca2+ remained elevated. The maintained thio-IP3-induced Ca2+ increase was dependent on the presence of external Ca2+, suggesting an activation of Ca2+ influx. Reintroduction of external Ca2+ in the presence of 5 microM phenylephrine to Ca(2+)-depleted cells resulted in a 2-fold greater rate of rise in the cytosolic Ca2+ compared to the rate observed upon Ca2+ addition to cells Ca(2+)-depleted by preatement with thapsigargin. The rate of Ca2+ rise upon Ca2+ addition to cells microinjected with thio-IP3 was similar to that observed with phenylephrine. Coinjection of the cells with thio-IP3 plus heparin reduced the rate of Ca2+ rise upon Ca2+ addition to that observed in thapsigargin-treated cells. These data indicate that the mechanism responsible for receptor-mediated stimulation of Ca2+ entry into hepatocytes involves not only capacitative Ca2+ entry but also an additional component mediated directly by Ins-1,4,5-P3.
Hansen
C A
CA
Department of Biochemistry and Biophysics, University of Pennsylvania School of Medicine, Philadelphia 19104.
Yang
L J
LJ
Williamson
J R
JR
D000818:Animals; D002478:Cells, Cultured; D003600:Cytosol / Q000378:metabolism; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000494:pharmacology; D007295:Inositol Phosphates / Q000378:metabolism; D007700:Kinetics; D008099:Liver / Q000378:metabolism*; D008297:Male; D008845:Microinjections; D010656:Phenylephrine / Q000494:pharmacology; D051381:Rats; D011919:Rats, Inbred Strains; D011956:Receptors, Cell Surface / Q000378:metabolism*; D015398:Signal Transduction
D007295:Inositol Phosphates; D011956:Receptors, Cell Surface; D010656:Phenylephrine; D016244:Guanosine 5'-O-(3-Thiotriphosphate)