P2-purinoceptor regulation of surfactant phosphatidylcholine secretion. Relative roles of calcium and protein kinase C.
Ca2+ and protein kinase C have both been proposed as intracellular signals for subsequent phosphatidylcholine secretion by alveolar Type II cells. We have determined the relative roles of Ca2+ and protein kinase C in regulating surfactant phosphatidylcholine secretion by utilizing exogenous ATP and the phorbol ester TPA (12-O-tetradecanoylphorbol 13-acetate) as secretagogues, along with MAPTAM to chelate intracellular Ca2+ and sphingosine to inhibit endogenous protein kinase C. Exposure of Type II cells to the P2-purinoceptor agonist, ATP, results in a dose-dependent increase in surfactant phosphatidylcholine secretion from isolated alveolar Type II cells with an EC50 (concn. producing 50% of maximal response) of 2 microM. Administration of exogenous ATP to Type II cells also results in a dose-dependent increase in inositol trisphosphate production, Ca2+ mobilization and [3H]phorbol 12,13-dibutyrate ([3H]PDBu) binding as a measure of protein kinase C translocation. The EC50 in each case is 1-5 microM, indicating association of these events with surfactant phosphatidylcholine secretion. Loading Type II cells with non-hydrolysable GTP analogue (GTP[S]) inhibited ATP-induced Ca2+ mobilization, supporting the hypothesis that Type II cell P2-purinoceptors are coupled to phospholipase C via a GTP-binding protein. The ATP-induced elevation of cytosolic Ca2+ was also inhibited by MAPTAM (a cell-permeant EGTA analogue) by 90%, but MAPTAM was without effect on surfactant phosphatidylcholine secretion induced by ATP. Sphingosine inhibited both ATP- and TPA-induced surfactant phosphatidylcholine secretion as well as [3H]PDBu binding with a similar IC50 (concn. producing 50% of maximal inhibition) (10 microM). Sphingosine did not affect surfactant phosphatidylcholine secretion induced by terbutaline and did not have a significant effect on Ca2+ mobilization induced by exogenous ATP. These results are consistent with a prominent role for protein kinase C in regulation of P2-purinoceptor-induced surfactant phosphatidylcholine secretion, and indicate that Ca2+ mobilization is not a necessary step for ATP-induced surfactant phosphatidylcholine secretion.
Rice
W R
WR
University of Cincinnati, College of Medicine, OH 45267-0541.
Dorn
C C
CC
Singleton
F M
FM
D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D002118:Calcium / Q000502:physiology*; D004075:Diglycerides / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000378:metabolism; D007295:Inositol Phosphates / Q000378:metabolism; D008297:Male; D015240:Phorbol 12,13-Dibutyrate / Q000378:metabolism; D010713:Phosphatidylcholines / Q000378:metabolism*; D011493:Protein Kinase C / Q000502:physiology*; D011650:Pulmonary Alveoli / Q000502:physiology*; D011663:Pulmonary Surfactants / Q000378:metabolism*; D051381:Rats; D011919:Rats, Inbred Strains; D011983:Receptors, Purinergic / Q000502:physiology*; D013110:Sphingosine / Q000494:pharmacology; D013726:Terbutaline / Q000494:pharmacology
D004075:Diglycerides; D007295:Inositol Phosphates; D010713:Phosphatidylcholines; D011663:Pulmonary Surfactants; D011983:Receptors, Purinergic; D015240:Phorbol 12,13-Dibutyrate; D006160:Guanosine Triphosphate; D000255:Adenosine Triphosphate; D011493:Protein Kinase C; D013726:Terbutaline; D013110:Sphingosine; D002118:Calcium