Activation of phospholipase D in rat type II pneumocytes by ATP and other surfactant secretagogues.

Surfactant phospholipid secretion can be stimulated by a variety of agonists acting via a number of signal-transduction mechanisms. To determine whether phospholipase D has a role in surfactant secretion, we examined phosphatidylethanol formation in response to surfactant secretagogues in primary cultures of rat type II cells. Phosphatidylethanol formation was stimulated by ATP, 12-O-tetradecanoylphorbol-13 acetate (TPA), and dioctanoylglycerol, surfactant secretagogues that also activate protein kinase C. Surfactant secretagogues that act via other signaling mechanisms had no effect on phosphatidylethanol formation. The effect of ATP on phosphatidylethanol formation was dependent on time, with the maximum stimulation being achieved in approximately 10 min. It was also dependent on ATP concentration. The ATP concentration eliciting 50% of the maximum effect (EC50) was 2.45 x 10(-6) M. This was similar to the EC50 reported for ATP stimulation of surfactant secretion. ATP analogues also stimulated phosphatidylethanol formation with a potency order generally similar to that reported for surfactant secretion. The effects of ATP, TPA, and dioctanoylglycerol were antagonized by protein kinase C inhibitors. We speculate that activation of protein kinase C either directly by TPA and dioctanoylglycerol or indirectly subsequent to phosphoinositide-specific phospholipase C activation by ATP leads to initial stimulation of surfactant secretion as well as activation of phospholipase D. The action of phospholipase D on cellular phospholipids then leads to further generation of diacylglycerols, continued activation of protein kinase C, and sustained surfactant secretion.