Distinct signaling pathways mediate cardiomyocyte phospholipase D stimulation by endothelin-1 and thrombin.

Abstract Several G protein-coupled receptors which stimulate phospholipase C (PLC) also activate phospholipase D (PLD) in cardiomyocytes. Here, we characterized PLD activation in neonatal rat cardiomyocytes by the PLC-stimulatory thrombin receptor PAR1, in comparison to the endothelin-1 receptor ET A R, which induces PLD stimulation by activation of protein kinase C (PKC) δ and ϵ . Similar to ET A R, activation of PAR1 induced PLD stimulation, which, however, was insensitive to PKC inhibition. Furthermore, in contrast to ET A R, PLD stimulation by PAR1 was suppressed by overexpression of regulators of G protein signaling specific for G 12 -type G proteins and treatment with brefeldin A, an inhibitor of guanine nucleotide exchange factors for ADP-ribosylation factor (ARF) GTPases. On the other hand, inactivation of Rho GTPases by Clostridium difficile toxin B and treatment with general tyrosine kinase inhibitors suppressed PAR1- and ET A R- as well as phorbol ester-induced PLD stimulation and was associated with a fall in the cellular level of phosphatidylinositol 4,5-bisphosphate (PIP 2 ). We conclude that, in contrast to ET A R-PLD coupling, PAR1-induced cardiomyocyte PLD stimulation is PKC-independent and mediated by G 12 -type G proteins and ARF GTPases, while Rho and tyrosine kinases regulate PLD stimulation by either receptor, apparently by controlling the cellular level of PIP 2 , a common regulator of PLD activity.