Abstract 39: The Transient Receptor-Potential Channel 6 Regulates Thromboxane A2 Receptor-Operated Calcium Entry and Plays a Critical Role in Platelet Function

Abstract

Although changes in the intracellular levels of calcium is a central step in platelet activation, the underlying mechanism of changes that are dependent on receptor-operated calcium entry (ROCE) remains to be defined. Furthermore, it was proposed, though never proven, that the Transient Receptor-Potential Channels (TRPCs) may play a key role in this process. In this connection, we have previously shown that TRPC6 plays a vital role in physiological hemostasis and thrombogenesis. However, the underlying mechanism by which TRPC6 modulates these processes also remains to be determined. Based on these considerations, we hypothesized that TRPC6 plays an essential role in ROCE and hence platelet function. Using a (genetic) TRPC6 knockout (KO) mouse approach, our preliminary studies revealed that these platelets exhibited a defect in platelet aggregation induced by the thromboxane A 2 receptor (TPR) agonist U46619. Conversely, the aggregation response triggered by ADP or the thrombin receptor activating peptide 4 (TRAP4) was found to be comparable to that of the wild-type (WT) platelets. Separate studies revealed that the TRPC6 deficient platelets were also found to exhibit a defect in TPR-mediated dense granule (ATP) secretion, whereas that of ADP and TRAP4 was normal. Moreover, we observed a defect in integrin GP IIb-IIIa activation that was again specific to TPR (normal activation in response to ADP and TRAP4), suggesting a defect in inside-out signaling. Finally, TPR-dependent CE was also found to be deficient in the TRPC6 KO platelets, unlike that stimulated by ADP or TRAP4. Future studies will further investigate the molecular mechanism of TRPC6-regulated platelet function and CE. Taken together, these findings demonstrate for the first time that TRPC6 regulates CE in a TPR-dependent manner and that this regulation consequently modulates platelet aggregation, secretion, as well as GP IIb-IIIa activation. These studies may define a new therapeutic target for managing multiple thrombosis-based disorders.