UC San Diego

Sphingosine-1-phoshpate (S1P) is a lysophospholipid that can signal through multiple G protein-coupled receptor subtypes. S1P is generated and released at sites of tissue injury in the heart and can act on S1P₁, S1P₂, and S1P₃ receptor subtypes to effect cardiovascular responses. We established that S1P causes little phosphatidylinositol hydrolysis and does not induce hypertrophy in neonatal rat ventricular myocytes (NRVMs), indicating that it does not cause receptor coupling to Gq. We previously demonstrated that S1P confers cardioprotection against ischemia/ reperfusion (I/R) and mediates this cardioprotection by activating RhoA and its downstream effector PKD. The S1P receptors and G proteins that regulate RhoA activation and downstream response in the heart have not been determined. By inhibiting different G proteins in NRVMs, we established that S1P regulates RhoA activation through G[alpha]₁₃. Knockdown of the three S1P receptors using siRNA demonstrated a requirement for S1P₃ in RhoA activation and subsequent PKD phosphorylation, and was confirmed in studies using isolated hearts from S1P₃ knockout (KO) mice. To investigate the functional role of S1P₃ receptors, we subjected isolated wild-type (WT) or S1P₃ KO mouse hearts to ex vivo I/R. Addition of S1P prior to I/R significantly reduced infarct size in WT hearts; this protection was abolished in the S1P₃ KO. To confirm the role of S1P₃ in cardioprotection, WT mouse hearts were perfused with S1P₃-specific agonist CYM-51736; infarct development was attenuated to a degree similar to that achieved by S1P. These findings suggest that S1P₃- and G[alpha]₁₃-mediated RhoA/PKD activation are responsible for protection against I/R