Sphingosine-1-phoshpate (S1P) is a bioactive lysophospholipid, generated and released at sites of tissue injury. S1P signals through a variety of G-protein coupled receptor subtypes and there are three major sub-types, S1P
1
, S1P
2
, and S1P
3
, to mediate cardiovascular responses. S1P
2
and S1P
3
receptors couple to Gα
i
, Gα
12
, Gα
13
and Gα
q
and we first examined the contribution of S1P
2
and S1P
3
to cardiac hypertrophy using S1P
2
and S1P
3
knockout (KO) mice and found that there is no difference in hypertrophy induced by pressure-overload. We previously showed that S1P provides cardioprotection against oxidative stress such as ischemia/reperfusion in which RhoA activation and its downstream effector PKD1 play an important role. It has not, however, been determined which S1P receptor subtype is responsible for S1P mediated cardioprotection. We knocked down the three major S1P receptors using siRNA in neonatal rat ventricular myocytes (NRVMs) and assessed RhoA and PKD1 activation induced by S1P. Knockdown of S1P
3
abolished RhoA activation and largely attenuated phosphorylation of PKD1 while knockdown of S1P
1
and S1P
2
did not. Using siRNA or pertussis toxin to inhibit different G-proteins, we further established that S1P
regulates RhoA activation through Gα
13
, but not Gα
12
, Gα
q
,
or Gα
i
. To investigate the role of S1P
3
receptors in the adult heart, hearts were isolated from wild-type or S1P
3
KO adult mice, perfused in the Langendorff mode and subjected to ex vivo ischemia/reperfusion. As previously reported, S1P perfusion significantly reduced infarct size induced by ischemia/reperfusion in WT hearts (by 50%), but this protection was abolished in the S1P
3
KO mouse heart. To further confirm the role of S1P
3
in cardioprotection we perfused WT mouse hearts with an S1P
3
-specific agonist CYM-51736. We observed that CYM-51736 attenuated the infarct size to a similar degree as that observed with S1P. Our findings reveal that activation of the S1P
3
receptor coupling to Gα
13
and subsequent RhoA activation is responsible for cardioprotection against ischemia/reperfusion. Accordingly specific drug targeting of S1P
3
receptors could provide therapeutic benefits in ischemic heart disease without the undesirable effects of global activation of other cardiac S1P receptors.
Endocannabinoids and Liver Disease. III. Endocannabinoid effects on immune cells: implications for inflammatory liver diseases
