LX-2931

It is well accepted that sphingolipids play an important role in the pathological process of cerebral ischemia. In the present study we have investigated the involvement of sphingosine 1-phosphate (S1P) pathway in two different in vitro models of global ischemia. In organotypic hippocampal slices exposed to oxygen and glucose deprivation (OGD) we evaluated the mRNA expression of S1P metabolic enzymes and receptors (S1P_1-5) by Real Time-PCR. In the same model we investigated the effect of the inhibitor of S1P lyase (SPL), LX2931, the selective antagonists of S1P₂, JTE-013, and S1P₃, CAY10444, quantifying the cell death in the CA1 region by propidium iodide fluorescence, and morphological and tissue organization alterations by immunohistochemistry and confocal microscopy. Moreover, we performed extracellular recordings of field excitatory postsynaptic potentials in acute slices exposed to OGD. In organotypic slices OGD induced a significant increase of SPL at mRNA level and of S1P₂ and S1P₃ at both mRNA and protein level. The incubation with LX2931, JTE-013 or CAY10444 was able to reduce CA1 damage induced by OGD in organotypic slices and provoked a significant delay of the onset of anoxic depolarization on acute slices. Moreover, S1P₂ and S1P₃ antagonists prevented the increase of TREM2 induced by OGD. Our results reveal a dual role of S1P pathway in brain ischemia: intracellular S1P, degraded via SPL, appears to be beneficial whereas signaling via S1P₂ and S1P₃ is detrimental to the disease. These findings support the notion that SPL, S1P₂ and S1P₃ are promising therapeutic targets in brain ischemia.