The cysteinyl
leukotrienes (CysLTs) are inflammatory mediators closely associated with neuronal injury after
brain ischemia through the activation of their receptors,
CysLT1R and CysLT2R. Here we investigated the involvement of both receptors in
oxygen-
glucose deprivation/recovery (OGD/R)-induced ischemic neuronal injury and the effect of the novel CysLT2R antagonist
HAMI 3379 [3-({[(1S,3S)-3- carboxycyclohexyl]amino}carbonyl)-4-(3-{4-[4-(cyclo-hexyloxy)butoxy]phenyl}propoxy)
benzoic acid] in comparison with the
CysLT1R antagonist
montelukast. In primary neurons, neither the nonselective agonist
leukotriene D4 (
LTD4) nor the CysLT2R agonist
N-methyl-leukotriene C4 (NMLTC4) induced neuronal injury, and
HAMI 3379 did not affect OGD/R-induced neuronal injury. However, in addition to OGD/R,
LTD4 and NMLTC4 induced cell injury and neuronal loss in mixed cultures of cortical cells, and neuronal loss and
necrosis in neuron-microglial cocultures. Moreover, they induced phagocytosis and
cytokine release (
interleukin-1β and
tumor necrosis factor-α) from primary microglia, and
conditioned medium from the treated microglia induced neuronal
necrosis.
HAMI 3379 inhibited all of these responses, and its effects were the same as those of CysLT2R interference by CysLT2R
short hairpin RNA, indicating CysLT2R dependence. In comparison,
montelukast moderately inhibited OGD/R-induced primary neuronal injury and most OGD/R- and LTD4-induced (but not NMLTC4-induced) responses in mixed cultures, cocultures, and microglia. The effects of
montelukast were both dependent and independent of CysLT1Rs because interference by
CysLT1R small interfering RNA had limited effects on neuronal injury in neuron-microglial cocultures and on
cytokine release from microglia. Our findings indicated that
HAMI 3379 effectively blocked CysLT2R-mediated microglial activation, thereby indirectly attenuating ischemic neuronal injury. Therefore, CysLT2R antagonists may represent a new type of therapeutic agent in the treatment of
ischemic stroke.