Ryanodine receptor inhibitor dantrolene reduces hypoxic-ischemic brain injury in neonatal mice

Abstract

Ryanodine receptors (RyR) located on the membrane of the endoplasmic reticulum (ER), are a potent regulator of intracellular calcium levels upon activation. Dysregulated Ca2+ homeostasis is characteristic of hypoxic-ischemic (HI) brain injury and ultimately leads to neurodegeneration. RyRs have thereby been implicated in the Ca2+ imbalance that occurs during and after HI. In this study, we investigated the effects of RyR antagonist, dantrolene, on HI brain injury in neonatal mice. We found that administration of dantrolene (i.p.) on postnatal day 7 mice reduced the infarction volume and morphological damage induced by HI, and improved functional recovery as assessed by neurobehavioral testing. The neuroprotective effect of dantrolene was further demonstrated in neuronal cell culture in vitro, where dantrolene significantly reduced oxygen-glucose deprivation (OGD)-induced cell death. Fura-2 calcium imaging confirmed that dantrolene reduced the intracellular calcium level in cultured cortical neurons in vitro. Finally, Western blot analysis showed that dantrolene treatment reduced cleaved caspase-3 and -9 apoptotic proteins, and elevated pro-survival protein kinase C (PKC) protein levels. Taken together, our results demonstrate that dantrolene exerts neuroprotective effects against neonatal HI brain injury. This suggests that RyRs play a role in mediating the ionic imbalance induced by HI and therefore represent a potential target for drug development.