N-Acetylcysteine and ebselen but not nifedipine protected cerebellar granule neurons against 4-hydroxynonenal-induced neuronal death

4-Hydroxynonenal (HNE), an aldehydic product of membrane lipid peroxidation, has been shown to induce neurotoxicity in various types of neurons. To clarify the mechanisms underlying HNE-induced neurotoxicity, the effects of antioxidants (N-acetylcysteine (NAC) and ebselen with or without NAC pretreatment) and Ca2+-related reagents were examined in cerebellar granule neurons. The decreases in neuronal survival and mitochondrial membrane potential induced by HNE were suppressed by pretreatment with NAC at concentrations of 500 and 1000 μM. HNE-induced protein modification and reactive oxygen species generation were also suppressed by pretreatment with NAC at 1000 μM. Although simultaneous application of ebselen (10 μM) did not protect against HNE-induced neurotoxicity, it completely suppressed HNE-induced injury after pretreatment with NAC at 300 μM. HNE increased [Ca2+]i levels, and this increase was significantly attenuated by simultaneous application of nifedipine (10 μM) or EGTA (1000 μM), but not by MK-801 or CNQX. However, none of these Ca2+-related reagents was able to prevent HNE-induced neuronal death or mitochondrial injury. These results suggest that pretreatment with a low concentration of NAC dramatically potentiates the neuroprotective activity of ebselen, and that HNE-induced increase in [Ca2+]i is not involved in HNE-induced neuronal death in cerebellar granule neurons.