Neuroprotective effect of dauricine in cortical neuron culture exposed to hypoxia and hypoglycemia: involvement of correcting perturbed calcium homeostasis.

We have previously reported that dauricine protects brain tissues from focal cerebral ischemia. To corroborate this effect, neurotoxicity due to hypoxia and hypoglycemia was assessed in primary cultures of rat cortical neurons by using a trypan blue exclusion method. To further clarify the mechanism, the intracellular Ca2+ concentration ([Ca2+]i) and mitochondrial membrane potential (Deltapsim) of dissociated rat cortical cells were monitored by fura-2 fluorescence measurements and flow cytometry, respectively. The results showed that 1 and 10 micromol/L dauricine significantly enhanced neuronal survival during 4 h of hypoxia and hypoglycemia. Dauricine inhibited the increase in [Ca2+]i and decrease in Deltapsim induced by 30 min of hypoxia and hypoglycemia. When exploring the pathway, we found that 1 micromol/L dauricine inhibited the [Ca2+]i increase induced by 7.5 nmol/L thapsigargin in either the presence or absence of extracellular Ca2+ and by 1 mmol/L L-glutamate in the presence of extracellular Ca2+. These results suggest that dauricine prevents neuronal loss from ischemia in vitro, which is in accordance with our previous research in vivo. In addition, by inhibiting Ca2+ release from the endoplasmic reticulum and Ca2+ influx from the extracellular space, dauricine suppressed the increase in [Ca2+]i and, subsequently, the decrease in Deltapsim induced by hypoxia and hypoglycemia. This effect may underlie the mechanism of action of dauricine on cerebral ischemia.