일차배양 뇌세포를 이용한 글루타메이트성 신경에 작용하는 천연물의 검색방법

Primary Cultured Brain Cells as Screening Methods for Natural Products Acting on Glutamatergic Neurons

Primary cultures of rat cortical and chicken embryonic brain cells were employed to establish a reliable screening method for natural products blocking or enhancing glutamate-induced neurotoxicity. Exposure of primary cultured rat cortical cells or chicken embryonic brain cells to high dose of glutamate resulted in the fragmentation of neurites and consequent neuronal death. The level of cytoplasmic lactate dehydrogenase(LDH), indicator for cell survival in cultures, was significantly reduced at exposure to glutamate. For the practical application of the methods, series of concentrations of plants extracts and positive control were applied prior to the glutamate insult on primary cultures of rat cortical and chicken embryonic brain cells. Relative LDH level in cells was measured for the estimation of the effect of the test materials on the glutamatergic neurons. The validity of the present screening method for natural products acting on glutamatergic neurons was examined with dextromethorphan, a known glutamatergic antagonist. The treatment of 100mcM dextromethorphan prevented the reduction of LDH in rat cortical and chicken embryonic, brain cells caused by glutamate insult keeping 60% and 90% of LDH level in normal control, respectively. Above results indicate that primary cultures of rat cortical and chicken embryonic brain cells could be proper systems for the screening of potential natural agents acting on glutamatergic neurons. Between the two types of cultures, primary culture of chicken embryonic brain cells seemed to be a better system for the primary screening, since it is technically easier and economical compared to that of rat cortical cells.