Effects of ketamine on glutamate-induced apoptosis in neuronal PC 12 cells

Objective To study the effects of ketamine on glutamate-induced apoptosis in neuronal cells using PC 12 pheochromocytoma cell line (provided by Chinese Academy of Phamacological Research) .Methods After being incubated in the culture medium containing 7S-NGF for 6 days. Over 95 % of the PC cells differentiated into neuron-type cells. The 7S-NGF induced differentiated neuronal PC 12 cells were seeded in 24-well plates pre-coated with poly-L-lysine(2×106 cells per well) .24 hours later the PC12 cells were exposed to glutamate 20 mrnol/ L(group A); glutamate 20 mmol/L + ketamine 0.1 mmo/L (group B); glutamate 20 mmol/L + ketamine 1.0 mmol/L (group C); glutamate 20 mmol/L + D-APS 100 )Ltmol/L(group D) and new culture medium containing no 7S-NGF(group E, control group), and incubated for 18 hours .The viability of the cells was evaluated by the ability of the cells to reduce the tetragotium derivative MTT into a blue formagan salt. DNA fragmentation indicative of apoptosis was detected using TUNEL technique. Results in group A following incubation with glutamate 20 mmol/L for 18 h , at 37℃, the viability was PC 12 cells was reduced to 37%± 6% However ketamine , when added to the culture medium to gather with glutamate , inhibited glutamate-induced cell death . The viability of PC 12 cell was 65 ± 7% in group B an 99±10% in group C. Ketamine appeared to attenuate the apoptotic process, because the number of the apoptotic cell bodies, determinated by YUNEL was also reduced by ketamine, with only 15-20% of neuronal cells staining positive after exposure to 20 mmol/L glutamate.The difference between group A and C was very significant (P 0.01).In addition D-APS 100μmol /L ( a competitive antagonist) also exerted significant protective effects on neuronal PC 12 cells. Conclusion Ketamine protects neuronal PC 12 cells from glutamate neurotoxicity induced apoptosis.