Different neuronal cells are found in olfactory bulb, cortex and cerebellum regions. Olfactory regulates complex behaviors, cerebellum has a great role in body balance and cortex mainly coordinate complex motor and sensory behaviors. The anesthetic is given to all patients (from birth to death) who undergo surgery. Midazolam is GABA agonist and used as an anesthesia. Glutamate an excitatory neurotransmitter can reach toxic levels in traumatic brain injury to cause schizophrenia and behavior disorders. In this study, we obtained 3 different olfactories, cortex and cerebellum cell culture. Glutamate 5-10 mM was added to all culture dishes except the negative control group. Midazolam was added in 4 different doses for 24th and then evaluation was done by MTT, TAC (Total antioxidant capacity), TOS (Total oxidant status) and flow cytometry (Annxin V-apoptosis marker). Our results show that although ketamine is NMDA antagonist there is an increased toxicity level of glutamate and reduce cell viability.