Quantification of Brain Neurotransmitters

Complete rat brain was dissected, washed with cold saline and stored at -70°C until estimation of neurotransmitters, i.e., dopamine and serotonin. For neurotransmitter analysis, tissues were homogenized in phosphate buffer and centrifuged at 14000 rpm for 10 min at 4°C. After centrifugation, the supernatant was filtered by a syringe filter (0.2 μm), and 20 μl of the sample was injected into the Agilent HPLC-1200. Isocratic mobile phase of 50 mM potassium phosphate buffer in 3% methanol was used for analysis. The flow rate was adjusted to 1.5 ml/min with total run time of 25 min. HPLC separation was carried out using a C18 (5 μm × 250 mm × 4.6 mm) column and detected at 270nm by a UV detector. Experiments were performed in triplicate. Neurotransmitter levels were quantified using standard linear regression equations [18 (link), 24 (link)].