To determine glutamate and GABA in the hippocampus, the hippocampus tissue was homogenized in an ice-cold PBS buffer and centrifuged at 10,000 rpm for 10 min at 37°C. The supernatant was used in the following assays.
The concentration of glutamate in the hippocampus was tested via the ultraviolet colorimetry method according to the instructions in the Glutamic acid measurement kit (Nanjing Jiancheng Bioengineering Institute, China).
The content of GABA was measured with the ELISA Kit for Gamma-Aminobutyric Acid (Cloud-Clone Corp., United States) in accordance with the manufacturer’s instructions. The sample was added into the prepared Detection Reagent A and incubated for 1 h at 37°C. The unbound conjugate was washed off, each microplate well was added to the prepared Detection Reagent B, and subsequently incubated for 30 min at 37°C. The substrate solution and the stop solution were used for color development reaction and termination. The absorbance was measured at 450 nm with a microplate reader (Beckman Coulter, United States). Each experiment was repeated in triplicate.
The concentration of glutamate in the hippocampus was tested via the ultraviolet colorimetry method according to the instructions in the Glutamic acid measurement kit (Nanjing Jiancheng Bioengineering Institute, China).
The content of GABA was measured with the ELISA Kit for Gamma-Aminobutyric Acid (Cloud-Clone Corp., United States) in accordance with the manufacturer’s instructions. The sample was added into the prepared Detection Reagent A and incubated for 1 h at 37°C. The unbound conjugate was washed off, each microplate well was added to the prepared Detection Reagent B, and subsequently incubated for 30 min at 37°C. The substrate solution and the stop solution were used for color development reaction and termination. The absorbance was measured at 450 nm with a microplate reader (Beckman Coulter, United States). Each experiment was repeated in triplicate.