Ne assay kit

The serum levels of CORT were measured using a radioimmunoassay with a Vitek Immune Diagnostic Assay System (Bio-RAD iMark, Berkeley, California, USA) according to the manufacturer’s instructions by using a CORT assay kit (HY-10063, Beijing Sino-UK Institute of Biological Technology, Beijing, China). The concentrations of serum NE were detected using an enzyme-linked immunesorbent assay (ELISA) according to the manufacturer’s instructions by using a NE assay kit (H096, Nanjing Jiancheng Bioengineering Institute, Nanjing, China). The samples were weighed and then homogenized on ice to detect the content of NO in the hypothalamus. After the solution was centrifuged at 1000×g for 10 min, the supernatant was collected, and the NO content was detected using the nitric-acid-reductase method according to the manufacturer’s instructions using a NO assay kit (A012–1-2, Nanjing Jiancheng Bioengineering Institute, Nanjing, China; n = 12 rats per group).

Following the behavioral tests, the mice were euthanized, and their brains were isolated and placed in centrifuge tubes immersed on ice, followed by the addition of nine times the volume of normal saline solution. After homogenization with a tissue precision homogenizer, the supernatant was obtained through centrifugation (4 °C, 4000 rpm, 10 min). The contents of GABA, 5-HT, NE, Glutamine (Glu), GAD67, GABAAR, SOD, IL-1β, iNOS in the mouse brain homogenate supernatant were examined by assay kit according to the instructions of the manuscripts. The mouse GABA ELISA Kit (Product code: JL12094), GABAAR ELISA Kit (Product code: JL50569), and GAD67 ELISA Kit (Product code: JL46227) were purchased from Shanghai Jianglaibio Science and Technology Ltd., China; The 5-HT assay Kit (Product code: H104), NE assay Kit (Product code: H096), NOS typed assay kit (Product code: A014-1-2), Glu measurement kit (Product code: A074-1-1), IL-1β Assay Kit (Product code: H002), SOD assay kit (Product code: A001-3-2) and Total protein quantitative assay kit (Product code: A045-2-2) were purchased from Nanjing Jiancheng Bioengineering Institute, China.