Cerebrospinal fluid samples from six mice per group were collected for ELISA. CSF samples (10 μL) were diluted 1:20 with artificial CSF (a buffer containing 103 mmol/L NaCl, 4.7 mmol/L KCl, 1.2 mmol/L KH2PO4, 1.2 mmol/L MgSO4, 25 mmol/L NaHCO3, 10 mmol/L glucose, 1 mmol/L sodium pyruvate, and 2.5 mmol/L CaCl2, pH 7.4) (Shen et al., 2020). Concentrations of nesfatin-1 in the CSF were determined by a mouse nesfatin-1 ELISA kit from JianglaiBIO (Beijing, China) according to the manufacturer’s instructions (Zhang et al., 2018).

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