Sleep-Aiding Effect of Casein-Derived Peptide TPTLN and Its Underlying Mechanism

Objective: To investigate the sleep-aiding effect of casein-derived peptide TPTLN and explore its possible mechanism of action. Methods: Thirty mice were divided equally into three groups, an experimental, a positive control and a blank control group, which were gavaged with TPTLN at a dose of 5 mg/kg m_b, intraperitoneally injected with diazepam at a dose of 1 mg/kg m_b, and gavaged with sterilized water at a dose of 5 mg/kg m_b, respectively. The body mass of all the mice was recorded every day. After 7 days of continuous administration, a sodium pentobarbital-induced sleep test was carried out to evaluate sleep quality. Following 4 more days of administration, the mice were euthanized and dissected to collect blood and tissue samples for analysis. Enzyme-linked immunosorbent assay (ELISA) was used to determine the levels of 5-hydroxytryptamine (5-HT), γ-aminobutyric acid (GABA), 5-hydroxyindoleacetic acid (5-HIAA), glutamate (Glu), β-endorphin (β-EP), cortisol, norepinephrine (NE) in the serum and brain tissue, and the GABA/Glu ratio was evaluated. Real-time quantitative polymerase chain reaction (qPCR) was used to detect the effect of TPTLN on the mRNA relative expression levels of γ-aminobutyric acid A receptor α1 (GABA_A-α1) and 5-hydroxytryptamine receptor 1A (5-HT1A) in the hypothalamus. Results: Compared with the control group, TPTLN and sodium pentobarbital synergistically increased the sleep onset rate to 100%, significantly shortened the sleep latency by 31.7% (P < 0.05), and extended the sleep duration 2.1 times (P < 0.05); increased the GABA/Glu ratio in the serum and brain tissue by 68.8% and 19.3%, respectively and significantly improved the excitation/inhibition imbalance (P < 0.001, P < 0.0001) as evidenced by a 49.4% increase in the serum GABA level (P < 0.001), a 7.6% increase in the brain 5-HT level, and a 24.3% and 23.4% decrease in the brain levels of cortisol and NE (P < 0.05); and significantly up-regulated the expression levels of the GABA_A-α1 and 5-HT1A genes 1.5 and 1.3 times, respectively (P < 0.01, P < 0.05). Conclusion: Casein-derived peptide TPTLN exhibited a favorable sleepaiding effect, and the underlying mechanism may be associated with the regulation of GABA_A-α1 and 5-HT1A expression levels, as well as the modulation of neurotransmitter contents including GABA, Glu, 5-HT, cortisol and NE in the brain.