Spinosin Ameliorates PCPA-Induced Insomnia and Anxiety-Like Behaviors via a TrkB-Dependent Restoration of Hypothalamic GABAergic Signaling

Background: Insomnia commonly presents alongside hyperarousal and anxiety and is often accompanied by a fundamental imbalance between inhibitory γ-aminobutyric acid (GABA) and excitatory glutamate (Glu) tissue levels. Current therapeutics often lack efficacy in addressing the neuroplastic deficits associated with chronic sleep disturbances. Spinosin is a C-glycoside flavonoid derived from Ziziphi Spinosae Semen with reported sedative activity, yet whether it improves insomnia-associated anxiety and which neurotrophic pathways are involved remain unclear. This study aimed to evaluate spinosin's effects on sleep indices and anxiety-like behaviors and tested whether activation of the Brain-derived neurotrophic factor (BDNF)-Tropomyosin receptor kinase B (TrkB)-cAMP response element-binding protein (CREB) axis contributes to recovery of hypothalamic GABAergic function. Methods: Insomnia-like phenotypes were established in male C57BL/6J mice using p-chlorophenylalanine (PCPA), after which spinosin was administered orally (10–40 mg/kg) once daily for 10 days. Sleep behavior was assessed with a pentobarbital challenge, and anxiety-related behaviors were evaluated via the open field, light-dark box, elevated plus maze, and novelty-suppressed feeding tests. Neurotransmitter levels, protein expression, and gene transcription in the hypothalamus were analyzed. To probe causality, we used corticosterone-injured HT22 cells and pharmacologically blocked TrkB with ANA-12 in vitro. Results: Across the tested doses, spinosin reduced sleep latency, prolonged sleep duration, and mitigated anxiety-like behaviors in PCPA-treated mice without impairing motor coordination. Mechanistically, spinosin restored the hypothalamic Glu/GABA balance by upregulating glutamate decarboxylase 67 (GAD67) and GABA_A receptor expression. These effects were accompanied by the reactivation of the hypothalamic BDNF-TrkB-CREB signaling pathway. Importantly, ANA-12 largely eliminated these spinosin-mediated changes in vitro, supporting a TrkB-dependent component. Conclusion: Spinosin effectively ameliorates insomnia and comorbid anxiety by re-establishing hypothalamic excitatory and inhibitory neurotransmitter levels. This therapeutic action is fundamentally driven by the activation of BDNF-TrkB signaling, which subsequently reinforces GABAergic signaling. Together, the findings suggest that spinosin may act as a multi-target candidate for insomnia accompanied by hyperarousal.