Gami-Guibitang Attenuates Anxiety-like Behaviors and Modulates Hippocampal Synaptic Signaling in a Valproic...

Autism spectrum disorder (ASD) is a neurodevelopmental condition characterized by social deficits, repetitive behaviors, and heightened anxiety. Despite extensive research, effective interventions targeting core symptoms remain limited. Gami-Guibitang (GBT), a traditional herbal formula, has been clinically prescribed for anxiety-related symptoms and cognitive complaints, yet its effects on ASD-associated behavioral and molecular abnormalities have not been fully elucidated.

This study aimed to evaluate the anxiolytic and neuroregulatory effects of GBT in a valproic acid (VPA)-induced ASD mouse model, focusing on behavioral outcomes and hippocampal synaptic protein expression.

Pregnant C57BL/6N mice received a single intraperitoneal injection of VPA (500 mg/kg) at embryonic day 12.5. Male offspring were administered GBT (150 mg/kg, p.o.) twice daily for 4 weeks from postnatal day 21 (PND 21). These mice were behaviorally evaluated by the open-field test, elevated plus maze, marble-burying test, Y-maze, three-chamber social interaction test, and Morris water maze. Western blot analysis was conducted to examine hippocampal expression of phosphorylated and total CREB and GluR1, PI3K/Akt signaling components, as well as GABRA1 and GABRB1.

VPA-exposed offspring exhibited increased anxiety-like behaviors, altered repetitive behaviors, dysregulated exploratory activity, and impaired spatial learning, and reduced spontaneous alternation performance in the Y-maze. GBT reduced anxiety-like behaviors in the elevated plus maze and marble burying tests, partially improved spatial learning acquisition in the Morris water maze, and normalized excessive locomotor activity, without significantly affecting short-term working memory performance. At the molecular level, GBT significantly attenuated VPA-induced hyperphosphorylation of CREB, GluR1, PI3K, and Akt, indicating suppression of aberrant synaptic signaling rather than global enhancement. In addition, GBT increased GABRA1 expression toward control levels and enhanced GABRB1 expression beyond baseline, suggesting selective modulation of GABAergic receptor subunit composition rather than simple normalization.

These findings provide preclinical evidence that GBT alleviates anxiety-like behavior and modulates hippocampal synaptic signaling disrupted by prenatal VPA exposure. By attenuating aberrant excitatory signaling and selectively regulating GABAergic receptor balance, GBT may represent a multi-target herbal candidate for modulating ASD-associated emotional dysregulation and domain-specific cognitive dysfunction, rather than acting as a broad cognitive enhancer.