Guilu Erxian Jiao remodels dendritic spine morphology through activation of the hippocampal TRPC6-CaMKIV-CREB signaling pathway and suppresses fear memory generalization in rats with post-traumatic stress disorder.

ETHNOPHARMACOLOGICAL RELEVANCE: Guilu Erxian Jiao (GLEXJ) is a renowned traditional Chinese herbal formula used to tonify the kidney. It is employed to treat psychiatric disorders, and alleviate memory impairment, cognitive dysfunction, and behavioral disorders. Modern pharmacological studies have demonstrated GLEXJ's ability to significantly inhibit the fear response in post-traumatic stress disorder (PTSD) and facilitate the extinction of fear memory. However, the underlying pharmacological mechanisms remain elusive.

AIM OF THE STUDY: Fear memory generalization, a fundamental characteristic of PTSD, remains poorly understood, and optimal pharmacological treatments are lacking. This study aimed to investigate GLEXJ's inhibitory effects on fear memory generalization in PTSD rats and elucidate its underlying mechanisms.

MATERIALS AND METHODS: PTSD rats were induced using the single prolonged stress and electrical stimulation (SPS&S) protocol and treated with GLEXJ or paroxetine (PRX). Fear memory generalization was assessed using a contextual fear memory test. Hippocampal dendritic spine morphology was analyzed using Golgi-Cox staining. The chemical composition of GLEXJ was determined using ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS). Network pharmacology was employed to predict GLEXJ's therapeutic mechanism in PTSD treatment. Western blotting and immunofluorescence were used to measure indicators of the transient receptor potential channel 6 (TRPC6)-mediated calcium/calmodulin-dependent protein kinase IV-cAMP response element-binding protein (CaMKIV-CREB) signaling pathway. In vitro, TRPC6 was suppressed in rat adrenal pheochromocytoma (PC12) cells using lentiviral vectors, and phalloidin staining was employed to examine changes in Fibros actin (F-actin), elucidating the mechanistic effects of GLEXJ-containing serum.

RESULTS: GLEXJ significantly mitigated fear memory generalization in PTSD rats, even with repeated stress exposure. It also alleviated abnormal hippocampal dendritic spine morphology. Network pharmacology analysis confirmed that GLEXJ was closely related to the Ca2+ signaling pathway in PTSD treatment. PTSD rats exhibited disrupted TRPC6-mediated CaMKIV-CREB signaling and impaired synaptic plasticity. GLEXJ upregulated TRPC6 expression, reactivated the CaMKIV-CREB pathway, and promoted synaptic remodeling. In vitro studies confirmed that TRPC6 suppression reduced F-actin levels while GLEXJ-containing serum increased TRPC6 expression and F-actin content.

CONCLUSIONS: GLEXJ activates CaMKIV-CREB signaling by upregulating TRPC6 in the hippocampus of PTSD rats, leading to the positive modulation of dendritic spine morphology and synaptic remodeling. This mechanism contributes to the attenuation of fear memory generalization. Given the limitations of current PTSD treatments, these findings offer potential avenues for developing more effective therapeutic strategies.