Buyang huanwu decoction improves synaptic plasticity of ischemic stroke by regulating the cAMP/PKA/CREB pathway

Ischemic stroke is an acute central nervous system disease that poses a threat to human health. It induces a series of severe pathological mechanisms, ultimately leading to neuronal cell death in the brain due to local ischemia and hypoxia. Buyang Huanwu decoction (BYHWD), as a representative formula for treating ischemic stroke, has shown good therapeutic effects in stroke patients. This study aimed to explore the mechanism of BYHWD in promoting neural remodeling after ischemic stroke from the perspective of neuronal synaptic plasticity, based on the cAMP/PKA/CREB signaling pathway. A modified suture technique was employed to establish a rat model of MCAO. The rats were divided into sham, model, and BYHWD (20 g/kg) groups. After the corresponding intervention, rat brains from each group were collected. TMT quantitative proteomics technology was employed for the research. Following proteomics studies, we investigated the mechanism of BYHWD in the intervention of ischemic stroke through animal experiments and cell experiments. The experimental animals were divided into sham, model, and BYHWD (5 g/kg, 10 g/kg, and 20 g/kg) groups. Infarct volume and severity of brain injury were measured by TTC staining. HE staining was utilized to evaluate alterations in tissue morphology. The Golgi staining was used to observe changes in cell body, dendrites, and dendritic spines. Transmission electron microscopy was used to observe the ultrastructure of synapses in the cortex and hippocampus. TUNEL staining was conducted to identify apoptotic neurons. Meanwhile, a stable and reliable (OGD/R) SH-SY5Y cell model was established. The effect of BYHWD-containing serum on SH-SY5Y cell viability was measured by CCK-8 kit. The apoptosis situation of SH-SY5Y cells was determined by Annexin V-FITC/PI. Immunofluorescence was employed to measure the fluorescence intensity of synaptic-related factors Syt1, Psd95, and Syn1. Synaptic plasticity pathways were assessed by using RT-qPCR and Western blot to determine the expression levels of cAMP, Psd95, Prkacb, Creb1/p-Creb1, BDNF, Shank2, Syn1, Syt1, Bcl-2, Bcl-2/Bax mRNA and proteins. After treatment with BYHWD, notable alterations were detected in the signaling pathways linked to synaptic plasticity and the cAMP signaling pathway-related targets among the intervention targets. This trend of change was also reflected in other bioinformatics analyses, indicating the important role of synaptic plasticity changes before and after modeling a