Shenzhi Jiannao formula ameliorates vascular dementia in vivo and in vitro by inhibition glutamate neurotoxicity via promoting clathrin-mediated endocytosis

Jingfeng Lin,Ganlu Liu,Yuchun Wang,Danfeng Tian,Dandan Zhang,Zhenyun Han,Rui Su,Yangyang Guo,Qiang Gao,Ze Chang

doi:10.1186/s13020-021-00477-4

Abstract

BackgroundSynaptic damage and glutamate excitotoxicity have been implicated in the pathogenesis of vascular dementia (VD). Clathrin, RAB5B and N-methyl-d-aspartic acid receptor 1 (NMDAR1) proteins play a vital role in endocytosis of synaptic vesicles in neurons and glutamate over accumulation. Previous researches have been confirmed that Shenzhi Jiannao (SZJN) formula has an anti-apoptotic and neuroprotective effect in VD, but the underlying mechanisms are still unclear. In this study, we aimed to explore the effect of SZJN formula on cognitive impairment and glutamate excitotoxicity via clathrin-mediated endocytosis (CME) in vivo and in vitro.MethodsSZJN formula consists of Panax ginseng C.A.Mey., Anemarrhena asphodeloides Bunge, and Paeonia anomala subsp. veitchii (Lynch) D.Y.Hong & K.Y.Pan. All herbs were prepared into granules. Both common carotid arteries were permanent occluded (2‐vessel occlusion, 2VO) in male Sprague Dawley (SD) rats to model VD. One day after operation, the rats began daily treatment with SZJN formula for 2 weeks. The neuroprotective effects of SZJN formula was subsequently assessed by the novel object recognition test, Morris water maze, hematoxylin–eosin (HE) staining and Nissl staining. Glutamate cytotoxicity was assessed by detecting cell viability and cell death of PC12 cells. Immunohistochemistry, immunofluorescence, Western blot, and quantitative real‐time PCR were used to detect the expression levels of clathrin, RAB5B, and NMDAR1.ResultsAdministration of SZJN formula effectively improved short-term memory and spatial memory. SZJN formula treatment significantly reduced hippocampal neuronal loss, and recovered the arrangement and morphology of neurons and Nissl bodies. Moreover, SZJN formula promoted the proliferation of PC12 cells and inhibited glutamate-induced cell death. The down-regulation of clathrin and RAB5B, as well as the upregulation of NMDAR1 in the brain induced by 2VO or glutamate was also notably reversed by SZJN formula at both the protein and mRNA levels, which may contribute to SZJN formula induced improved neurological function.ConclusionsTaken together, our findings provide evidence that the neuroprotective effects of SZJN formula in experimental VD maybe mediated through promoting the expression of clathrin-mediated endocytosis and reducing NMDARs‐associated glutamate excitotoxicity. SZJN formula serves as a promising alternative therapy and may be a useful herbal medicine for preventing progression of VD.Graphic abstract

Highlights

Vascular dementia (VD) is the second most common type of senile dementia and maybe the main dementia type in East Asia [1]
Shenzhi Jiannao (SZJN) increased clathrin and RAB5B expression, whereas reduced cytomembrane N-methyl-d-aspartic acid receptor 1 (NMDAR1) expression in the hippocampus of vascular dementia (VD) rats We evaluated whether SZJN formula could inhibit glutamate toxicity and promote the clathrin-mediated endocytosis (CME) process associated with upregulating clathrin and RAB5B, and downregulating NMDAR1 in the hippocampus
Our study showed that SZJN formula could significantly up-regulate clathrin and RAB5B expression, and we speculated that was associated with the significant reduction of surface NMDAR1 expression, which may be closely involved in promoting endocytosis of NMDARs and attenuating toxic glutamate accumulation

Summary

Introduction

Vascular dementia (VD) is the second most common type of senile dementia and maybe the main dementia type in East Asia [1]. Studies have confirmed that the excessive release of glutamate in the synaptic cleft and subsequent C a2+ influx via NMDA-subtype glutamate receptors could lead to an intracellular cascade of cytotoxic events [7, 8]. The excessive activation of NMDA receptors evokes the impairment of synaptic plasticity, neuronal dysfunction and death [9, 10]. Synaptic damage and glutamate excitotoxicity have been implicated in the pathogenesis of vascular dementia (VD). RAB5B and N-methyl-d-aspartic acid receptor 1 (NMDAR1) proteins play a vital role in endocytosis of synaptic vesicles in neurons and glutamate over accumulation. We aimed to explore the effect of SZJN formula on cognitive impairment and glutamate excitotoxicity via clathrin-mediated endocytosis (CME) in vivo and in vitro

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