Neuroprotective effect of a novel Chinese herbal decoction on cultured neurons and cerebral ischemic rats.
BACKGROUND
Historically, traditional Chinese medicine has been widely used to treat stroke. Based on the theory of Chinese medicine and the modern pharmacological knowledge of herbal medicines, we have designed a neuroprotective formula called Post-Stroke Rehabilitation (PSR), comprising seven herbs - Astragalus membranaceus (Fisch.) Bunge, Salvia miltiorrhiza Bunge, Paeonia lactiflora Pall., Cassia obtusifolia L., Ligusticum chuanxiong Hort., Angelica sinensis (Oliv.) Diels, and Glycyrrhiza uralensis Fisch. We aim to examine the neuroprotective activity of PSR in vitro and in vivo, and to explore the underlying molecular mechanisms, to better understand its therapeutic effect and to further optimize its efficacy.


METHODS
PSR extract or vehicle was applied to primary rat neurons to examine their survival effects against N-methyl-D-aspartate (NMDA)-elicited excitotoxicity. Whole-cell patch-clamp recording was conducted to examine the NMDA-induced current in the presence of PSR. ERK- and CREB-activation were revealed by western blot analysis. Furthermore, PSR was tested for CRE promoter activation in neurons transfected with a luciferase reporter. The protective effect of PSR was then studied in the rat middle cerebral artery occlusion (MCAO) model. MCAO rats were either treated with PSR extract or vehicle, and their neurobehavioral deficit and cerebral infarct were evaluated. Statistical differences were analyzed by ANOVA or t-test.


RESULTS
PSR prominently reduced the death of cultured neurons caused by NMDA excitotoxicity in a dose-dependent manner, indicating its neuroprotective property. Furthermore, PSR significantly reduced NMDA-evoked current reversibly and activated phosphorylation of ERK and CREB with distinct time courses, with the latter's kinetics slower. PSR also triggered CRE-promoter activity as revealed by the increased expression of luciferase reporter in transfected neurons. PSR effectively reduced cerebral infarct and deficit in neurological behavior in MCAO rats when PSR decoction was administered starting either 6 days before or 6 h after onset of ischemia.


CONCLUSIONS
PSR is neuroprotective both in vitro and in vivo - it protects cultured neurons against NMDA excitotoxicity, and effectively reduces ischemic injury and neurobehavioral deficit in MCAO rats in both the pre- and post-treatment regimens. The underlying neuroprotective mechanisms may involve inhibition of NMDA receptor current and activation of ERK and CREB. This study provides important preclinical data necessary for the further development of PSR for stroke treatment.
Division of Life Science, The Hong Kong University of Science and Technology, Clear Water Bay, Hong Kong, China.|State Key Laboratory of Molecular Neuroscience, The Hong Kong University of Science and Technology, Clear Water Bay, Hong Kong, China.|Guangdong Key Laboratory of Brain Science, Disease and Drug Development, HKUST Shenzhen Research Institute, Shenzhen, Guangdong, China.
Fanny Chui-Fun
FC
Ip
Division of Life Science, The Hong Kong University of Science and Technology, Clear Water Bay, Hong Kong, China.|Present Address: Department of Pharmacology, School of Basic Medical Sciences, Capital Medical University, Beijing, 100069, China.
Yu-Ming
YM
Zhao
Division of Life Science, The Hong Kong University of Science and Technology, Clear Water Bay, Hong Kong, China.|State Key Laboratory of Molecular Neuroscience, The Hong Kong University of Science and Technology, Clear Water Bay, Hong Kong, China.
Kim-Wan
KW
Chan
Division of Life Science, The Hong Kong University of Science and Technology, Clear Water Bay, Hong Kong, China.|State Key Laboratory of Molecular Neuroscience, The Hong Kong University of Science and Technology, Clear Water Bay, Hong Kong, China.
Elaine Yee-Ling
EY
Cheng
Division of Life Science, The Hong Kong University of Science and Technology, Clear Water Bay, Hong Kong, China.|State Key Laboratory of Molecular Neuroscience, The Hong Kong University of Science and Technology, Clear Water Bay, Hong Kong, China.
Estella Pui-Sze
EP
Tong
Division of Life Science, The Hong Kong University of Science and Technology, Clear Water Bay, Hong Kong, China.
Oormila
O
Chandrashekar
Division of Life Science, The Hong Kong University of Science and Technology, Clear Water Bay, Hong Kong, China.|State Key Laboratory of Molecular Neuroscience, The Hong Kong University of Science and Technology, Clear Water Bay, Hong Kong, China.
Guang-Miao
GM
Fu
School of Chinese Medicine, Hong Kong Baptist University, Hong Kong, China.
Zhong-Zhen
ZZ
Zhao
Division of Life Science, The Hong Kong University of Science and Technology, Clear Water Bay, Hong Kong, China. boip@ust.hk.|State Key Laboratory of Molecular Neuroscience, The Hong Kong University of Science and Technology, Clear Water Bay, Hong Kong, China. boip@ust.hk.|Guangdong Key Laboratory of Brain Science, Disease and Drug Development, HKUST Shenzhen Research Institute, Shenzhen, Guangdong, China. boip@ust.hk.
Nancy Yuk-Yu
NY
Ip
D000818:Animals; D002545:Brain Ischemia / Q000378:metabolism* / Q000473:pathology; D002478:Cells, Cultured; D004365:Drugs, Chinese Herbal / Q000737:chemistry / Q000494:pharmacology*; D020244:Infarction, Middle Cerebral Artery / Q000378:metabolism; D008297:Male; D009474:Neurons / Q000166:cytology / Q000187:drug effects*; D018696:Neuroprotective Agents / Q000737:chemistry / Q000494:pharmacology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D015398:Signal Transduction
D004365:Drugs, Chinese Herbal; D018696:Neuroprotective Agents
CREB; ERK; Excitotoxicity; MCAO; NMDA receptor; Neuroprotection; Stroke; TCM