The Effect Of Donepezil Hydrochloride On The Changes About Extracellular Signal-regulated Kinase In The Hippocampus Of Mice With Vascular Dementia

Objectives: Vascular dementia (VD) is a clinical syndrome of intelligence and recognition impairment caused by cerebrovascular diseases (CVD). Because of the proportion of aged people in the population increases and the changes of spectrum of diseases as well as the diagnostic level elevates day by day, its morbility presents an obviously increase tendency, which gives a heavy threaten to the physical and mental health of the aged people, and a heavy burden of spirit and economy to the families and society. The World Health Organization has wrote it on the list of the important study items in 21 century. At present, the pathogenesis of VD has not been fully explained, so, approach the pathogensis of VD and use nootropics on the base of the treatment of primary disease, can be hoped to improve the quality of VD patients'lives. Extracellular signal-regulated kinase (ERK)is one of the important members in the family of mitogen-activated protein kinases (MAPKs). Initially, it is considered to be concerned with growth, development and differentiation of cells, but recent studies indicate that ERK also has a close relationship with learning and memory. It is known to all, the important syndrome of VD is the disorders of learning and memory. Hippocampus is an important structure concerned with learning and memory, then, what are the changes about ERK in the hippocampus of VD mice? We haven't seen the reports yet. In this study, the changes of ERK1, ERK2 and p-ERK in the hippocampus and the effect of donepezil hydrochloride (nootropics, a cholinesterase inhibitor) were observed through established VD mice models. We provided the experimental evidence for clinical medication as well as revealed the pathogenesis of VD.Methods: This study was made up of two sections. Section one was to establish VD mice models. Kunming mice were subjected for three times ischemia-reperfusion through the ligation of the bilateral commom carotid arteries, accompanied by sham group and drug group (donepezil hydrochloride). The capabilities of learning and memory of mice were evaluated by step-down test and water maze test, and the behavioral differences were observed after mice had been bred for 28 days. In section two, the mice's cerebral tissue were fixed up via perfusion of 4% paraformaldehyde solution, and dyed by immunohistochemistry technique. Then we respectively counted positive neurons of ERK1 and ERK2 in the same area of hippocampus CA1, and measured average optical density of the positive neurons with Image-Pro Plu software; We measured average optical density in hippocampus CA3 to observe the expression of p-ERK in each group. One-way analysis of variance was carried out with SPSS 10.0 statistics software, LSD-t test was used to compare the differences among the three groups. P<0.05 meant statistical significance.Results: (1) Step-down test: The behavioral grades of learning and memory in model group decreased evidently, which were reflected by the prolongation of response period (P<0.05) and the increase of error times (P<0.05) during the learning phase, and by the decrease of latent period (P<0.05) and the increase of error times (P<0.05) during the memory phase. However, the records of learning and memory in drug group improved obviously compared with model group (P<0.05), and no significant distinction was discovered between drug group and sham group (P>0.05). (2) Water maze test: The notably reduced records of learning and memory in model group were shown through the prolongation of whole course swimming time and the increase of error times during both learning phase (P<0.05) and memory phase (P<0.05). Whereas, the records of drug group improved obviously compared with model group (P<0.05), and no significant distinction was discovered between drug group and sham group (P>0.05). (3) HE stainning in hippocampus: The arrangement of pyramydal neurons in hippocampus CA1 in sham group was tight and in order, nucleus were large and round, and the chromatospherites were evident; In VD model group, the arrangement of pyramydal neurons was loose and not in order, pyramydal neurons lacked and inflammatory cells infiltrated, cell bodies shrinked, karyopycnosis and cytoplast dyed densely; The arrangement of pyramidal neurons was better than model group in drug group, the phenomenon of karyopycnosis and inflammatory cells infiltrate was less. (4) The changes about ERK1 and ERK2 in hippocampus CA1: In sham group, the layer in hippocampus CA1 was clear, there were abundant positive neurons, the positive substance located mainly at cell membrane and intracytoplasm, the positive neurons ranged tightly, brown and yellow, the cell bodies were round or ellipse, and prominences were observed in some bodies; However, in model group, the number of positive neurons was obviously decreased, the pyramidal neurons ranged loosely, their bodies shrinked and karyopycnosis; In drug group, the number of positive neurons increased, ranged tightly and cell bodies were round compared with model group. Through analyzed, we discovered that the number of positive neurons of ERK1 and ERK2 in the same area of hippocampus CA1 and the average optical density of positive neurons in model group was less than that in sham group(P<0.05), this condition was improved in drug group compared with model group(P<0.05), and no significant distinction was found between drug group and sham group(P>0.05). (5) The changes about p-ERK in hippocampus CA3: The positive substances of p-ERK mainly located at CA3 stratum radiatum (dendrite), the color was dark brown, dendrites were dense, and collateral branches were abundant in sham group; But they were dyed lightly in model group, dendrites were sparse, collateral branches were less, and the average optical density was less than that in sham group(P<0.05); This situation was obviously improved in drug group(P<0.05), and there was no significant distinction between drug group and sham group(P>0.05).Conclusions: (1) This study successfuly established the VD mice models, which could imitate the clinical intelligence impairment of VD patients, the models were confirmed by step-down test and water maze test, and their pyramidal cells in hippocampus CA1 were damaged. At the same time, the effect of donepezil hydrochloride was proved in the study. (2) It was confirmed that the decreased expression of ERK1, ERK2 and p-ERK in hippocampus was related to the cognitive impairment of VD mice, donepezil hydrochloride could increase the expression of ERK1, ERK2 and p-ERK, and improve the capabilities of learning and memory by many pathways.