Study On The Effects And Mechanisms Of Potassium 2-(1-Hydroxypentyl)-benzoate On Animal Senile Dementia

Potassium 2-(1-hydroxypentyl)-benzoate(dl-PHPB) was a new compound synthesized by our institute,which is the prodrug of 3-n-butylphthalide(dl-NBP).dl-NBP is a first-class anti-ischemic stroke drug which had many nervous protective effects for ischemic brain tissue We deduced that it might have therapeutic action for senile dementia.But dl-PHPB has many advantages compared with dl-NBP.In this study,we adopted three dementia models to investigate the improvement of dl-PHPB on the cognitive deficits of dementia animals and observe its possible mechanisms.PARTⅠEffects and mechanisms of dl-PHPB on the learning and memory deficits of cerebral hypoperfusion ratsIn the present study,we administered drugs on day 10 to 30 after permanent occlusion of common carotid arteries(BCCAO) of rats.On the last six days,the behavioural test was performed,and then we determined some biochemical and immunohistochemical indexes in brains of dementia rats.The results are as follows:dl-PHPB 39 and 129mg/kg,dl-NBP 100mg/kg and piracetam 600mg/kg reduced the latencies of dementia rats with different extent.Meanwhile,dl-PHPB 39mg/kg group used more linear and tendency modes than vehicle group.In the probe trial,except for dl-PHPB 129mg/kg and dl-NBP 100mg/kg,other groups spent more time in the target-quandrant than the vehicle,especially dl-PHPB 39mg/kg.In addition,the first crossing-platform time of dl-PHPB 39mg/kg group was significantly shorter than that of vehicle group.The above showed that dl-PHPB could significantly improve rats' cognition,especially large and middle doses.We found that the ChAT activity in hippocampus had significant decreasing trend after BCCAO with biochemistry method,dl-PHPB 129mg/kg significantly increased the hippocampus ChAT activities of ischemic rats.In cortex,the SOD activities of dl-PHPB 13 and 39mg/kg,dl-NBP 100mg/kg,and piracetam 600mg/kg were lower than that of vehicle group.Besides piracetam,other groups showed decrease trend on cortex CAT activity.The GSH-Px activity didn't show obvious differences among all groups,dl-PHPB and dl-NBP significantly decreased cortex MDA content.In hippocampus,cerebral ischemia and the drugs didn't affect SOD and MDA.In addition,there were no significant differences on ATPase activity,and LD and NO content among all groups.HE-staining showed that three doses of dl-PHPB improved the abnormalities in cortex and hippocampus CA1 and CA3 with different extent,especially 39mg/kg.We also found that they had improvement on the pathological changes in corpus callosum and optic tract with K-B staining,especially 39mg/kg.GFAP-positive astrocytes were investigated with immunohistochemistry method.In cortex, there was only a tendancy that GFAP-positive astrocytes in vehicle rats exceeded that of sham-operated rats,but all drugs could reduce astrocytes,especially dl-PHPB 39mg/kg.In hippocampus,all drugs also decreased the astrocytes,especially dl-PHPB 39 and 129mg/kg. In corpus callosum,GFAP-positive astrocytes in dl-PHPB 39mg/kg group were less than that of vehicle group.In optic tract,there were such results similar to that in hippocampus.dl-PHPB 39mg/kg increased the MAP2 area in cortex and hippocampus CA1 region significantly.But the drugs didn't affect the MAP2 density.The BDNF areas in rats' brains of all groups had no significant differences.But dl-PHPB obviously increased the BDNF density in cortex,especially 39mg/kg,but in hippocampus,only dl-PHPB 39mg/kg increased it significantly.PARTⅡImprovement and mechanisms of dl-PHPB on the cognitive deficits of dementia rats induced by Aβ_(25-35)In this study,we adopted dementia rats induced by Aβ_(25-35) through intracerebroventricular injection(i.c.v.).After administering for two weeks,rats' learning and memory was detected with Morris water maze.The related biochemical indexes and phospho-tau protein were examined to elucidate the action mechanisms of dl-PHPB.The results are as follows:On day 9-13 after administration,water maze test was performed,dl-PHPB 39 and 129mg/kg dose-dependently reduced latency compared with the vehicle group.In the probe trial,dl-PHPB 129mg/kg group spent more time in the target-quandrant.In addition,the drug had a tendancy of reducing first crossing-platform time.These showed that large dose of dl-PHPB could significantly improve rats' learning and memory abilities.dl-PHPB decreased the cortex SOD activity and MDA content dose-dependently.But in hippocampus,SOD activity and MDA content had no obvious differences among four groups.dl-PHPB 39mg/kg increased the cortex ChAT activity compared with vehicle group. Meanwhile,dl-PHPB did not affect the cortex AChE activity and there were also no significant differences on hippocampus ChAT and AChE activity among all groups.Meanwhile,we determined the phosphorylated tau protein of Ser396 site with Western blotting,dl-PHPB reduced dose-dependently tau protein expression in cortex.However,it did not significantly affect it in hippocampus.PARTⅢImprovement and mechanisms of dl-PHPB on the cognitive deficits of senescence-accelerated mouse(SAM)1.Comparative study on the learning and memory of different month-age SAM,and brain tissue biochemistry and morpholoyWe investigated the cognition,and brain biochemistry and morphology of five,ten,and fifteen-month-old SAMP8.In step-down test,there was a tendancy that the passive avoidance response ability of five-month-old SAMP8 was lower than that of same-age SAMR1.The abilities of ten and fifteen-month-old SAMP8 were significantly lower than that of same-age SAMR1.The difference on this between SAMP8 and SAMR1 increased gradually from five to ten-month age,but it did not increase from ten to fifteen-month age.In water maze test,the spatial cognitive ability of five-month-old SAMP8 was slightly lower than that of same-age SAMR1,but the abilities of ten and fifteen-month-old SAMP8 were significantly lower than that of same-age SAMR1.We also found that their cognitive abilities decreased gradually from five to ten-month age and did not change obviously from ten to fifteen-month age.The difference on cognitive ability between them increased from five to ten-month age,but did not increase further from ten to fifteen-month age.Decreasing value of fluorescence intensity was used to show mitochondria membrane potential indirectly.The values of three month-age SAMP8 were significantly less than that of same-age SAMR1.We also found that the mitochondria membrain potentials of SAMP8 and SAMR1 decreased gradually and their difference increased slightly with month-age increase.Meanwhile,with the increase of month age,the mitochondria ATPase activity decreased gradually,but it did not decrease further after ten-month age.Moreover,the values of all month-age SAMP8 were significantly smaller than those of same-age SAMR1.The brain ChAT activities of five and ten-month-old SAM were in high level,but it decreased obviously in fifteen-month-old.In addition,the hippocampus ChAT activity of ten-month-old SAMP8 was significantly lower than that of same-age SAMR1.In cortex,there were no differences on AChE activity between them.In hippocampus,the AChE activities of five and ten-month-old SAMP8 were significantly higher than that of same-age SAMR1.But the AChE activity of fifteen-month-old SAMP8 was lower than that of SAMR1. With the increase of month age,the serum SOD activity of SAM decreased gradually.The SOD activity of ten-month-old SAMP8 was slightly higher than that of same-age SAMR1. However,in fifteen-month-old SAMP8,it was significantly lower than that of SAMR1.From five to ten-month age,the MDA level increased gradually,but there was no obvious difference between ten and fifteen-month age.Meanwhile,there was no significant difference between SAMP8 and SAMR1.2.Improvement of dl-PHPB on the cognitive deficits of SAMP8We administered two doses of dl-PHPB to ten-month-old SAMP8 for five weeks.On the last five days,we performed behavioral test to detect animals' cognitive abilities,and determined related biochemical indexes and phospho-tau protein of brain.We found that dl-PHPB improved dose-dependently the passive avoidance response ability of SAMP8 in step-down test.Meanwhile,dl-PHPB reduced the number of entering non-exit and the latency of finding safe steps of SAMP8 in water maze trial.Meanwhile,dl-PHPB reduced the hippocampus SOD activity of SAMP8 and had a strong tendency of decreasing the hippocampus MDA content.On the other hand,dl-PHPB increased dose-dependently the hippocampus ChAT activity and had a tendency of decreasing the hippocampus AChE activity.In addition,large dose of dl-PHPB had weak effect increasing the cortex mitochondria ATPase activity.We determined the tau protein of one phosphorylation site(Ser396) with Western blotting. Compared with control,dl-PHPB decreased dose-dependently the expression of phospho-tau protein in the hippocampus of SAMP8,but it didn't affect it in cortex.Taken together,we confirmed the therapeutic effects of dl-PHPB on VD,Aβ-induced dementia and the cognitive deficits in SAMP8.Meanwhile,these results indicated that such effects were mainly mediated by improving the pathologic changes,reducing active astrocytes, and increasing MAP2 and BDNF of VD rats' brains,alleviating the oxidative stress damage and enhangcing the cholinergic nerve function in brains of three animal models,and reducing the hyperphosphorylation tau protein in brain tissues of the later two animal models.In a word,dl-PHPB affected many pathologic courses of dementia and was a potentially beneficial and promising drug for the treatment of dementia.