Effects of GBEE on memory impairment induced by scopolamine in mice and activity of AChE in cerebral cortex

OBJECTIVE The fruit of Ginkgo biloba L.is also known as Ginkgo nuts. Ginkgo has the effect of warming the lung, boosting qi, downbear phlegm, dispersing toxin, kil ing worms and etc, which also recorded in ancient books on tumor treatment. The scientific name of succulent skin is exocarp in Gingko nuts. Research shows that GBEE(Ginkgo biloba exocarp extracts) has anti-tumor,anti-aging and immune promoting activity. As an M receptor blocker, scopolamine can block the excitatory effect of acetylcholine on M receptors, causing learning and memory dysfunction. It can partially simulate some features of the cholinergic system of Alzheimer disease(AD). The learning and memory impairment model of scopolamine is the classic screening model for AD drug research. In this paper, the effects of GBEE on scopolamine induced learning and memory impairment in mice and its mechanism were preliminarily studied. METHODS GBEE was prepared by the patented method(patent: ZL201610916394.4). Infrared Spectroscopy(IR) analysis shows that the proteoglycan in GBEE is ester linkage. The total content of proteoglycan in GBEE was 62.6%-64.8%, which was measured by phenol sulfuric acid method and brilliant blue method. Pre-column derivatization high-performance liquid chromatography(HPLC) detection showed that the proteoglycan in GBEE contains 6 kinds of monosaccharides including mannose, rhamnose, galacturonic acid, glucose, galactose, arabinose and 14 kinds of amino acids including aspartic acid, glutamic acid, glycine,serine, threonine, alanine, proline, valine, methionine,isoleucine, leucine, phenylalanine, tryptophan and lysine.60 ICR mice, half male and half female, 6-8 weeks old and weight(18-22) g, were randomly divided into six groups: blank control group(normal saline), model group(normal saline), positive drug group( donepezi 1.25 mg·kg-1)and GBEE low-does(50 mg·kg-1), medium-does(100 mg·kg-1) and high-does(200 mg·kg-1) groups. The drug were infused to stomach of mice once a day for 14 d,after that model of learning disorder and dysmnesia were made by intraperitoneal injection of scopolamine hydrobromide(3 mg·kg-1) in six groups except the blank control group on days 15-19. Morris water maze experiment was performed by using the mice′s instinct of escaping from the water to find rest platform. The mean escape latency(s), in other words, the time needed for searching platform was detected on days 15-18, the time in platform annulus(s) was detected on day 19. Each test was performed 30 min after intraperitoneal injection and 1 h after intragastric administration. The mean escape latency(s) and the time in platform annulus(s) were correlated with the memory ability of mice, the former was negatively correlated, the latter was positively correlated, so as to evaluate the changes of learning and memory ability of each group of mice. After the behavioral experiment, the brain tissues of mice were taken out immediately, and the cerebral cortex and hippocampus were cut and stored in the refrigerator at-80℃ with the help of the mouse brain mold. During the test, the brain homogenate was prepared by ultrasonic method, and the activity of acetylcholin esterase(AChE) in the cerebral cortex and hippocampus of each group of mice was measured according to the instructions of AChE test box. RESULTS Scopolamine can significantly prolong the mean escape latency(s) of Morris water maze experiment in mice, and shorten the time in platform annulus(s), which is statistically significant compared with the control group(P<0.05,P<0.01). The results showed that scopolamine caused the decrease of learning and memory ability in the model group. The time required to search for the platform in mice of positive drug group and the GBEE of the three dose groups was significantly reduced on the third and fourth days of Morris water maze experiment test. Except for the GBEE low-dose group, the comparison with the model group was statistically significant(P<0.05, P<0.01). On day 19, positive medicine and three dose groups GBEE can significantly extend the time in platform annulus(s), compared with model group(P<0.05,P<0.01), the effect of medium-dose GBEE group is better than that of high dose, close to the positive drug, the results showed that it could improve the learning and memory ability of scopolamine-induced memory impairment modle mice. While the memory ability of the scopolamine model group was decreased, AChE activity in the cerebral cortex of the mice was significantly increased compared with the control group(P<0.05). Donepezil and GBEE in each dose could reduce Ach E activity in the cerebral cortex of the mice. Except for the high-dose GBEE group, there were significant differences between the other groups and the model group(P<0.05). The results of AChE activity in hippocampi need to be repeated further because of the large difference. CONCLUSION GBEE can improve the learning and memory ability of scopolamine induced memory impairment model mice,and the mechanism may be related to reducing the activity of cerebral AChE and reducing the inactivation of cholinergic neurotransmitter ACh in memory impairment mice.AD is a kind of neural degenerative disease, which can easily be seen in the middle-aged and old people, the early disease mainly for hypomnesis, dysmnesia and so on, accompanied by the reduction of acetylcholine(ACh), the increasing of AChE activity. Early stage of AD is the best treatment stage, by improving the memory function, which can effectively prevent the further development of AD, slow disease progression and reduce disease severity. This study shows that GBEE has certain potential in the treatment of AD, which lays a foundation for further research on the effect and mechanism of GBEE on AD.