Effect Of Maternal Deprivation On Learning And Memory Ability And Hippocampal Nerve Regeneration Of Hypoxic-ischemic Brain Damage Rats

ObjectiveTo observe the effect of maternal deprivation(MD) on long-term behavior development, hippocampal pathology, hypothalamic pituitary adrenocortical axis, and protein or nuclein associated with hippocampal nerve regeneration of hypoxic-ischemic brain damage(HIBD) rats, to investigate the influence of early MD on HIBD rats and the possible neuromechanisms, and provide theoretical basis for guiding the clinical work and early intervention.MethodsThe 7-day old SD rats' models of HIBD were established by the method of Rice,they were subjected to the right carotid artery ligation and put into the hypoxia device with 80 m L /L air mixture(oxygen and nitrogen) for 2.5 h. Then the rats with HIBD were divided randomly into 4 groups: control female group(CF), control male group(CM), maternal deprivation female group(MF), maternal deprivation male group(MM). In addition, the sham-operation female group(SF) and the sham-operation male group(SM) were set up by only subjecting to the right carotid artery separation without ligation and systemic hypoxia. The MD group rats were deprived from their mother 3 hours per day from the second day after modeling to the 14 th day. After 28 postnatal days, Morris water maze was used to evaluate the learning and memory ability of rats, and elevated plus maze(EPM) was used to evaluate anxiety level of rats.HE staining and nissl staining were employed to observe the moophorlogical changes and the number of neurons in right hippocampus of rats. The immunohistochemical stain and RT-PCR were respectively used to detect the number of nestin/AQP-4positive cells and the level of their m RNA in right hippocampus of rats. Western-blot was used to detected nestin protein expression in right hippocampus of rats. The enzyme-linked immunosorbnent assay was used to detected the serum concentration ofcorticotropin-releasing hormone(CRH), adrenocorticotropic hormone(ACTH), and corticoserone(CORT).Results1. Morris water maze test results showed that, rats of group MF and group MM had lower scores than those of group CF and group CM(P<0.05) about incubation period and times of penetrating the platform, and rats of group SF and SM had higher scores than those of group CF and group CM(P<0.05); by intra-group comparison,there were no significant difference in group MF and group MM(P>0.05).2. The elevated plus maze test showed that, the scores of group MF and group MM were significantly lower than those of group CF and group CM(P < 0.05), and the scores of group SF and group SM were higher than those of group CF and group CM(P < 0.05); by intra-group comparison, the scores of group CF were higher than that of group CM(P<0.05), and scores of MF were higher than that of group MM(P<0.05).3. ELISA test results showed that, serum of rats in group CF and group CM had higher contents of ACTH, CORT and CRH than those of group SF and group SM(P<0.05); the contents of group MF and group MM were higher than those of group CF and group CM(P<0.05); by intra-group comparison, contents of group CF were higher than that of group CM(P<0.05), contents of group MF were higher than that of group MM(P<0.05), serum hormone of female rats had a higher level than that of male rats(P<0.05).4. The results of HE stain showed that the pathology in right-sided hippocampus in group SF and group SM were normal and neurons were well-arranged, and that in group CF and group CM were minimal abnormal, the neurons almost arranged orderly and remained normal. While the cell morphology in group MF and the group MM were obviously abnormal, the neurons arranged disorderly, many neurons lost.5. Nissl stain of nerve cell showed that in group MF and MM, the surviving numbers of nerve cell in right hippocampus of brain were less than those in group CF and group CM(P<0.05), the numbers of group SF and group SM were higher than those of group CF and group CM(P<0.05); by intra-group comparison, there were no significant differences in group MF and group MM(P>0.05).6. Immunohistochemical staining showed that, in group CF and group CM,numbers of nestin positive cell in group CF and group CM were higher than that of group SF and group SM(P<0.05), numbers of group MF and group MM were lower than that of group CF and group CM(P<0.05); numbers of AQP-4positive cell in group CF and group CM were higher than that in group SF and group SM(P<0.05),numbers of group MF and group MM were higher than that of group CF and group CM(P<0.05); the difference between group MF and group MM was significant(P<0.05).7. RT-PCR detection results showed that the expressions of nestin m RNA in group MF and group MM were significantly lower than that of group CF and group CM(P<0.05), while the expressions of m RNA AQP-4 expression in group MF and group MM were significantly higher than that of group CF and group CM(P<0.05).8. Western-blot results showed that transcript level of nestin in group MF and group MM were lower than that in group CF and group CM(P<0.05); the difference between group MF and group MM was significant(P<0.05).Conclusion1. In tests of water maze and elevated plus maze, scores of rats in group MD were lower than those of group CF and group CM, scores of group CF and group CM were lower than those of group SF and group SM, which indicated that MD aggravated the damage of anxiety and long-term learning and memory ability of HIBD rats; in the elevated plus maze test, male rats performed poorer than female rats, which showed that male rats suffered from anxiety more seriously than female male, and indicated that early stress events had a greater influence on male rats.2. In comparison with group CF and group CM, the contents of CRH, ACTH and CORT in rats serum of group MF and group MM raised, which stated that as stress events, MD caused the modification of HPA axis function; hormonal content in the serum of female rats was higher than that of male rats, which presented that as stress events, MD or only separating right common carotid artery would have a greater influence on HPA axis function of male rats, or transformation of HPA axis function made female rats more sensitive to stress events than male rats; by combining with the results of the elevated plus maze test, the ability of change and adjust anxiety offemale rats was stronger than that of male rats.3. In comparison with group CF and group CM, the pathological lesions in right hippocampus of rats of group MF and group MM were more serious, the surviving numbers of nerve cell in right hippocampus decreased, protein expression of nestin group related to nerve regeneration reduced, protein expression of AQP-4 group increased. After MD, regeneration of right hippocampal neurons was restrained, which was not good for brain damage. The restraint of nerve regeneration might be one of the important mechanisms for MD to aggravate the damage of learning and memory ability of HIBD rats.