Changes Of BDNF,NSE On Cochlear Nucleusin Rat Offsprings After Prenatal Stress And Intervention Of Fluoxetine

A lot of evidents have proved that:the early life environment has a far-reaching impact on the development of the fetus, newborn and adult individuals. As social competition intensifies, people are affected by the pressures from their families, work, and environment. Pregnant women, as a special group of people, are paid great attention. Stress during pregnancy is mainly inhibitory stress, and the most common is anxiety and depression. This incidence can reach 16.5%. People have recognized that prenatal stress can not only affect the growth and development of fetal brains, cause certain physical defects of the offspring,which will result to long-term impairment of the offsprings, but also have an influence on the offsprings'auditory capabilities. However, the precise mechanism is unknown.In recent years, it is reported that chronic stress has something to do with the decreation of the expression of brain-derived neurotrophic factor (brain-derived neurotrophic factor, BDNF) both at home and abroad. But the BDNF plays an important role as a protective factor in stress. However, there is not any evidence of diversification of BDNF in prenatal stress leading to the changes of auditory nerve system in the offspring. Neuron specific enolase (neuron specific enolase, NSE) is not only the specific protein of the central nervous system, but also the mostly sensitive indicator of the damage nerve cell, which can be used as the maker of neuronal damage. However, whether the NSE in prenatal stress has changed, whether this change is related to the offspring hearing impairments, there is no report.At the same time,in some researches, it has shown that antidepressant fluoxetine has the effect of treatment and protection to the disease caused by stress. However, fluoxetine for the treatment of hearing impairment in offspring after prenatal stress has not been reported at home and abroad.The stress time is fixed in the required time in the previous restraint stress model and so the animal is easy to be inertia. In this study, the stress time will be randomly assigned in order to make the animal model more perfect. ObjectiveThe part we will completed in this experiment is to build improved rat prenatal restraint stress model, intervened with fluoxetine, observed the changes of BDNF and NSE in the offspring cochlear nucleus tissue by immunohistochemistry, guarded the success of prenatal restraint stress animal model, and further investigated the internal mechanism of offspring cochlear nucleus damage after prenatal stress and the treatment produced by fluoxetine. This will not only provide us a theoretical basis of maternal psychosocial health, but also is of a great importance on prevention and treatment to the disearses caused by prenatal stress.Materials and Methods136 adult female SD rats (210-250g).5 adult male SD rats(270-320g).Male and female rats were caged together, with its full view of sperm found in estrus as day 0 of pregnancy. Female rats were randomly divided into six groups, control group (group A, n=6), pregnancy stress group (group B, n=6), pregnancy stress+high dose of fluoxetine group (group C, n=6, fluoxetine dose of 1.2 mg/kg), pregnancy stress+middle dose of fluoxetine group (group D, n=6, fluoxetine dose of 0.8mg/kg), prenatal stress+low dose of fluoxetine group (group E, n=6, fluoxetine dose of 0.4mg/kg), prenatal stress+saline control group (group F, n=6). Group A is normal pregnancy without any treatment. Use restraint stress model, give the rat restraint stress in the first 1-14 days of pregnancy,3 times a day, each time lasts 45 minutes. Since the first day, each treatment group (C, D, E group) was fed with fluoxetine, by gavage volume of 10ml/kg daily for the first time before 1 hour of restraint stress. Group F was fed with the same volume of saline. Litters were randomly culled to 1-2 offsprings from every nest. According to adult groups, the offspring groups were divided into groups OA, OB, OC, OD, OE, OF with each group of ten.2 The liquid consumption of the pregnant female rats were observed in each group.3 The auditory brainstem response (ABR) of the offsprings (14 days old) were tested before they were sacrificed, recording ABR response threshold,Ⅰ,Ⅱ,Ⅲ-wave latency andⅠ-Ⅱ,Ⅱ-Ⅲ,Ⅰ-Ⅲ-wave intervals.4 Offsprings in each group were killed after the ABR test. Brain tissues were picked out, and specific planes, that is, the cochlear nucleus, were selected to slice. A total of four sets were selected:a set of line HE staining, a line of BDNF immunohistochemistry, a line of NSE immunohistochemical staining, another set as a negative control.5 German Leica image acquisition system was applied. The average optical density value of each group offspring cochlear nucleus BDNF and NSE immunohistochemical staining were analysed by Biosens Digital Imaging Systems V1.6 software (Scientific Instruments Co., Ltd. Shanghai Murayama).6 Statistical analysis of the experimental data was performed with SPSS13.0 software. All data was shown with the mean±standard deviation (x±s). A number of groups of samples were compared using ANOVA. T is used to be the comparison between the two groups. The test level isα=0.05.Results1 After 14 days'stress, the sugar consumption and preference to sugar of prenatal stress group pregnant rats were decreased markedly. Compared with the blank control group, the difference was statistically significant (P<0.01). The sugar consumption and preference to sugar of fluoxetine pregnancy intervention group were increased, in which the increase of 1.2mg/kg group was most obvious. Compared with the prenatal stress normal saline control group, the difference was statistically significant (P<0.01).2 Prenatal stress group offspring ABR response threshold increased.Ⅰ,Ⅱ,Ⅲ-wave latency andⅠ-Ⅱ,Ⅱ-Ⅲ,Ⅰ-Ⅲ-wave interval was prolonged. Compared with the blank control group, the difference was statistically significant(P <0.01). Offsprings'ABR response threshold of the fluoxetine 1.2mg/kg group decreased.Ⅰ,Ⅱ,Ⅲ-wave latency andⅠ-Ⅱ,Ⅱ-Ⅲ,Ⅰ-Ⅲ-wave interval were shortened. Compared with the production pre-stress saline control group, the difference was statistically significant (P<0.01).3 The cell swelling of prenatal stress group offspring cochlear nucleus neurons HE staining significantly reduced, cytoplasm and nuclei enriched, Nissl reduced; the number of neuronal cells of fluoxetine 1.2mg/kg in the intervention group offspring cochlear nucleus HE staining increased. Cells developed normally. The sub-nuclear cells arranged in neat rows. A handful of cytoplasm and nucleus appeared shrunken and vacuolated changes.4 Immune reactants of cochlear nucleus organization BDNF turned brownish-yellow in the light microscope. NSE immune reactants was brown. The optical density value of prenatal stress group offspring cochlear nucleus BDNF and NSE was significantly lower than control group, that is, the expression of BDNF and NSE significantly decreased, and the difference was statistically significant (P<0.01); fluoxetine intervention group offspring cochlear nucleus BDNF and NSE immunohistochemistry increased in average optical density value, namely, the expression of BDNF and NSE increased, in which the difference between the group of fluoxetine 1.2mg/kg, and 0.8mg/kg and the prenatal stress saline control group was statistically significant (P<0.05).ConclusionsWhen prenatal stress happened, the expression of BDNF and NSE decreased in the cochlear nucleus of rat offsprings and the cochlear nucleus of rat offspring was injured. Fluoxetine can increase the expression of BDNF and NSE of the cochlear nucleus of the rat offsprings who have prenatal stress. This can protect the cochlear nucleus of the rat offspring from injury to some extent.