RESUMO
Objective:To study the effect of β-amyloid(Aβ) on calcium homeostasis and endoplasmic reticulum calcium storage of hippocampal neurons in rats.Methods:A total of 60 adult male SD rats were randomly divided into six groups by body mass matching method, with 10 rats in each group.Three groups were injected Aβ 25-35 into the hippocampus(2 μL per side), and divided into low dose group(2.5 μg/μL), medium dose group(5.0 μg/μL) and high dose group(7.5 μg/μL) respectively, and the other 3 groups were set up as the normal saline group(2 μL 0.9% sodium chloride solution), sham-operated group(rats craniotomy without injection) and normal control group(normal feeding without any treatment). The rats were fed until 14 days after operation, and the behavior and state of the rats were observed and recorded, as well as the body weight and total food intake ratio.And the rat hippocampal cells endoplasmic reticulum pathological change were observed by using the electron microscope and light microscope, meanwhile the concentration of intracellular free Ca 2+ ions was detected by the laser scanning confocal microscope and the expression level of PS, SERCA and RyR mRNA and protein by real-time PCR and Western blot methods respectively.The experimental results were analyzed using SPSS 25.0 software for statistical analysis. Repeated measurement ANOVA and one-way ANOVA were used for multi-group comparison, and Dunnett test and Tukey test were used for further pairwise comparison. Results:(1) The body weight of rats in each group was analyzed by repeated measurement ANOVA, and the difference of time effect was statistically significant( F=153.15, P<0.001), but there was no significant difference between the intergroup effects and interaction effects( F=1.547, P=0.374; F=1.598, P=0.113). The body weight of high, medium and low dose Aβ 25-35 groups at 7, 14 days after injection had no significant difference compared with the control group(all P>0.05). The food utilization rates of the high, medium and low doses of Aβ 25-35 groups were(22.9±4.0)%, (23.0±4.2)% and(22.6±3.2)%, respectively, and there was no significant difference compared with the control group((23.7±5.0)%, P>0.05). Within 14 days after injection, listlessness and lethargy were observed in rats in the high dose Aβ 25-35 group.(2) Pathological observation results showed that the endoplasmic reticulum of rat hippocampal cells in the high dose and medium dose groups of Aβ 25-35 was expanded and swelled, and the mitochondria were swollen and deformed.(3) 14 days after Aβ 25-35 injection, the fluorescence intensity of free calcium in hippocampus of rats in high, medium and low dose groups were(820.43±6.89), (720.12±4.30) and (680.50±4.32), respectively, which were all higher than that in the control group(592.17±3.97)(all P<0.001). (4) RT-PCR and Western blot results showed that compared with the control group, high dose and medium dose Aβ 25-35 injection could up-regulate the expression of PS and SERCA mRNA and protein in hippocampal cells(all P<0.05), while down-regulate the expression of RyR mRNA and protein in hippocampal cells(all P<0.05). Conclusion:The deposition of Aβ 25-35 in hippocampal tissues can disrupt the homeostasis of calcium ions in hippocampal tissues, and then cause the increase of free calcium and its related proteins, thus playing the neurotoxic role.