Expression Changes Of NR2A、NR2B、GluR1、PKC-γ In The Hippocampal Tissue Of Diabetic Rats Induced By STZ

[Objective] To investigate the localization and expression changes of NR2A, NR2B, GluR1and PKC-y in the hippocampus of diabetic rats. development process of cognitive impairment in diabetes (CID). The study would help to better understand the pathogenesis of CID.[Methods] Seventy five male SD rats were randomly divided into4groups:15rats were used as the normal control group (NC),20rats each were used as experimental groups at different time points after diabetes mellitus (diabetes mellitus for3/6/9weeks, DM3/6/9W). In the experimental groups, type1diabetes mellitus was induced by large-dose of STZ (65mg/kg of body weight). Meanwhile, the control group rats were given an equal volume of lemon acid buffer. The learning and memory ability of each group rats was assessed with the Morris water maze test. The brain tissue sections of each group were stained with Hematoxylin-Eosin (HE) and Immunocytochemistry. The hippocampal tissues were subjected to Western blotting analysis for NR2A、NR2B、 GluRland PKC-y expression.[Results] Morris Water Maze of learning scores (escape latency):the escape latency was increased gradually in the course of development of DM. The escape latency at DM9W was to some extent decreased compared to the DM6W. The escape latency showed a significant increase at DM6and9W (P<0.01) in comparison to the NC. It showed a significant increase at DM6W (P<0.01or P<0.05) compared to DM3and9W). Morris Water Maze of learning scores (the frequency in crossing the platform):the frequency in crossing the platform was reduced gradually following the course of development of DM. It was moderately increased at DM6W. In comparison to the NC, the frequency in crossing the platform was significantly decreased (P<0.01) at DM3,6and9W. There was no significant change in the number of crossing the platform at DM3,6and9W (P>0.05) through pairwise comparison. Morris Water Maze of learning scores (the time of staying at the target area):the time of staying at the target area was shortened gradually through the course of development of DM. It was not reduced siginficantly at DM3W (P>0.05) compared with that of NC. The duration of staying at the target area was not reduced siginficantly at DM6W (P>0.05) compared with that of DM9W. The time of staying at the target area was reduced significantly at DM6and9W (P<0.01or P<0.05) compared with that of NC and DM3W. HE staining showed that there was no significant histological change in the hippocampus of DM3W. However, pathological changes were evident with the progression of the disease. Histological alteration included loss of hippocampal pyramidal neurons, disarrayed neuronal arrangement and necrosis. By immunohistochemistry, NR2A cells, NR2B cells, GluRl and PKC-y were mainly localized in the hippocampal pyramidal layer. NR2A, NR2B, GluRl and PKC-y protein expression was weak in the NC rats, but the expression of these markers was gradually enhanced following the course development of DM. Image analysis of immunohistochemical stained sections showed that NR2A, NR2B, GluR1and PKC-γ protein expression was significantly increased in gray intensity level at DM3,6and9W (P<0.01) compared to NC level; NR2A protein expression was significantly increased in gray intensity level at DM9W (P<0.01) in comparison to the DM3W levels; NR2B, GluRl and PKC-y protein expression showed a significant increase in gray intensity level (P <0.01) through pairwise comparison. The results of western blotting showed that the NR2A, NR2B, GluR1and PKC-y protein expression levels were weak in the hippocampal tissue of NC rats. On the other hand, the expression levels of these markers were increased gradually following the course development of DM. By western analysis, NR2A, NR2B and GluR1protein expression showed significant increase in optical density at DM3W (P<0.05) compared to NC level; NR2A, NR2B, GluR1and PKC-γ protein expression showed a significant increase in optical density at DM6and9W (P<0.01or P<0.05) compared to NC level; NR2B and GluR1protein expression showed significant changes in optical density at DM3,6and9W (P<0.01or P<0.05) through pairwise comparison.NR2A an dPKC-γ protein expression was not significantly altered in optical density at DM3,6and9W (P>0.05) through pairwise comparison.[Conclusions]1. In the CID, learning and memory behaviors displayed different degrees of reduction with the development of diabetes and over a protracted duration.2.NR2A, NR2B, GluRl and PKC-y protein expressions in the hippocampal tissue were increased gradually following the course development of DM. It is suggested that the four markers have close relationship or are linked to the occurrence and development process of CID. They would be useful molecular biomarkers either collectively or individually separately to serve for early prediction and diagnosis of CID.