Effect of tyrosine kinase receptor binding protein B3 on the synapse-associated protein and synaptic structure in the hippocampus of epileptic rats model
10.3760/cma.j.cn371468-20250321-00106
- VernacularTitle:癫痫模型大鼠海马区酪氨酸激酶受体结合蛋白B3对突触相关蛋白及突触结构的影响
- Author:
Tiantian LIU
1
;
Hengfang LIU
;
Yanjie JIA
Author Information
1. 郑州大学第一附属医院神经内科，郑州 450000
- Publication Type:Journal Article
- Keywords:
Epilepsy;
Tyrosine kinase receptor binding protein B3;
N-methyl-D-aspartic acid;
Postsynaptic density protein 95;
Synaptic plasticity
- From:
Chinese Journal of Behavioral Medicine and Brain Science
2025;34(9):774-782
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To investigate the effect of tyrosine kinase receptor binding protein B3 (Ephrin-B3) on the expressions of postsynaptic density protein 95 (PSD95), N-methyl-D-aspartic acid (NMDA) receptor subunit 2A and 2B(NR2A/NR2B) and synaptic ultrastructure of hippocampal neurons in the dentate gyrus (DG)area of epileptic rats.Methods:A total of 96 SPF-grade adult male Sprague Dawley(SD) rats were divided into blank group, epilepsy group, lentiviral vectors control group and Ephrin-B3 overexpression group( n=24 in each group) according to the random number table.Another 36 adult male SD rats were randomly divided into recombinant adeno-associated virus control group and Ephrin-B3 suppression group( n=18 in each group). The viruses were stereotaxically injected into bilateral hippocampus DG area of rats and epileptic model was established by intraperitoneal injection of lithium chloride and pilocarpine.Racine standard grading method was used to evaluate seizure rate and the duration and latency of epileptic rats were observed. Immunofluorescence staining was used to observe expression levels of PSD95, NR2A and NR2B protein in hippocampus DG area of rats.PCR and Western blot were used to detect the expression levels of PSD95, NR2A and NR2B mRNA and protein in hippocampus of rats. Transmission electron microscopy was used to observe the ultrastructure of hippocampal neurons in rats. GraphPad Prsim 8 software was used for statistical analysis. Results:(1) The results of Ephrin-B3 overexpression experiment: there were statistically significant differences in the seizure latency and duration among the four groups( F=368.30, 120.00, both P<0.01). The seizure latency in Ephrin-B3 overexpression group was longer than that of the epilepsy group and lentiviral vectors control group((31.32±1.10)d, (24.09±2.02)d, (21.42±0.79)d)(all P<0.01), while the seizure duration in Ephrin-B3 overexpression group was shorter than that of the epilepsy group and lentiviral vectors control group((26.85±1.14)s, (40.40±1.26)s, (36.50±5.50)s)(all P<0.05). Immunofluorescence staining showed the average fluorescence intensity of PSD95, NR2A and NR2B in Ephrin-B3 overexpression group were higher than that in the lentiviral vectors control group(all P<0.01). PCR and Western blot both showed that the Ephrin-B3 overexpression group had higher mRNA expression levels of PSD95, NR2A and NR2B than that in lentiviral vectors control group(all P<0.05). The results of transmission electron microscopy showed the number of synapses in the Ephrin-B3 overexpression group was greater than that in lentiviral vectors control group((9.00±1.00), (6.00±1.00))( P<0.05), synaptic gap was narrower than that in lentiviral vectors control group ((31.60±1.45)nm, (36.43±2.22)nm)(all P<0.05). (2)The results of the Ephrin-B3 suppression experiment: the evaluation of seizures in rats showed that there were statistically significant differences in the seizure latency and duration between recombinant adeno-associated virus control group and Ephrin-B3 suppression groups( t=3.88, 3.93, both P<0.05). The seizure latency in Ephrin-B3 suppression group was shorter than that of recombinant adeno-associated virus control group ((17.10±1.88)d, (23.50±2.15)d)( P<0.05). The seizure duration in Ephrin-B3 suppression group was longer than that of recombinant adeno-associated virus control group((55.16±5.48)s, (42.06±1.83)s)( P<0.05). Western blot results showed that the Ephrin-B3 suppression group had lower protein expression levels of PSD95, NR2A and NR2B than those in recombinant adeno-associated virus control group(all P<0.01). The results of transmission electron microscopy showed the numbers of synapses in the Ephrin-B3 suppression group were less than recombinant adeno-associated virus control group((3.33±0.58), (4.66±0.58))( P<0.05), synaptic gap were narrower than recombinant adeno-associated virus control group((37.27±0.97)nm, (33.33±1.46)nm)( P<0.05). Conclusion:Ephrin-B3 can attenuate seizures and upregulate the synaptic related proteins and glutamate receptor expression, improving synaptic structure.
