摘要目的 探讨慢性点燃癫癎大鼠脑内腺苷受体A1(AIR)及A2a(A2aR)表达的动态演变,评价腺苷受体表达改变在癫癎发病机制中的作用.方法 选用氯化锂-匹鲁卡品慢性点燃小鼠模型,分为模型组、对照组和正常组.采用逆转录PCR(RT-PCR)、免疫荧光及Western blot监测发作后24 h,1、6个月时,脑内AlR、A2aR表达的动态改变.结果 点燃后24 h,小鼠脑内A1R表达增高[受体mRNA(1.1483±0.1182);Western blot(0.7872±0.0621);免疫荧光阳性细胞(76.17±4.62)个/高倍镜视野],A2aR表达降低[受体mRNA(0.8338±0.0572);Western blot(0.2098±0.0257);免疫荧光阳性细胞(43.83.4-5.12)个/高倍镜视野],与正常组及对照组比较差异均有统计学意义(P<0.05).在点燃后1个月、6个月时,A1R表达较点燃前下降,A2aR则明显增高,差异亦具有统计学意义(P<0.01);Jg与1个月时比较,6个月时A1R表达更低[受体mRNA(0.5682±0.0443);Western blot(0.7749±0.0262);免疫荧光阳性细胞(38.50±4.81)个/高倍镜视野],A2aR表达更高[受体mRNA(1.2169 ±0.0332);Western blot(0.7080±0.0371);免疫荧光阳性细胞(114.50 ±4.04)个/高倍镜视野],即慢性期A1R表达降低,A2aR表达增高,且随时间延长越来越明显(t=-19.02-13.28,P<0.05).结论 腺苷受体表达失衡在点燃过程中呈现双向改变特征,急性期适应性调整以最大限度抑制癫癎发作,慢性反复发作则导致A1R及A2aR的表达失衡,可能是导致癫癎发作无法控制的重要因素.

abstractsObjective To investigate the dynamic changes of adenosine receptors, A1 (A1R) and A2a (A2aR) in the brain from the acute to chronic phase after kindling and to explore the correlation between seizure and expression level of A1R and A2aR. Methods Rats were randomly selected into the testing model, reference and normal control groups. Testing rats were kindled by lithium choride-pilocarpine, reference rats were treated with saline, and no treatment was given in normal control group. The dynamic expression of A1R and A2aR were detected by RT-PCR, immunofluorescence staining and Western blot at time-points of 24 hours, 1 month and 6 months post-kindling. Results In the acute phase of 24 hours after kindling, the A1R expression level (mRNA level was (1. 1483 ±0. 1182); Western blot result was ( 0. 7872± 0. 0621 ) ; immunofluorescence staining count was ( 76. 17 ± 4. 62 )/HP) was increased and A2aR (mRNA level was (0. 8338±0. 0572) ; Western blot result was (0. 2098 ±0. 0257) ; immunofluorescence staining count was (43. 83 ± 5. 12 )/HP) was decreased. The results showed statistically difference compared with the reference and normal groups (P< 0. 05 ). In the later chronic phase of 1 month and 6 months after kindling, the expression level of A1R was decreased and A2aR was increased. These data revealed statistically significant difference (P <0. 01 ). Furthermore, the comparison of the results in 1 month and 6 months after kindling found that the expression of AIR was lower in 6 months (mRNA level was (0. 5682 ±0. 0443) ; Western blot result was (0. 7749 ±0. 0262) ; immunofluorescence staining count was (38. 50 ±4. 81 )/HP) than in 1 month and that of A2aR was higher in 6 months (mRNA level was (1. 2169±0. 0332) ; Western blot result was (0. 7080 ±0. 0371 ); immunofluorescence staining count was (114. 50 ± 4. 04)/HP). The differences were statistical significant (t = - 19. 02--13.28, P < 0. 05). Conclusions The expressions of A1R and A2AR during and after kindling presents a bidirectional change. In the acute phas the expression of AR is regulated to suppress seizures. While in the chronic phase, the repeated seizures result in the change of A1R and A2aR expression in the opposite direction. This mechanism plays an important role in refractory seizures.