| Objective To investigate the expression of NMDA receptor type 2 receptor B (NR2B) and extracellular signal regulated protein kinase (ERK) signal transduction pathway and the significance in the hippocampus of rats after epileptic seizures induced by penicillin (PEN), detect serum levels of tumor necrosis factor (TNF), interleukin(IL)-1, IL-6 and IL-8 by radioimmunoassay, and clarify the possible antiepileptic mechanism of oxymartrine ( OXY) in the pathogenesis of epilepsy. Methods 1. Sprague Dawley rats were randomly divided into PEN group, sodium phenobarbital (PBS)+PEN group, and oxymatrine+PEN group. Histopathological damage and ultrastructure of morphological deformation of neurons in CA1 and CA3 of the rat hippocampus were observed through HE staining. Changes in p-ERK1/2 and NR2B protein expression in the hippocampus were evaluated at 1, 2, 4, 6 and 8h after seizures by immunohistochemistry staining and Western blot. 2. The content of serum TNF, IL-1, IL-6 and IL-8 was measured by radioimmunoassay in PEN rats. Results 1. Histopathological damage in hippocampal neurons: Light microscopic observation of the HE stained hippocampal CA1 and CA3 showed that the neurons were round or oval in shape. Neurons in PEN group were most seriously damaged, presenting as necrosis and condensation of nuclei with vague nucleoli, and varying degrees of degeneration and necrosis of neurons. Some degenerated and necrosed neurons were seen in hippocampal CA1 and CA3, while the degree of damage in OXY and PBS groups was much lower. 2. Immunohistochemical results of phosphorylated extracellular signal-regulated protein kinase (p-ERK1 / 2) and NR2B: Strong immunoreactivity of p-ERK1/2 was detected in hippocampal neurons of PEN-treated rats at 1h after penicillin injection. In addition, p-ERK1/2 began increasing at 1h after penicillin injection, reached the peak at 2h, began declining at 4h, and descended to the lowest level at 8h. The number of immunoreactive position cells of hippocampal CA1 and CA3 in PBS and OXY groups were lower than that of PEN group (P﹤0.05). There was no significant difference in the number of immunoreactive position cells of hippocampal CA1 and CA3 between PBS and OXY groups. NR 2B began increasing at 1h, reached the peak at 6h, and declined slightly at 8h. The number of immunoreactive position cells of hippocampal CA1 and CA3 in PBS and OXY groups was significantly lower than that in PEN group (P﹤0.05). There was no significant difference in the number of immunoreactive position cells of hippocampal CA1 and CA3 between PBS and OXY groups. 3. Western blot of p-ERK1 / 2 and NMDA receptor type 2 receptor B (NR2B): The results of Western blot showed that p-ERK1/2 and NR2B in the hippocampus of epileptic rats increased significantly as compared with those in the control rats (P<0.05) at the different designated time points. The expression of p-ERK1/2 was higher in 2h group than that in the other groups. The expression of p-ERK1/2 decreased to some extent in PBS and OXY groups compared with that in PEN group. The expression of NR2B was higher in 6h group than that in the other groups. The expression of NR2B decreased to some extent in PBS and OXY groups compared with PEN group. The results were similar to those obtained by immunohistochemical staining. 4. The content of serum TNF, IL-1, IL-6 and IL-8: The content of serum TNF, IL-1, IL-6 and IL-8 in OXY group was significantly lower than that in PEN group(P﹤0.05).There was no significant difference between PBS and OXY groups. Conclusions 1. Oxymartrine could obviously prolong the latency of epileptic seizure, and reduce seizure degrees compared with penicillin group. 2. Following PEN -induced seizure, activation of ERK in rats in the hippocampus changed at different time points, probably because OXY inhibited the ERK signal pathway, thus reducing the expression of NR2B and changing the process of pathologic and physiologic reactions in PEN-induced epileptic seizure. 3. OXY could reduce the content of serum tumor necrosis factor, interleukin-1, interleukin-6 and interleukin-8. Cytokines may take part in the Oxy antiepileptic mechanism... |
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