Investigation Of Mitochondrial Fission And Fusion In Hippocampi Of Rats With Status Epilepticus

Background:Epilepsy is one of the most common neurological disorders. There are about6500-9100thousand epileptic patients in China, and as many as20%～40%epilepsy patients are intractable epilepsy. Recurrent seizures affected the life quality of the patients seriously and cause great economic burden to the patients and the whole society. At present, the mechanisms underlying the neuronal injury caused by seizures have not been clarified. Because of undertaking energy metabolism and signal transduction duties in the cell, mitochondria became the heart of epilepsy research. Mitochondria are highly dynamic organelles. The change of mitochondrial form is controlled by two contrasting progresses named "mitochondrial fusion and fission". Mitochondrial fusion makes mitochondria a long pipe network, while mitochondrial fission collapses the mitochondrial reticular structure, makes mitochondria become fragmented with multiple point and granular short mitochondria. In normal conditions, mitochondrial fission and fusion keep in balance, which is very important to maintain mitochondrial morphology and mitochondrial function. However, in abnormal conditions, such as oxidative stress, ischemia and aging can disturb the balance of mitochondrial fusion and fission. The disorder of mitochondrial fission and fusion is an important factor in the process of neuronal injury in many neurodegenerative diseases, such as Alzheimer’s disease, Parkinson’s disease, Huntington’s disease and Amyotrophic lateral sclerosis (ALS). A variety of studies have confirmed that mitochondrial oxidative stress participate in the pathological process of epilepsy. However, at present, the alteration of mitochondrial fission and fusion in hippocampal after seizures are still unknown.Objectives:To detect the alterations of mitochondrial fission proteins Drp1and hFisl and mitochondrial fusion proteins Mfnl and Opal after SE and to investigate the neuroprotective effect of mdivi-1in SE, as well as discus the possible mechanism.Methods:Healthy adult male Wistar rats (weighing220-250g) were given lithium-pilocarpine intraperitoneally to induce status epilepticus (SE) model. Seizures were allowed to last for60min and then were terminated by administration of diazepam. All the rats were randomly divided into normal control group (N), lithium-pilocarpine induced epileptic group (pilo), mdivi-1+NS group and pilo+mdivi-1group. The pilo group was divided into four subgroups of3,6,24and48h after SE and the pilo+mdivi-1group included the epileptic rats intervened by mdivi-1at24h after SE. Western blot, PCR and Immunohistochemical staining were used to detect the alteration of Drpl, hFisl, Mfnl and Opal in hippocampus of rats after SE. Western blot was used to detect cytochrome c and caspase3expression in hippocampal neurons after SE. Elisa was used to detect the content of8-oHdG and SOD commercial assay kits was used to investigate the activity of SOD in hippocampus of rats after SE. Nissl staining was used to detect the neuronal injury induced by seizures.Research results:Comparing with the normal control group, mitochondrial fission was up-regulated and mitochondrial fusion was down-regulated after SE, shown by that both Drpl and hFisl began to increase at3h after SE (P<0.05), peaked at24h (P<0.05) and still at a higher level at48h after SE (P<0.05). While Mfnl and Opal showed a temporary increase at3h and thereafter decreased sharply, with the bottom level at24h after SE (P<0.05). Mdivi-1significantly decreased the expression of Drpl in hippocampal mitochondria and suppressed the expression of cytochrome c and capase3in hippocampus after SE, as well as attenuating the8-oHdG content and increasing SOD activity after SE. Moreover, mdivi-1increased the surviving neural numbers in hippocampal CA3region after SE (P<0.05)Conclusion:SE induced impaired balance of mitochondrial fission and fusion, shown by the elevated fission and the decreased fusion. Mdivi-1is neuroprotective against SE-induced neuronal injury. Mdivi-1suppressed mitochondrial fission after SE, reduced the oxidative stress, attenuated neuronal apoptosis and neuronal injury and increased the surviving neurons after SE. The possible mechanism may be through the mitochondria/ROS/cytochrome c pathway.