Study Of The Role Of Acid Sensing Ion Channel 2a In Neurological Injury

Neurological damage like ischemic brain injury and spinal cord injury becomes more and more as the economic level develops, human habit changes and traffic break up. These neurological damages not only make huge damage to personal life but also bring huge burden to family and society. And now, there is no effective treatment to ischemic brain damage and spinal cord injury. As reported, after ischemic brain damage and spinal cord injury, there will be locality acidosis caused by inflammation and acid excreta accumulation. As locality acidosis develops, acid sensing ion channels will be activated. There are 7 subunits in ASIC family and they are 1a, 1b1, 1b2, 2a, 2b, 3 and 4. Many reports suggest that ASIC1 a can accelerate neural death after neurological damage. Our study also suggests that ASIC1 a mediates secondry injury of neurons after spinal cord injury. But there are few report mention the function of ASIC2 a after ischemic brain injury and spinal cord injury. So this research is to find the mechanism how ASIC2 a achieve its function after ischemic brain injury and spinal cord injury.Objective:To observe the change of ASIC2 a level after ischemic brain injury and to find the function of ASIC2 a in ischemic brain injury.To observe the change of ASIC2 a level after spinal cord injury and to find the function of ASIC2 a in spinal cord injury.To find the possible mechanism of ASIC2 a in neural disorder.Method1.Use histopathology, behavioristics score, evoke potential, immunofluorescence and TTC staining to expound the change after MCAO and SCI.2.Use western-blotting to observe the change of ASIC2 a level after MCAO and SCI.3.Use lentilentivirus vector to regulate the expression of ASIC2 a, state the role of ASIC2 a in MCAO and SCI by TTC staining, MTT staining, evoke potential and behavioristics score.Results1. After MCAO, rats have a significant edema in brain tissue. The ischemic region enlarges in 24 hours and rats showed a bad behave of circling warking, epilepsy even died. 45% rats after surgery died in 24 hours. Western-blotting showed that ASIC2 a levels increased after MCAO. ASIC2 a level continue increasing, and after 12 hours, the change is significant(p<0.05). After lentivirus infection, the expression of ASIC2 a changes significantly. After ASIC2 a has been down-regulated, barin tissue edema significantly gets worse and HE staining showed shrunken cytoplasm and pyknotic nuclei. TTC staining showed that after MCAO, ischemic area enlarges in 24 hours and gets largest at 24 hours. When ASIC2 a expression has been down-regulated, ischemic area enlarges significantly. The infarct rate in this group gets 37.4% at 12 hours after surgery. But at 24 hours, the infarct area won’t be lager any more. Modified Bederson score showed that 2.3, 2.4 and 3.2 in MCAO group, MCAO+Blank lentivirus group and MCAO+Down-regu group, respectively. Neural function after ASIC2 a has been down-regulated significantly decreased.2. Allen’s SCI model is made by A rod which weighs 10 grams with diameter of 2.5 mm was dropped from 10 mm high onto the exposed spinal cord. Then the peri-injury site will be significant edema. After SCI, rats would loss motor function immediately and recover slowly in 28 days. At injury site(0.5cm), ASIC2 a expression was significantly decreased 12 hours after SCI, and the expression level continued to fall without recovery. On the contrary, Western blot analysis showed that ASIC2 a expression was markedly increased until 7 days after spinal cord injury at peri-injury site(0.5cm each ends). After refected by lentivirus, expression of ASIC2 a changes significantly. After ASIC2 a has been up-regulated, tissue injury decreased, showed light edema and survival neurons increased. On the contrary injury became hard. In the OGD model, both the number of axons and the volume of neurons were markedly reduced, in accompany with disassociation of the intercellular junctions. MTT assay showed that up-regulation of ASIC2 a can relieve this effection and down-regulation of ASIC2 a can aggravate it. SEP showed normal rats’ incubation period is 5.5 ms. SCI can make this incubation period 21 ms. Up-regulation of ASIC2 a can shorten the incubation period to 15.3 ms and on the contrary it will be 25.6 ms. BBB score showed sham rats 21 points, SCI+blank virus 1 and 10.0±0.2 points at 1 and 28 day after surgery. After down-regulation of ASIC2 a, rats got 0 piont at 3 day after surgery and 5.8±0.1 points at 28 days after surgery. After expression of ASIC2 a has been up-regulated, score would be 2.6 and 12.6±0.2 at 3 and 28 days after surgery.3. ASIC2 a can increase the tolerance of neurons to OGD. And it can prevent Ca2+ from flowing inside neurons.Conclusion1. After down-regulation of ASIC2 a, MCAO made lager infarct area and animal showed more hemiplegia. Animals with lower ASIC2 a level showed worse moter and sensing function after SCI, tissue injury became worse and neurons got more sensitive to oxygen deficit. On the contrary, up-regulate the expression of ASIC2 a can remit these symptoms and made neurons survive from oxygen deficit.2. ASIC2 a may be a endogenic neurological protective factor. It can protect neurons from ischemic brain injury and spinal cord injury. ASIC2 a may be a new therapeutic target to ischemic brain injury and spinal cord injury in clinical works.