|Budget Amount *help
¥3,400,000 (Direct Cost: ¥3,400,000)
Fiscal Year 2000: ¥1,200,000 (Direct Cost: ¥1,200,000)
Fiscal Year 1999: ¥2,200,000 (Direct Cost: ¥2,200,000)
Excessive release of excitatory amino acids (glutamate), increased generation of reactive oxygen species and free radical (NO) are regarded as important factors in the pathogenesis of cerebral ischemia-reperfusion injury. Reciprocal actions between these factors also involved. Ascorbate, a major antioxidant in the brain, is highly concentrated in neuropils. Its extracellular release is closely related to that of glutamate, and ascorbate-mediated protection from excitotoxins has been demonstrated in vitro. Thus, 1) after establishment of method to measure the dynamics of extracellular ascorbate release in vivo, 2) the relationship between extracellular release of ascorbate and glutamate, 3) the effect of NOS inhibitor (L-NAME), 4) the effect of extended time of ischemia, 5) the effect of postischemic hypothermia (from 36 to 32 ℃) on extracellular ascorbate and glutamate release were investigated in vivo using a microdialysis biosensor system during the early stage of forebrain ischemia-
reperfusion in rat.
METHODS : Two probes of a microdialysis biosensor electrode were inserted stereotaxically in the bilateral striatum or hippocampus of male Wistar rats. Mean arterial pressure (MAP), cortical CBF measured by laser-Doppler flowmeter, and rectal and cortical temperatures were continuously recorded. Forebrain ischemia-reperfusion was performed by bilateral carotid artery occlusion with hemorrhagic hypotension (MAP=30mmHg) for 10 min followed by reperfusion for 60 min.
RESULTS : 1) Using the microdialysis biosensor, continuous real-time measurement of extracellular ascorbate can be obtained in rat striatum in vivo and a novel dialysis electrode can follow the changes in extracellular ascorbate during reperfusion, 2) the marked increase of ascorbate during reperfusion was associated with the rapid decrease in glutamate, 3) in L-NAME treated rats, glutamate was higher than saline treated rats throughout the experiment, 4) the extended time of ischemia (5 min) caused significant inhibition of glutamate re-uptake and ascorbate release during reperfusion, 5) the post-ischemic hypothermia caused significant increase in ascorbate release and glutamate re-uptake during reperfusion along with significantly higher MAP and cortical CBF.
CONCLUSION : These results suggest that 1) the heteroexchange of ascorbate with glutamate, 2) L-NAME did not prevent marked intraischemic glutamate accumulation, moreover, increased its level during reperfusion, 3) the changes in extracellular ascorbate could be considered a marker of glutamate re-uptake during the early stage of reperfusion after forebrain cerebral ischemia, 4) the marked extracellular release of ascorbate by post-ischemic hypothermia consistent with the reported ascorbate-mediated protection from glutamate citotoxicity. Less