| Budget Amount *help |
¥2,800,000 (Direct Cost: ¥2,800,000)
Fiscal Year 2002: ¥700,000 (Direct Cost: ¥700,000)
Fiscal Year 2001: ¥2,100,000 (Direct Cost: ¥2,100,000)
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| Research Abstract |
The measurement of oxygen tension (PO_2) in the brain is quite important in the study of ischemic diseases. To date, most PO_2 measurements have been performed using oxygen microelectrodes. However, the microelectrode is not always a suitable tool for studying brain oxygenation because insertion of the electrode into the brain per se causes hemorrhage and hence leads to erroneous PO_2 measurement. Furthermore, microelectrodes only measure PO_2 in the brain surface as epifluorescent microscopic patterns, using the oxygen-sensitive fluorescent membrane technique with proper modifications
An oxygen-quenching fluorescence dye, tris (1,10-phenanthroline) ruthenium (II) chloride hexahydrate, was immobilized in silica-gel beads 3 um in diameter and the dyed heads were embedded in a highly gas-permeable (〜20 um thick), opaque silicone-rubber film formed on a microscope coverslip. Unlike the original method, intended for transparent mesenteric tissue, he opaque silicone elastomer was adopted to
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minimize the effect of the redox state of the brain tissue on the optical measurement of PO_2. The feasibility of this technique for use in brain microcirculatory studies was assessed using Pulsinelli's four-vessel occlusion rat. Under pentobarbital anesthesia (35 mg/Kg. I.p.), male wistar rats (300-350g, n=8) underwent bilateral electrocauterization of the vertebral arteries. After 24 hours recovery, two cranial windows were made in the skull under anesthesia (15% urethane, 10 ml/Kg, s.c.) and the oxygen-sensitive membrane was placed on the brain, with the PO_2-sensing side in very gentle contactwith the brain surface through one cranial window. Under artificial ventilation, the animals were subjected to occlusion of the common carotid arteries, which resulted in reversible brain ischemia as monitored by a Laser-Doppler flowmeter through the other window. During the ischemia/reperfusion periods, the fluorescence pattern on the oxygen-sensitive membrane was video-recorded continuously and subjected to subsequent playback analysis and conversion to PO_2 values
The present method enabled us to visualize the PO_2 gradient on the rat brain surface without causing cortical injury. In ischemia/reperfusion protocol, heterogeneity in the PO_2 distribution on the brain surface was detected, in particular between the periarteiolar region and other regions, and this heterogeneity was observed to increase in association with ischemia Less
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