SYNAPTOSOMAL NA,K-ATPASE DURING FOREBRAIN ISCHEMIA IN MONGOLIAN GERBILS

We studied the activity and kinetic parameters of synaptosomal Na,K-AT Pase during 15 min of forebrain ischemia and following 60 min of reper fusion produced by reversible common carotid occlusion in Mongolian ge rbils. A synaptosomal fraction was obtained by both differential centr ifugation of brain tissue homogenate and centrifugation of crude mitoc hondrial fraction at a discontinual sucrose density gradient. We found two components of ATP concentration dependence of ATP hydrolysis that represent two types of ATP-binding sites: high affinity and low affin ity. Neither ischemia nor reperfusion affected kinetic parameters of a high-affinity site. However, low-affinity site parameters were affect ed by both ischemia and ischemia followed by reperfusion. Maximal velo city (V-max) decreased by 43 and 42% after ischemia and after ischemia /reperfusion, respectively. The apparent K-m for ATP decreased by 52% after ischemia and by 47% after ischemia/reperfusion. The apparent aff inities for K+ and Na+ were determined from the ATP hydrolysis rate as a function of Na+ and K+ concentrations. We found the half-maximal ac tivation constant for K+ (KaK+) increased by 60% after ischemia and by 146% after ischemia/reperfusion. On the other hand, we found that KaN a+ decreased significantly after ischemia/reperfusion (16%). We conclu ded that it is the dephosphorylation step of the ATPase reaction cycle that is primarily affected by both ischemia and ischemia/reperfusion. This might be caused by alteration of the protein molecule and/or its surroundings subsequent to ischemia.