Ultracytochemical study of Ca2+-ATPase activity in isolated, superoxide-treated rat brains

The relationship between changes in Ca2+-adenosine triphosphatase (ATPase) activity and plasma membrane damage was histochemically studied by enzymatic electron microscopy in rat brains with and without superoxide treatment. The brains were obtained from male Wistar rats after decapitation, and the control brains were examined immediately. Brains not treated with superoxide were incubated at 20 degrees C for 3, 6, or 12 hours. The superoxide-treated brains were immersed in a hypoxanthine-xanthine oxidase system for 20, 60, or 120 minutes. Ca2+-ATPase activity in the cerebral cortex and hippocampal CA1 was studied by the lead citrate method. Control brains showed Ca2+-ATPase activity in the membrane of the nerve cell body and dendrites, the basement membrane of endothelial cells, and the erythrocyte membrane. In untreated brains, enzymatic activity gradually decreased but was still detected after 12 hours. In those treated with superoxide, enzymatic activity gradually decreased but was still observed after 120 minutes in fragments of the plasma membrane. These findings show that the plasma membrane is more affected by treatment with superoxide than by decapitation itself, and that plasma membrane damage precedes the disappearance of Ca2+-ATPase activity. Residual Ca2+-ATPase activity does not necessarily imply reversibility of cerebral ischemia.