Specific High‐Affinity Binding of L‐[ 3 H]Aspartate to Rat Brain Membranes

The binding of L‐[ 3 H]aspartate was investigated in washed membranes prepared from whole rat brain. We were able to differentiate two separate binding sites differing in their Na dependence. The Na‐independent binding was saturable, reversible, and optimal at 20°C and at pHs in the neutral range. The dissociation constant (K d ) at 20°C was about 200 n M . This binding site seemed to be modulated by magnesium and calcium at physiological concentrations. None of the amino acids tested was a potent competitor for Na‐independent L‐[ 3 H]aspartate binding. This binding site was unevenly distributed in the rat central nervous system: cerebellum = cerebral cortex > ponsmedulla > spinal cord. Destruction of the intrinsic neurons of the cerebellum by injecting kainic acid 30 days before sacrifice resulted in a 53% reduction in Na‐independent binding in this region. The Na‐dependent binding of L‐[ 3 H]‐aspartate (K d = 484 n M ) was strongly inhibited by D‐aspartate, L‐glutamate, D,L‐aspartate β‐hydroxamate; was unaffected by calcium and magnesium; and showed a different pattern of distribution: cerebral cortex > cerebellum = pons‐medulla = spinal cord. This binding in cerebellum was unaffected by injections of kainic acid.