Molecular Pharmacology, Vol 15, 492-505, Copyright © 1979 by the American Society for Pharmacology and Experimental Therapeutics

Specific Binding of [³H]Kainic Acid to Receptor Sites in Rat Brain

¹ Departments of Pharmacology and Experimental Therapeutics, and Psychiatry and the Behavioral Sciences, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21205

The binding of [³H]kainic acid to washed membranes from rat forebrain was saturable with dissociation constants at 2° of 4-16 nM and 27-66 nM for high and low affinity binding, respectively. The Hill coefficients for binding to high and low affinity sites were 0.99 and 0.85, respectively. At 2° [³H]kainic acid receptor binding reached equilibrium by 60 minutes. Dissociation of the ligand-receptor complex at 2° was biphasic. The kinetically-determined dissociation constant for the high affinity binding site corresponded well with the value obtained in equilibrium studies. Of the substances tested, kainic acid and quisqualic acid were the most potent in competing for [³H]kainic acid binding sites. L-glutamic acid was 40-fold less potent than kainic acid itself but 2500 times more effective than D-glutamic acid; dihydrokainic acid was 200-fold less potent than kainic acid at the low affinity site and 1000-fold less active in competing for binding to the high affinity site. Among the poor displacers of [³H]kainic acid (IC₅₀ > 0.1 mM at 50 nM [³H]kainic acid) were D- and L-aspartic acids as well as several proposed glutamate antagonists. In brain, there was considerable variation in the regional distribution of high affinity binding sites: striatum > frontal cortex = hippocampus > cerebellum > medulla-pons; more than 90% of the total specific binding in the cerebellum and medulla-pons was to low affinity sites. Thus, on the basis of the kinetics of binding, differential affinity for competitors and differences in regional distribution in brain, the high and low affinity binding sites represent independent entities. Negligible specific binding was obtained (< 1 fmole/mg tissue) in liver, lung, kidney and intestine. Ablation of the intrinsic neurons of the caudate nucleus by injecting 10 nmole of kainic acid 28 days before sacrifice was associated with a 57-64% reduction in high and low affinity binding in this region.

Note:
ACKNOWLEDGMENTS We thank A. Hunter Thompson for providing excellent technical assistance and Victoria Rhodes, Princie Campbell and Carol Kenyon for preparing the manuscript. We are also grateful to Dr. Morley Hollenberg for his helpful comments and Dr. Kathleen Biziere for her contributions.

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