The binding properties of the 1,4-dihydropyridine calcium channel antagonist, [3H](+)PN 200-110, were studied in rat cerebral cortical and cardiac homogenates (37 degrees C, Krebs phosphate buffer). Specific binding of [3H](+)PN 200-110 was saturable, reversible, and of high affinity (Kd values are 35 and 64 pM for the cerebral cortex and heart, respectively). In parallel studies with [3H](+)PN 200-110, the dissociation constant of [3H]nitrendipine was 10-12 times higher. Substituted dihydropyridine calcium channel antagonists and agonists competitively inhibited specific [3H](+)PN 200-110 binding, but d-cis diltiazem enhanced and verapamil incompletely inhibited [3H](+)PN 200-110 binding in both the cerebral cortex and the heart. The effects of diltiazem and verapamil on [3H](+)PN 200-110 binding were due mainly to alterations in the dissociation constant (Kd), without alterations in the binding density (Bmax). The new [3H](+)PN 200-110 receptor binding assay is remarkable for its low degree of nonspecific binding as compared to [3H]nitrendipine at physiological temperatures. [3H](+)PN 200-110 is a useful ligand for the further analysis of the dihydropyridine binding sites associated with calcium channels.