Potency of 5-hydroxytryptamine(1A) agonists to inhibit adenylyl cyclase activity is a function of affinity for the 'low-affinity' state of [³H]8- hydroxy-N,N-dipropylaminotetralin ([³H]8-OH-DPAT) binding

In this study, the radiolabeled 5-hydroxytryptamine(1A) agonist, [³H]8- hydroxy-N,N-dipropylamino tetralin ([³H]8-OH-DPAT), was shown to have both a high (K(d), 0.7 ± 0.2 nM) and a low (K(d), 17 ± 4 nM) affinity binding component in rat hippocampal homogenate preparations in the absence of guanine nucleotides. The high-affinity binding component was markedly reduced by the elimination of Mg⁺⁺ from the incubation medium and the addition of both the nonhydrolyzable guanine nucleotide guanylylimidodiphosphate (Gpp(NH)p) (100 μM) and 1.0 mM EDTA to the incubation medium. Under these latter conditions, a single binding affinity component was observed with a K(d) of 11 ± 1 nM, a value in good agreement with the value for the low- affinity component measured in the absence of Gpp(NH)p. Further, the B(max) value for the single low-affinity binding component measured in the presence of Gpp(NH)p was essentially equivalent to the total of the two B(max) values found in the absence of Gpp(NH)p. A binding assay was developed using 15 nM [³H]8-OH-DPAT to determine the affinities of serotonergic drugs for the low- affinity component of [³H]8-OH-DPAT binding and these values were compared with their affinities for the high-affinity binding component as well as their potencies in a hippocampal adenylyl cyclase assay. For agonists, the K(i) value for the high-affinity binding component was always less than the low-affinity K(i) value, whereas the antagonist spiperone had similar values for both the high-affinity and low-affinity binding components. Furthermore, when K(i) values of the agonists for the high-affinity [³H]8-OH-DPAT binding component were compared to their EC₅₀ values for inhibition of forskolin- stimulated adenylyl cyclase activity, K(i)/EC₅₀ ratios were 15-100. By contrast, the ratio of the K(i) values for the low-affinity component to the EC₅₀ values were near unity for these drugs. It appears that the potency of agonists to elicit 5-hydroxytryptamine(1A)-mediated responses is quantitatively better related to their affinity for the low-affinity state of [³H]8-OH-DPAT binding than for the high-affinity state.