Modulation of 5-Hydroxytryptamine1AReceptor Density by Nonhydrolyzable GTP Analogues

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Abstract
Co-incubation of rat cortical membranes with 10-4 M GTP results in a competitive inhibition of 5-hydroxytryptamine1A (5-HT1A) receptor binding sites labeled by [3H]8-hydroxy-2-(di-n-propylamino)tetralin ([3H]8-OH-DPAT). Preincubation of cortical membranes with 10-4 M GTP does not significantly change either KD or Bmax values, indicating that the effect of GTP is reversible. By contrast, GTP.gamma.S and 5''-guanylylimidodiphosphate (GppNHp) are nonhydrolyzable analogues of GTP which lengthen the time course of guanine nucleotide activation of guanine nucleotide binding proteins (G proteins) and thereby alter G protein-receptor interactions. These nonhydrolyzable GTP analogues were used to characterize the effects of persistent alterations in G proteins on [3H]8-OH-DPAT binding to 5-HT1A receptors. Co-incubation of rat cortical membranes with either 10-4 M GTP.gamma.S or GppNHp results in a decrease in both the affinity and apparent density of 5-HT1A binding sites. Co-incubation with the nonhydrolyzable nucleotides reduces the affinity of [3H]8-OH-DPAT binding by 65-70% and lowers the density of the binding site by 53-61%. Similarly, preincubation of membranes with a 10-4 M concentration of either GTP.gamma.S or GppNHp significantly increases the KD value and reduces the Bmax value of [3H]8--OH-DPAT binding. These results indicate that GTP.gamma.S and GppNHp induce persistent changes in 5-HT1A receptor-G protein interactions that are reflected as a decrease in the density of binding sites labeled by [3H]8-OH-DPAT.