Lack of apparent receptor reserve at postsynaptic 5-hydroxytryptamine1A receptors negatively coupled to adenylyl cyclase activity in rat hippocampal membranes
Yocca, F.D.; Iben, L.; Meller, E.
Molecular Pharmacology 41(6): 1066-1072
1992
ISSN/ISBN: 0026-895X PMID: 1352034 Document Number: 396153
Previous studies have demonstrated the existence of a large receptor reserve for agonists at somatodendritic 5hydroxytryptamine-1A (5-HT-1A) serotonin receptors in the raphe nuclei of the rat. 5-HT-1A agonists with anxiolytic properties (e.g., buspirone, gepirone, and ipsapirone) display full intrinsic activity at these receptors but are partial agonists at postsynaptic 5HT-1A receptors, which suggests that the latter sites may be devoid of a receptor reserve. In the present studies, this was directly determined by examining the relationship between receptor occupancy and response at postsynaptic 5-HT-1A receptors, in rat hippocampus, mediating the inhibition of forskolin-stimulated adenylyl cyclase activity, using the method of partial irreversible receptor inactivation. Rats were treated with vehicle or the irreversible antagonist N-ethoxycarbonyl-2-ethoxy-1,2-dihydroquinoline (EEDQ), and 24 hr later hippocampi were removed for saturation analysis of (3H)8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT) binding to 5-HT-1A receptors or for adenylyl cyclase assays. EEDQ (1 and 6 mg/kg) dose-dependently reduced the maximal number of (3H)8-OH-DPAT binding sites by 68.5 and 80%, respectively, without altering the K-d. Concentration-response curves were generated for inhibition of forskolin-stimulated adenylyl cyclase activity by 5-HT and the selective 5HT-1A agonist N,N-dipropyl-5-carboxamidotryptamine (DP-5-CT). EEDQ treatment dose-dependently reduced the maximal inhibitory effect of 5-HT (percentage of inhibition: control, 23.6; EEDQ (1 mg/kg), 13.4; EEDQ (6 mg/kg), 8.9), without altering either the slope factor (1.01) or the EC-50 (96.4 nm). Analogous results were obtained with DP-5-CT (percentage of maximal inhibition: control, 24.1; EEDQ (1 mg/kg), 15.2; EEDQ (6 mg/kg), 10.7), again without changes in slope factor (0.89) or EC-50 (9.9 nm). Analysis of double-reciprocal plots of equieffective concentrations of agonist, followed by calculation of fractional receptor occupancy, revealed a linear relationship between receptor occupancy and response for both 5-HT and DP-5-CT (i.e., an absence of receptor reserve). The receptor specificity of the effect of EEDQ was demonstrated in two ways. First, it was shown that pretreatment of rats with the selective 5-HT-1A partial agonist BMY 7378 (10 mg/kg) before EEDQ afforded substantial protection (about 75%) against loss of the inhibitory effect of DP-5-CT on forskolin-stimulated adenylyl cyclase activity. Second, EEDQ did not alter the inhibition of forskolin-stimulated adenylyl cyclase activity induced by the adenosine Al receptor agonist phenylisopropyladenosine (PIA). Because the inhibition of forskolin-stimulated adenylyl cyclase activity by PIA and 5-HT in the hippocampus appears to be mediated via a common pool of guanine nucleotide-binding regulatory protein(s) interacting with the same effector, EEDQ likely elicits its effects by inactivation of the 5-HT-1A receptor active site and not by interference with intracellular proteins essential for signal transduction. The results support the hypothesis that the differential potency and efficacy of 5-HT-1A agonists at pre- and postsynaptic 5-HT-1A receptors reflects variation in the extent of receptor reserve at the two loci, rather than actions at different receptors.