Clozapine and other competitive antagonists reactivate risperidone-inactivated h5-HT7 receptors: radioligand binding and functional evidence for GPCR homodimer protomer interactions

• Published in: Psychopharmacologia Vol. 212; no. 4; pp. 687 - 697
• Main Authors: Teitler, Milt, Toohey, Nicole, Knight, Jessica A., Klein, Michael T., Smith, Carol
• Format: Journal Article
• Language: English
• Published: Berlin/Heidelberg Springer-Verlag 01.12.2010; Springer; Springer Nature B.V
• Subjects: Antagonist drugs; Binding, Competitive; Biological and medical sciences; Biomedical and Life Sciences; Biomedicine; Biosensing Techniques; Cell Line; Clozapine - metabolism; Clozapine - pharmacology; Cyclic AMP - metabolism; Dose-Response Relationship, Drug; Humans; Isoxazoles - pharmacology; Medical sciences; Neuropharmacology; Neurosciences; Original Investigation; Paliperidone Palmitate; Pharmacology. Drug treatments; Pharmacology/Toxicology; Protein Binding; Protein Multimerization; Protein Subunits; Psychiatry; Psycholeptics: tranquillizer, neuroleptic; Psychology. Psychoanalysis. Psychiatry; Psychopharmacology; Psychotropic drugs; Pyrimidines - pharmacology; Radioligand Assay; Receptors, Serotonin - chemistry; Receptors, Serotonin - drug effects; Receptors, Serotonin - genetics; Receptors, Serotonin - metabolism; Risperidone - metabolism; Risperidone - pharmacology; Serotonin; Serotonin Antagonists - metabolism; Serotonin Antagonists - pharmacology; Transfection; Reactivation; Serotonin; 9-OH-risperidone; Serotonin receptor; Protomer–protomer interactions; Homodimer; GPCR; Clozapine; 5-HT; Inactivation; Risperidone; Dibenzodiazepine derivatives; Atypical antipsychotic; Neuroleptic; Psychotropic; Serotonine receptor; Protomer-protomer interactions; G protein coupled receptor; Benzisoxazole derivatives; Biological receptor; Dopamine antagonist; Serotonin antagonist; Interaction; Biological fixation; D2 Dopamine receptor; Neurotransmitter
• ISSN: 0033-3158; 1432-2072; 1432-2072
• DOI: 10.1007/s00213-010-2001-x

Rationale The h5-HT 7 receptor is subject to inactivation by risperidone and 9-OH-risperidone, apparently through a pseudo-irreversible complex formed between these drugs and the receptor. Although risperidone and 9-OH-risperidone (“inactivating antagonists”) completely inactivate the receptor, only 50% of the receptors form a pseudo-irreversible complex with these drugs. Objectives This study aims to more fully determine the mechanism(s) responsible for the novel effects of risperidone and 9-OH-risperidone and to determine if the inactivation can be reversed (reactivation). Methods The ability of non-inactivating drugs (competitive antagonists) to dissociate wash-resistant [ 3 H]risperidone binding from h5-HT 7 receptors was investigated. Also, the ability of non-inactivating drugs to reactivate inactivated h5-HT 7 receptors was investigated, using cAMP accumulation as a functional endpoint. Results The competitive (non-inactivating) antagonists clozapine and mesulergine released the wash-resistant [ 3 H]risperidone binding to the h5-HT 7 receptor. The competitive antagonists clozapine, SB269970, mianserin, cyproheptadine, mesulergine, and ICI169369 reactivated the risperidone-inactivated h5-HT 7 receptors in a concentration-dependent manner. The potencies for reactivation closely match the affinities of these drugs for the h5-HT 7 receptor ( r 2  = 0.95), indicating that the reactivating antagonists are binding to and producing their effects through the orthosteric binding site of the h5-HT 7 receptor. Bioluminescence resonance energy transfer analyses indicate that the h5-HT 7 receptor forms homodimers. Conclusions The ability of the non-inactivating drugs to bind h5-HT 7 orthosteric sites and reverse the wash-resistant effects of risperidone or 9-OH-risperidone, also bound to h5-HT 7 orthosteric sites, is evidence for protomer–protomer interactions between h5-HT 7 homodimers. This is the first demonstration of a non-mutated G-protein-coupled receptor homodimer engaging in protomer–protomer interactions in an intact cell preparation.