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The highly efficacious actions of N-desmethylclozapine at muscarinic receptors are unique and not a common property of either typical or atypical antipsychotic drugs: is M1 agonism a pre-requisite for mimicking clozapine’s actions?



Recent studies have suggested that the salutary actions of clozapine in schizophrenia may be due to selective activation of M1 muscarinic receptors by clozapine and/or its major active metabolite N-desmethylclozapine.


We systematically tested this hypothesis by screening a large number of psychoactive compounds, including many atypical antipsychotic drugs, for agonist activity at cloned, human M1, M3 and M5 muscarinic receptors.


Only three of the 14 atypical antipsychotic drugs we tested were found to possess partial agonist actions at M1 muscarinic receptors (fluperlapine, JL13, clozapine). A few additional miscellaneous compounds had a modest degree of M1 agonist actions. Only carbachol and N-desmethylclozapine had appreciable M3 muscarinic agonism at M3 muscarinic receptors, although several were M5 partial agonists including MK-212, N-desmethylclozapine and xanomeline.


Although M1 muscarinic receptor-selective partial agonists have shown promise in some preclinical antipsychotic drug models, these studies indicate that it is unlikely that the salutary actions of clozapine and similar atypical antipsychotic drugs are mediated solely by M1 muscarinic receptor activation. It is possible, however, that the M1 agonism of N-desmethylclozapine contributes to the uniquely beneficial actions of clozapine. Thus, these results are consistent with the notion that a balanced degree of activity at multiple biogenic amine receptors, including M1 muscarinic agonism, is responsible for the uniquely beneficial actions of clozapine.

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  1. Altar CA, Wasley AM, Neale RF, Stone GA (1986) Typical and atypical antipsychotic occupancy of D2 and S2 receptors: an autoradiographic analysis in rat brain. Brain Res Bull 16:517–525

  2. Ancellin N, Preisser L, Le Maout S, Barbado M, Creminon C, Corman B, Morel A (1999) Homologous and heterologous phosphorylation of the vasopressin V1a receptor. Cell Signal 11:743–751

  3. Bolden C, Cusack B, Richelson E (1992) Antagonism by antimuscarinic and neuroleptic compounds at the five cloned human muscarinic cholinergic receptors expressed in Chinese hamster ovary cells. J Pharmacol Exp Ther 260:576–580

  4. Bymaster FP, Calligaro DO, Falcone JF, Marsh RD, Moore NA, Tye NC, Seeman P, Wong DT (1996) Radioreceptor binding profile of the atypical antipsychotic olanzapine. Neuropsychopharmacology 14:87–96

  5. Bymaster FP, Shannon HE, Rasmussen K, Delapp NW, Mitch CH, Ward JS, Calligaro DO, Ludvigsen TS, Sheardown MJ, Olesen PH, Swedberg MD, Sauerberg P, Fink-Jensen A (1998) Unexpected antipsychotic-like activity with the muscarinic receptor ligand (5R,6R)6-(3-propylthio-1,2,5-thiadiazol-4-yl)-1-azabicyclo[3.2.1]octane. Eur J Pharmacol 356:109–119

  6. Bymaster FP, Felder C, Ahmed S, McKinzie D (2002) Muscarinic receptors as a target for drugs treating schizophrenia. Curr Drug Target CNS Neurol Disord 1:163–181

  7. Chavkin C, Sud S, Jin W, Steward J, Zjawiony JK, Siebert D, Toth BA, Hufeisen SJ, Roth BL (2004) Salvinorin A, an active component of the hallucinogenic sage salvia divinorum, is a highly efficacious kappa opioid receptor agonist: structural and functional considerations. J Pharmacol Exp Ther 368:1197–2003

  8. Crook JM, Tomaskovic-Crook E, Copolov DL, Dean B (2001) Low muscarinic receptor binding in prefrontal cortex from subjects with schizophrenia: a study of Brodmann’s areas 8, 9, 10, and 46 and the effects of neuroleptic drug treatment. Am J Psychiatry 158:918–925

  9. de Paulis T (2001) M-100907 (Aventis). Curr Opin Invest Drugs 2:123–132

  10. Dean B, McLeod M, Keriakous D, McKenzie J, Scarr E (2002) Decreased muscarinic1 receptors in the dorsolateral prefrontal cortex of subjects with schizophrenia. Mol Psychiatry 7:1083–1091

  11. Dean B, Bymaster FP, Scarr E (2003) Muscarinic receptors in schizophrenia. Curr Mol Med 3:419–426

  12. Gewirtz GR, Gorman JM, Volavka J, Macaluso J, Gribkoff G, Taylor DP, Borison R (1994) BMY14802, a sigma receptor ligand for the treatment of schizophrenia. Neuropsychopharmacology 10:37–40

  13. Harrison PJ, Owen MJ (2003) Genes for schizophrenia? Recent findings and their pathophysiological implications. Lancet 361:417–419

  14. Hodder P, Mull R, Cassaday J, Berry K, Strulovici B (2004) Miniaturization of intracellular calcium assays to 1536-well plate format using a fluorometric imaging plate reader. J Biomol Screen 9:417–426

  15. Janssen PA, Niemegeers CJ, Awouters F, Schellenkens KH, Megens AA, Meert TF (1988) Pharmacology of risperidone (R 64,766) a new antipsychotic with serotonin-S2 and dopamine-D2 antagonistic properties. J Pharmacol Exp Ther 244:685–693

  16. Kahn RS, Siever L, Davidson M, Greenwald C, Moore C (1993) Haloperidol and clozapine treatment and their effect on M-chlorophenylpiperazine-mediated responses in schizophrenia: implications for the mechanism of action of clozapine. Psychopharmacology 112:S90–S94

  17. Kane J, Honigfield G, Singer J, Meltzer HY, Group at CCS (1988) Clozapine for the treatment-resistant schizophrenic. Arch Gen Psychiatry 45:789–796

  18. Karle J, Clemmesen L, Hansen L, Andersen M, Andersen J, Fensbo C, Sloth-Nielsen M, Skrumsager BK, Lublin H, Gerlach J (1995) NNC 01-0687, a selective dopamine D1 receptor antagonist, in the treatment of schizophrenia. Psychopharmacology 121:328–329

  19. Kassack MU, Hofgen B, Lehmann J, Eckstein J, Quillan JM, Sadee W (2002) Functional screening of G protein coupled receptors by measuring intracellular calcium with a fluorescence microplate reader. J Biomol Screen 7:233–246

  20. Kenakin T (1997) Differences between natural and recombinant G protein-coupled receptor systems with varying receptor/G protein stoichiometry. Trends Pharmacol Sci 18:456–464

  21. Kohen R, Metcalf MA, Khan N, Druck T, Huebner K, Lachowicz JE, Meltzer HY, Sibley DR, Roth BL, Hamblin MW (1996) Cloning, characterization, and chromosomal localization of a human 5-HT6 serotonin receptor. J Neurochem 66:47–56

  22. Kramer MS, Last B, Getson A, Reines SA (1997) The effects of a selective D4 dopamine receptor antagonist (L-745,870) in acutely psychotic inpatients with schizophrenia. D4 Dopamine Antagonist Group. Arch Gen Psychiatry 54:567

  23. Kroeze WK, Sheffler DJ, Roth BL (2003) G-protein-coupled receptors at a glance. J Cell Sci 116:4867–4869

  24. Kuoppamaki M, Syvalahti E, Hietala J (1993) Clozapine and N-desmethylclozapine are potent 5-HT1C receptor antagonists. Eur J Pharmacol 245:179–182

  25. Lahti AC, Weiler M, Carlsson A, Tamminga CA (1998) Effects of the D3 and autoreceptor-preferring dopamine antagonist (+)-UH232 in schizophrenia. J Neural Transm 105:719–734

  26. Leucht S, Pitschel-Walz G, Engel RR, Kissling W (2002) Amisulpride, an unusual “atypical” antipsychotic: a meta-analysis of randomized controlled trials. Am J Psychiatry 159:180–190

  27. Leucht S, Wahlbeck K, Hamann J, Kissling W (2003) New generation antipsychotics versus low-potency conventional antipsychotics: a systematic review and meta-analysis. Lancet 361:1581–1589

  28. Liao DL, Hong CJ, Chen HM, Chen YE, Lee SM, Chang CY, Chen H, Tsai SJ (2003) Association of muscarinic m1 receptor genetic polymorphisms with psychiatric symptoms and cognitive function in schizophrenic patients. Neuropsychobiology 48:72–76

  29. Meltzer HY, McGurk SR (1999) The effects of clozapine, risperidone, and olanzapine on cognitive function in schizophrenia. Schizophr Bull 25:233–255

  30. Meltzer HY, Matsubara S, Lee J-C (1989) Classification of typical and atypical antipsychotic drugs on the basis of dopamine D-1, D-2 and serotonin2 pKi values. J Pharmacol Exp Ther 251:238–246

  31. Meltzer HY, Alphs L, Green AI, Altamura AC, Anand R, Bertoldi A, Bourgeois M, Chouinard G, Islam MZ, Kane J, Krishnan R, Lindenmayer JP, Potkin S (2003) Clozapine treatment for suicidality in schizophrenia: international suicide prevention trial (InterSePT). Arch Gen Psychiatry 60:82–91

  32. Meltzer H, Arvantis L, Bauer D, Rein W, Group MS (2004) A placebo-controlled evaluation of four novel compounds for the treatment of schizophrenia and schizoaffective disorder. Arch Gen Psychiatry (in press)

  33. Miller RJ, Hiley CR (1974) Anti-muscarinic properties of neuroleptics and drug-induced parkinsonism. Nature (London) 248:546–547

  34. Muller MJ, Grunder G, Wetzel H, Muller-Siecheneder F, Marx-Dannigkeit P, Benkert O (1999) Antipsychotic effects and tolerability of the sigma ligand EMD 57445 (panamesine) and its metabolites in acute schizophrenia: an open clinical trial. Psychiatry Res 89:275–280

  35. Newcomer JW, Faustman WO, Zipursky RB, Csernansky JG (1992) Zacopride in schizophrenia: a single-blind serotonin type 3 antagonist trial. Arch Gen Psychiatry 49:751–752

  36. Newman-Tancredi A, Gavaudan S, Conte C, Chaput C, Touzard M, Verriele L, Audinot V, Millan MJ (1998) Agonist and antagonist actions of antipsychotic agents at 5-HT1A receptors: a [35S]GTPgammaS binding study. Eur J Pharmacol 355:245–256

  37. Olianas MC, Maullu C, Onali P (1999) Mixed agonist-antagonist properties of clozapine at different human cloned muscarinic receptor subtypes expressed in Chinese hamster ovary cells. Neuropsychopharmacology 20:263–270

  38. Pfeiffer CC, Jenney EH (1957) The inhibition of the conditioned response and the counteraction of schizophrenia by muscarinic stimulation of the brain. Ann N Y Acad Sci 66:753–764

  39. Potkin SG, Shipley J, Bera R, Carreon D, Fallon J, Alva G, Keator D (2001) Clinical and PET Effects of M100907, a selective 5HT-2A receptor antagonist. Schizophr Res 49:242

  40. Poyurovsky M, Weizman A (1999) Lack of efficacy of the 5-HT3 receptor antagonist granisetron in the treatment of acute neuroleptic-induced akathisia. Int Clin Psychopharmacol 14:357–360

  41. Raedler TJ, Knable MB, Jones DW, Urbina RA, Egan MF, Weinberger DR (2003a) Central muscarinic acetylcholine receptor availability in patients treated with clozapine. Neuropsychopharmacology 28:1531–1537

  42. Raedler TJ, Knable MB, Jones DW, Urbina RA, Gorey JG, Lee KS, Egan MF, Coppola R, Weinberger DR (2003b) In vivo determination of muscarinic acetylcholine receptor availability in schizophrenia. Am J Psychiatry 160:118–127

  43. Rasmussen T, Fink-Jensen A, Sauerberg P, Swedberg MD, Thomsen C, Sheardown MJ, Jeppesen L, Calligaro DO, DeLapp NW, Whitesitt C, Ward JS, Shannon HE, Bymaster FP (2001) The muscarinic receptor agonist BuTAC, a novel potential antipsychotic, does not impair learning and memory in mouse passive avoidance. Schizophr Res 49:193–201

  44. Rauser L, Savage JE, Meltzer HY, Roth BL (2001) Inverse agonist actions of typical and atypical antipsychotic drugs at the human 5-hydroxytryptamine(2C) receptor. J Pharmacol Exp Ther 299:83–89

  45. Remington G, Kapur S (2001) SB-277011 GlaxoSmithKline. Curr Opin Invest Drugs 2:946–949

  46. Roth BL, Ciaranello RD, Meltzer HY (1992) Binding of typical and atypical antipsychotic agents to transiently expressed 5-HT1C receptors. J Pharmacol Exp Ther 260:1361–1365

  47. Roth BL, Craigo SC, Choudhary MS, Uluer A, Monsma FJ Jr, Shen Y, Meltzer HY, Sibley DR (1994) Binding of typical and atypical antipsychotic agents to 5-hydroxytryptamine-6 and 5-hydroxytryptamine-7 receptors. J Pharmacol Exp Ther 268:1403–1410

  48. Roth BL, Tandra S, Burgess LH, Sibley DR, Meltzer HY (1995) D4 dopamine receptor binding affinity does not distinguish between typical and atypical antipsychotic drugs. Psychopharmacology 120:365–368

  49. Roth BL, Baner K, Westkaemper R, Siebert D, Rice KC, Steinberg S, Ernsberger P, Rothman RB (2002) Salvinorin A: a potent naturally occurring nonnitrogenous kappa opioid selective agonist. Proc Natl Acad Sci USA 99:11934–11939

  50. Roth BL, Sheffler DJ, Potkin S (2003) Atypical antipsychotic drugs: unitary or multiple mechanisms for “atypicality”. Clin Neurosci Res 3:108–117

  51. Roth BL, Kroeze WK, Sheffler DJ (2004) Magic bullets vs magic shotguns: why selectively non-selective drugs are better for treating schizophrenia and depression. Nat Rev Drug Discov 3:359–364

  52. Rothman RB, Vu N, Partilla JS, Roth BL, Hufeisen SJ, Compton-Toth BA, Birkes J, Young R, Glennon RA (2003) In vitro characterization of ephedrine-related stereoisomers at biogenic amine transporters and the receptorome reveals selective actions as norepinephrine transporter substrates. J Pharmacol Exp Ther 307:138–145

  53. Schoemaker H, Claustre Y, Fage D, Rouquier L, Chergui K, Curet O, Oblin A, Gonon F, Carter C, Benavides J, Scatton B (1997) Neurochemical characteristics of amisulpride, an atypical dopamine D2/D3 receptor antagonist with both presynaptic and limbic selectivity. J Pharmacol Exp Ther 280:83–97

  54. Seeger TF, Seymour PA, Schmidt AW, Zorn SH, Schulz DW, Lebel LA, McLean S, Guanowsky V, Howard HR, Lowe JA III et al. (1995) Ziprasidone (CP-88,059): a new antipsychotic with combined dopamine and serotonin receptor antagonist activity. J Pharmacol Exp Ther 275:101–113

  55. Seibyl JP, Krystal JH, Pricer LH, Woods SW, Heninger GR, Charney DS (1989) 5-HT function in the biochemical and behavioral effects of MCPP in healthy subjects and schizophrenics. Soc Neurosci Abstr 15:1236

  56. Setola V, Hufeisen SJ, Grande-Allen KJ, Vesely I, Glennon RA, Blough B, Rothman RB, Roth BL (2003) 3,4-Methylenedioxymethamphetamine (MDMA, “Ecstasy”) induces fenfluramine-like proliferative actions on human cardiac valvular interstitial cells in vitro. Mol Pharmacol 63:1223–1229

  57. Shannon HE, Rasmussen K, Bymaster FP, Hart JC, Peters SC, Swedberg MD, Jeppesen L, Sheardown MJ, Sauerberg P, Fink-Jensen A (2000) Xanomeline, an M(1)/M(4) preferring muscarinic cholinergic receptor agonist, produces antipsychotic-like activity in rats and mice. Schizophr Res 42:249–259

  58. Shapiro DA, Renock S, Arrington E, Chiodo LA, Liu LX, Sibley DR, Roth BL, Mailman R (2003) Aripiprazole, a novel atypical antipsychotic drug with a unique and robust pharmacology. Neuropsychopharmacology 28:1400–1411

  59. Snyder S, Greenberg D, Yamamura HI (1974) Antischizophrenic drugs and brain cholinergic receptors. Affinity for muscarinic sites predicts extrapyramidal effects. Arch Gen Psychiatry 31:58–61

  60. Sur C, Mallorga PJ, Wittmann M, Jacobson MA, Pascarella D, Williams JB, Brandish PE, Pettibone DJ, Scolnick EM, Conn PJ (2003) N-desmethylclozapine, an allosteric agonist at muscarinic 1 receptor, potentiates N-methyl-d-aspartate receptor activity. Proc Natl Acad Sci USA 100:13674–13679

  61. Tandon R (1999) Cholinergic aspects of schizophrenia. Br J Psychiatry Suppl 37:7–11

  62. Tandon R, Greden JF (1987) Trihexyphenidyl treatment of negative schizophrenic symptoms. Acta Psychiatr Scand 76:732

  63. Tandon R, Greden JF (1989) Cholinergic hyperactivity and negative schizophrenic symptoms. A model of cholinergic/dopaminergic interactions in schizophrenia. Arch Gen Psychiatry 46:745–753

  64. Tuunainen A, Wahlbeck K, Gilbody S (2002) Newer atypical antipsychotic medication in comparison to clozapine: a systematic review of randomized trials. Schizophr Res 56:1–10

  65. Van Tol HH, Bunzow JR, Guan HC, Sunahara RK, Seeman P, Niznik HB, Civelli O (1991) Cloning of the gene for a human dopamine D4 receptor with high affinity for the antipsychotic clozapine. Nature 350:610–614

  66. Watson J, Brough S, Coldwell MC, Gager T, Ho M, Hunter AJ, Jerman J, Middlemiss DN, Riley GJ, Brown AM (1998) Functional effects of the muscarinic receptor agonist, xanomeline, at 5-HT1 and 5-HT2 receptors. Br J Pharmacol 125:1413–1420

  67. Wetzel H, Szegedi A, Hain C, Wiesner J, Schlegel S, Benkert O (1995) Seroquel (ICI 204 636), a putative “atypical” antipsychotic, in schizophrenia with positive symptomatology: results of an open clinical trial and changes of neuroendocrinological and EEG parameters. Psychopharmacology 119:231–238

  68. Zeng XP, Le F, Richelson E (1997) Muscarinic m4 receptor activation by some atypical antipsychotic drugs. Eur J Pharmacol 321:349–354

  69. Zorn SH, Jones SB, Ward KM, Liston DR (1994) Clozapine is a potent and selective muscarinic M4 receptor agonist. Eur J Pharmacol 269:R1–R2

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This work was supported in part by RO1MH57635, KO2MH01366 and the NIMH Psychoactive Drug Screening Program to B.L.R. M.A.D. was supported in part by a supplement to RO1MH57635.

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Correspondence to Bryan L. Roth.

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Davies, M.A., Compton-Toth, B.A., Hufeisen, S.J. et al. The highly efficacious actions of N-desmethylclozapine at muscarinic receptors are unique and not a common property of either typical or atypical antipsychotic drugs: is M1 agonism a pre-requisite for mimicking clozapine’s actions?. Psychopharmacology 178, 451–460 (2005). https://doi.org/10.1007/s00213-004-2017-1

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  • Clozapine
  • Atypical antipsychotic
  • N-desmethylclozapine
  • Muscarinic