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We found that 3-cyano-N-(1,3-diphenyl-1H-pyrazol-5-yl)benzamide (CDPPB) is a potent and selective positive allosteric modulator of the metabotropic glutamate receptor subtype 5 (mGluR5). In Chinese hamster ovary cells expressing human mGluR5, CDPPB potentiated threshold responses to glutamate in fluorometric Ca 2ϩ assays more than 7-fold with an EC 50 value of approximately 27 nM. At 1 M, CDPPB shifted mGluR5 agonist concentration response curves to glutamate, quisqualate, and (R,S)-3,5-dihydroxyphenylglycine 3-to 9-fold to the left. At higher concentrations, CDPPB exhibited agonist-like activity on cells expressing mGluR5. No other activity was observed on any other mGluR or cell type at concentrations up to 10 M. CDPPB had no effect on [ 3 H]quisqualate binding to mGluR5 but did compete for binding of [ 3 H]methoxyPEPy, an analog of the selective mGluR5 negative allosteric modulator MPEP. CDPPB was found to be brain penetrant and reversed amphetamine-induced locomotor activity and amphetamineinduced deficits in prepulse inhibition in rats, two models sensitive to antipsychotic drug treatment. These results demonstrate that positive allosteric modulation of mGluR5 produces behavioral effects, suggesting that such modulation serves as a viable approach to increasing mGluR5 activity in vivo. These effects are consistent with the hypothesis that allosteric potentiation of mGluR5 may provide a novel approach for development of antipsychotic agents.Glutamate, the predominant excitatory neurotransmitter in the mammalian central nervous system (CNS), exerts its effects through two classes of receptors. The first class of receptor, ionotropic glutamate receptors, are postsynaptic, multimeric ligand-gated ion channels classified into three groups named for group-selective agonists: N-methyl-D-aspartate (NMDA), ␣-amino-3-hydroxy-5-methyl-4-isoxazopropionic acid, and kainate receptors. The NMDA receptor (NMDAR) is known to play an important role in processes related to schizophrenia. NMDAR antagonists, such as phencyclidine and ketamine, induce positive, negative, and cognitive symptoms reminiscent of schizophrenia in human volunteers and worsen existing symptoms in schizophrenic patients. This observation has led to the hypothesis that changes in CNS circuits induced by NMDAR hypofunction may play a key role in the development and/or in the underlying symptoms of schizophrenia (Olney et al., 1999). Therefore, development of compounds that selectively increase NMDAR function could be used to test this hypothesis.The metabotropic glutamate receptors (mGluRs), the second class of glutamate receptor, are members of family C of the G-protein-coupled receptors and are characterized by a large extracellular agonist binding domain on the aminoterminal end of the receptor that is distinct from the seventransmembrane domain characteristic of all G-protein-couArticle, publication date, and citation information can be found at

The molecular and neuronal substrates conferring on clozapine its unique and superior efficacy in the treatment of schizophrenia remain elusive. The interaction of clozapine with many G proteincoupled receptors is well documented but less is known about its biologically active metabolite, N-desmethylclozapine. Recent clinical and preclinical evidences of the antipsychotic activity of the muscarinic agonist xanomeline prompted us to investigate the effects of N-desmethylclozapine on cloned human M1-M5 muscarinic receptors. N-desmethylclozapine preferentially bound to M1 muscarinic receptors with an IC 50 of 55 nM and was a more potent partial agonist (EC50, 115 nM and 50% of acetylcholine response) at this receptor than clozapine. Furthermore, pharmacological and site-directed mutagenesis studies suggested that N-desmethylclozapine preferentially activated M1 receptors by interacting with a site that does not fully overlap with the acetylcholine orthosteric site. As hypofunction of N-methyl-D-aspartate (NMDA) receptordriven neuronal ensembles has been implicated in psychotic disorders, the neuronal activity of N-desmethylclozapine was electrophysiologically investigated in hippocampal rat brain slices. N-desmethylclozapine was shown to dose-dependently potentiate NMDA receptor currents in CA1 pyramidal cells by 53% at 100 nM, an effect largely mediated by activation of muscarinic receptors. Altogether, our observations provide direct evidence that the brain penetrant metabolite N-desmethylclozapine is a potent, allosteric agonist at human M1 receptors and is able to potentiate hippocampal NMDA receptor currents through M1 receptor activation. These observations raise the possibility that N-desmethylclozapine contributes to clozapine's clinical activity in schizophrenics through modulation of both muscarinic and glutamatergic neurotransmission.

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