Clozapine reverses phencyclidine-induced desynchronization of prefrontal cortex through a 5-HT 1A receptor-dependent mechanism

The non-competitive NMDA receptor (NMDA-R) antagonist phencyclidine (PCP) - used as a pharmacological model of schizophrenia - disrupts prefrontal cortex (PFC) activity. PCP markedly increased the discharge rate of pyramidal neurons and reduced slow cortical oscillations (SCO; 0.15-4 Hz) in rat PFC. Both effects were reversed by classical (haloperidol) and atypical (clozapine) antipsychotic drugs. Here we extended these observations to mice brain and examined the potential involvement of 5-HT 2A and 5-HT 1A receptors (5-HT 2AR and 5-HT 1AR, respectively) in the reversal by clozapine of PCP actions. Clozapine shows high in vitro affinity for 5-HT 2AR and behaves as partial agonist in vivo at 5-HT 1AR. We used wild-type (WT) mice and 5-HT 1AR and 5-HT 2AR knockout mice of the same background (C57BL/6) (KO-1A and KO-2A, respectively). Local field potentials (LFPs) were recorded in the PFC of WT, KO-1A, and KO-2A mice. PCP (10 mg/kg, intraperitoneally) reduced SCO equally in WT, KO-2A, and KO-1A mice (58±4%, 42±7%, and 63±7% of pre-drug values, n = 23, 13, 11, respectively; p < 0.0003). Clozapine (0.5 mg/kg, intraperitoneally) significantly reversed PCP effect in WT and KO-2A mice, but not in KO-1A mice nor in WT mice pretreated with the selective 5-HT 1AR antagonist WAY-100635.The PCP-induced disorganization of PFC activity does not appear to depend on serotonergic function. However, the lack of effect of clozapine in KO-1A mice and the prevention by WAY-100635 indicates that its therapeutic action involves 5-HT 1AR activation without the need to block 5-HT 2AR, as observed with clozapine-induced cortical dopamine release. © 2012 American College of Neuropsychopharmacology. All rights reserved.The work leading to these results has received funding from the Innovative Medicines Initiative Joint Undertaking (IMI) under Grant Agreement No. 115008 (NEWMEDS). This work was supported by Instituto de Salud Carlos III, Centro de Investigacion Biomedica en Red de Salud Mental (CIBERSAM) and Grants SAF 2007-62378, FIS PI09/1245 (PN de I + D + I 2008-2011, ISCIII-Subdireccion General de Evaluacion y Fomento de la Investigacion), CIBERSAM (P82, 11INT3), and SENY Fundacio. PC is supported by the Researcher Stabilization Program of the Health Department of the Generalitat de Catalunya. LK was recipient of a predoctoral fellowship from the Ministry of Science and Education.Peer Reviewe

Overstimulation of NMDA Receptors Impairs Early Brain Development in vivo

BACKGROUND: Brains of patients with schizophrenia show both neurodevelopmental and functional deficits that suggest aberrant glutamate neurotransmission. Evidence from both genetic and pharmacological studies suggests that glutamatergic dysfunction, particularly with involvement of NMDARs, plays a critical role in the pathophysiology of schizophrenia. However, how prenatal disturbance of NMDARs leads to schizophrenia-associated developmental defects is largely unknown. METHODOLOGY/PRINCIPAL FINDINGS: Glutamate transporter GLAST/GLT1 double-knockout (DKO) mice carrying the NMDA receptor 1 subunit (NR1)-null mutation were generated. Bouin-fixed and paraffin-embedded embryonic day 16.5 coronal brain sections were stained with hematoxylin, anti-microtubule-associated protein 2 (MAP2), and anti-L1 antibodies to visualize cortical, hippocampal, and olfactory bulb laminar structure, subplate neurons, and axonal projections. NR1 deletion in DKO mice almost completely rescued multiple brain defects including cortical, hippocampal, and olfactory bulb disorganization and defective corticothalamic and thalamocortical axonal projections. CONCLUSIONS/SIGNIFICANCE: Excess glutamatergic signaling in the prenatal stage compromises early brain development via overstimulation of NMDARs

Genetic association analysis of N-methyl-d-aspartate receptor subunit gene GRIN2B and clinical response to clozapine

OBJECTIVE: Approximately 30% of patients with schizophrenia fail to respond to antipsychotic therapy and are classified as having treatment-resistant schizophrenia. Clozapine is the most efficacious drug for treatment-resistant schizophrenia and may deliver superior therapeutic effects partly by modulating glutamate neurotransmission. Response to clozapine is highly variable and may depend on genetic factors as indicated by twin studies. We investigated eight polymorphisms in the N-methyl-d-aspartate glutamate receptor subunit gene GRIN2B with response to clozapine. METHODS: GRIN2B variants were genotyped using standard TaqMan procedures in 175 European patients with schizophrenia deemed resistant or intolerant to treatment. Response was assessed using change in Brief Psychiatric Rating Scale scores following six months of clozapine therapy. Categorical and continuous response was assessed using chi-squared test and analysis of covariance, respectively. RESULTS: No associations were observed between the variants and response to clozapine. A-allele carriers of rs1072388 responded marginally better to clozapine therapy than GG-homozygotes; however, the difference was not statistically significant (p = 0.067, uncorrected). CONCLUSIONS: Our findings do not support a role for these GRIN2B variants in altering response to clozapine in our sample. Investigation of additional glutamate variants in clozapine response is warranted. Copyright © 2016 John Wiley & Sons, Ltd

Papel de la corteza prefrontal en la fisiopatología de la esquizofrenia y en el mecanismo de acción de fármacos antipsicóticos

En la presente tesis hemos caracterizado los cambios de actividad cortical que ocurren en la corteza prefrontal (CPF) de animales de experimentación sometidos al tratamiento con fenciclidina (PCP), antagonista no competitivo de los R-NMDA, usada como modelo experimental de esquizofrenia. Así mismo, hemos estudiado la acción de fármacos antipsicóticos en este modelo, habiéndose encontrado que tanto el haloperidol (antipsicótico clásico) como la clozapina (antipsicótico atípico) normalizan las alteraciones de la actividad de la CPF inducidas por PCP. En concreto, la PCP aumenta marcadamente la frecuencia de descarga de un elevado porcentaje (45%) de neuronas piramidales y disminuye la actividad de una proporción menor (33%) del mismo tipo neuronal, produciendo un aumento global de la actividad de la CPF. Estas modificaciones de la actividad a nivel celular, se acompaña de una pérdida de la capacidad de la corteza de generar una actividad oscilatoria de baja frecuencia, que supondría una pérdida de la calidad del procesamiento de la información a nivel cortical y se traduciría en un deterioro de la comunicación de la CPF con otras áreas corticales y subcorticales, alterando de este modo el funcionamiento de circuitos córtico-límbicos. Estas alteraciones a nivel celular y poblacional suponen posiblemente el substrato neurobiológico de gran parte de la acción psicotomimética de la PCP. Asimismo se ha estudiado la posible reversión de los efectos de la PCP por parte de los antipsicóticos clásicos, antagonistas preferentes de receptores D2, como el haloperidol (HAL) y antipsicóticos atípicos, antagonistas preferentes de receptores 5-HT2A, como la clozapina (CLZ), que carece de los importantes efectos secundarios (motores y cognitivos) de los fármacos clásicos, derivados del bloqueo masivo de receptores D2. A pesar de su distinto perfil receptorial, ambos fármacos son capaces de revertir las disfunciones corticales ocasionadas por PCP. El efecto de HAL es muy probablemente debido a su bloqueo de receptores D2, bien a nivel local en la propia CPF, o a nivel distal, en circuitos que implican un retomo de la información a la CPF a través del tálamo. Además, experimentos en ratones carentes de receptores 5-HT2A y 5-HT1A muestran como CLZ precisa de estos últimos receptores para revertir las alteraciones producidas por PCP. En conjunto, los estudios realizados en esta Tesis doctoral representan un paso más en el entendimiento del mecanismo de acción de la CLZ, poniéndose de manifiesto que la modulación la actividad de la CPF juega un rol importante en la fitopatología de la EZ y el restablecimiento de la actividad cortical un punto fundamental del tratamiento, tanto con fármacos clásicos como con fármacos atípicos.Peer Reviewe

Role of prefrontal cortex in pharmacological models of schizophrenia and antipsychotic action

NMDA receptor (NMDA-R) antagonists are extensively used as schizophrenia models due to their ability to evoke positive and negative symptoms as well as cognitive deficits similar to those of the illness. Likewise, 5-HT2A receptor agonists display hallucinogen actions resembling psychotic symptoms. Overall, these drugs are useful models of schizophrenia for the screening of new antipsychotic drugs. However, the cellular and network elements involved in these actions are poorly known. Data obtained by several groups in recent years indicate that the prefrontal cortex (PFC) and anatomically related areas play a major role in these actions. This paper summarizes data obtained by the authors supporting that a) NMDA-R antagonists (phencyclidine –PCP–, dizocilpine –MK–801–) and 5-HT2A agonists (DOI) alter the function of PFC in a similar fashion, and b) antipsychotic drugs exert their therapeutic action, at least in part, by normalizing hyperactivity states in PFC. While the actions of NMDA-R antagonists may involve blockade of these receptors in PFC and subcortical areas, that of antipsychotic drugs, in particular atypical drugs like clozapine, appear to be mediated essentially by a local action in PFC. These results help to better understand the neurobiological basis of the action of pharmacological models of schizophrenia and the mode of action of antipsychotic drugs.Work supported by grants SAF2007-62378, FIS PI070111 and FIS FIS PI060264.Peer reviewe

Disruption of thalamocortical activity in schizophrenia models: relevance to antipsychotic drug action

Non-competitive NMDA receptor antagonists are widely used as pharmacological models of schizophrenia due to their ability to evoke the symptoms of the illness. Likewise, serotonergic hallucinogens, acting on 5-HT2A receptors, induce perceptual and behavioural alterations possibly related to psychotic symptoms. The neurobiological basis of these alterations is not fully elucidated. Data obtained in recent years revealed that the NMDA receptor antagonist phencyclidine (PCP) and the serotonergic hallucinogen 1-(2,5-dimethoxy-4-iodophenyl-2-aminopropane; DOI) produce a series of common actions in rodent prefrontal cortex (PFC) that may underlie psychotomimetic effects. Hence, both agents markedly disrupt PFC function by altering pyramidal neuron discharge (with an overall increase) and reducing the power of low frequency cortical oscillations (LFCO; < 4 Hz). In parallel, PCP increased c-fos expression in excitatory neurons of various cortical areas, the thalamus and other subcortical structures, such as the amygdala. Electrophysiological studies revealed that PCP altered similarly the function of the centromedial and mediodorsal nuclei of the thalamus, reciprocally connected with PFC, suggesting that its psychotomimetic properties are mediated by an alteration of thalamocortical activity (the effect of DOI was not examined in the thalamus). Interestingly, the observed effects were prevented or reversed by the antipsychotic drugs clozapine and haloperidol, supporting that the disruption of PFC activity is intimately related to the psychotomimetic activity of these agents. Overall, the present experimental model can be successfully used to elucidate the neurobiological basis of schizophrenia symptoms and to examine the potential antipsychotic activity of new drugs in development.Supported by the Innovative Medicines Initiative Joint Undertaking (IMI) under Grant Agreement No. 115008 (NEWMEDS). IMI is a public–private partnership between the European Union and the European Federation of Pharmaceutical Industries and Associations. Support from the following grants is also acknowledged: SAF 2012-35183 (Ministry of Economy and Competitiveness and European Regional Development Fund), PI09/1245 and PI12/00156 (PN de I+D+I 2008-2011, ISCIII-Subdireccion General de Evaluación y Fomento de la Investigación cofinanced by the European Regional Development Fund. ‘Una manera de hacer Europa’) and Centro de Investigación Biomédica en Red de Salud Mental, CIBERSAM (P82, 11INT3). Support from the Generalitat de Catalunya (SGR20093) is also acknowledged. P.C. is supported by the Researcher Stabilization Program of the Health Department of the Generalitat de Catalunya. M.R. is recipient of an IDIBAPS fellowship.Peer reviewe