Project 3: Elucidation of Biochemical and Behavioral Efficacies of Antipsychotics

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SPECIFIC AIMS In classical pharmacology, an agonist activates a single linear signal transduction pathway, whereas an antagonist blocks the action of the agonist and possesses no intrinsic activity. Over the past few years, it has become clear that signal transduction pathways are not merely linear cascades for signaling. Instead, they are organized into complex signaling networks that require high levels of regulation to generate precise and unique cellular responses. Hence, a given receptor, through various ligand-induced functional conformations, can engage multiple modalities through interaction with different signaling partners. In this way, a given ligand can bind a receptor and act as an antagonist for one signaling pathway while serving as an agonist at another or the converse. This property is established for several G protein-coupled receptors (GPCRs) - the most important targets for therapeutic intervention. Importantly, none of the drugs in clinical use have been developed with these multiple signaling considerations in mind. Additionally, agonists and antagonists are rarely completely selective and, for a given receptor, may alter signaling by influencing various receptor-mediated processes such as interaction with G proteins, desensitization, internalization, down-regulation, and receptor-mediated scaffolding of non-G protein signaling components. Unfortunately, the physiological relevance of these properties is not fully appreciated. Thus, identifying the functional selectivity of compounds may help reveal not only distinct biological processes, but also specific functional outcomes. Currently, the relevance of functional selectivity to psychiatry is unknown. This is particularly important for antipsychotic drugs, where dopamine (DA) D2 receptor (D2R) antagonism is essentially a prerequisite for all these drugs; however, their other intrinsic activities are obscure. The overall goal of the proposed research is to examine behavioral responses to antipsychotic compounds and to elucidate signal transduction mechanisms that are essential for antipsychotic efficacy in preclinical genetic and pharmacological mouse models of schizophrenia-like behaviors. For behavior, effects of antipsychotic compounds on motor activity, prepulse inhibition (PPI), latent inhibition (LI), and social behavior in DA transporter (DAT) knockout (KO), N-methyl-D-aspartate (NMDA) receptor NR1-subunit knockdown (KD), and C57BL/6 mice treated with amphetamine (AMPH) or phencyclidine (PCP) to reproduce schizophrenia-like states. Molecular fingerprinting of signal transduction pathways (MFSTP) will be performed to analyze effects of antipsychotic compounds on various signal transduction modalities that include the protein kinase A (PKA) and DA and cAMP-regulated phosphoprotein 32 (DARPP- 32), Akt/protein kinase B (PKB or Akt) and glycogen synthase kinase 33 (GSK3), phospholipase C (PLC), and extracellular signal-regulated mitogen activated protein kinase (ERK) pathways. The experiments in the present Project #3 will complement those in Projects #1 and #2 by providing preclinical models to test the in vivo selectivities and efficacies of various antipsychotic compounds on amelioration of schizophrenia-like behaviors and will correlate these responses to alterations in signal transduction. Our Project #3 will complement also the Core Project from Wyeth where antipsychotic responses will be analyzed in rat models of schizophrenia-like and antipsychotic-treated behaviors. Understanding the relevance of functional selectivity of antipsychotic drugs may provide novel targets with fewer side-effects, greater therapeutic selectivity, and enhanced efficacy for treating individuals with schizophrenia. Our Project #3 has three Specific Aims. Aim I. Role of hyperdopaminergia in responses to antipsychotic compounds in DAT-KO mice. A. To compare behavioral responses of DAT-KO mice to different antipsychotic compounds. B. To apply MFSTP to various brain regions to define the hyperdopaminergic phenotype and analyze responses to antipsychotic compounds. Aim II. Role of reduced NMD A receptor expression in responses to antipsychotics in NR1-KD mice. A. To compare behavioral responses of NR1 hypomorphic mice to different antipsychotic compounds. B. To apply MFSTP to various brain regions to define the phenotype of the NMDA receptor hypomorphic mice and analyze responses to antipsychotic compounds. Aim III. Roles of pharmacologically-induced hyperdopaminergia or hypoglutamatergia in responses to antipsychotic compounds in C57BU6 mice. A. To compare behavioral responses of mice treated with AMPH or PCP to different antipsychotics. B. To apply MFSTP to various brain regions to define pharmacologically-induced hyperdopaminergic or hypoglutamatergic states and analyze responses to antipsychotic compounds.