Conte Center

Vanderbilt / NIMH Silvio O. Conte Center for Neuroscience Research

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Vanderbilt University
465 21st Avenue South
Nashville, TN 37232
For more information,
please contact Denise Malone.

2009 Pilot Grant Recipients

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Paul Gresch, Ph.D.

5-HT2A Receptor Regulation of Presynaptic mGlu2 Receptors

This Conte Pilot Project proposal seeks funds for the development of two new techniques for the study of serotonin regulation of cortical glutamate neurotransmission. There is growing evidence for the interaction of serotonin 2A (5-HT2A) and metabotropic glutamate 2 (mGlu2) receptors that produces unique activation of signal transduction pathways and gene expression patterns. Furthermore, mGlu2 receptors can modulate 5-HT2A receptor-induced behaviors. For example, our laboratory demonstrated that chronic 5-HT2A receptor activation diminished the ability of an mGlu2/3 receptor agonist to suppress 5-HT2A mediated head twitch response. Our hypothesis is that continuous activation of 5-HT2A receptors produces a persistent hyperglutamatergic state that leads to mGlu2 receptor desensitization and/or downregulation. The first aim will use cortical synaptosomal preparations to study presynaptic mGlu2 receptor coupling to G-proteins after chronic 5-HT2A agonist treatment with (-)-DOB, in order to test if mGlu2 receptors are desensitized. In addition, the synaptosomal preparation will allow for the characterization of presynaptic mGlu2 and 5-HT2A receptors using traditional binding techniques. The second aim will establish a new technique for the principal investigator using an enzyme-based microelectrode array to measure glutamate neurotransmission dynamics with second-to-second temporal resolution in freely moving mice after chronic (-)-DOB treatment. Our long-term goal is to understand the mechanisms of the hyperglutamatergic state produced by repeated administration of 5-HT2A receptor agonists which may yield new strategies for controlling glutamatergic function and thus provide insight for the development of therapeutics for schizophrenia.

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Christine Saunders, Ph.D.

Regulation of Serotonin Homeostasis by RICTOR/mTOR2 Complex

Depression, a serotoninergic linked disorder, afflicts ~10% of the US population, whereas disproportionately more people with diabetes suffer from this devastating illness (~25%) that compromises their ability for self care. This link, coupled with evidence that insulin is a potent regulator of other monoaminergic systems leads to our overarching hypothesis: insulin signaling via phosphorylation of Akt controls SERT plasma membrane expression and thus 5HT homeostasis. Revealing the link between insulin and serotonin behavior (mood) will additionally lay the foundation for understanding serotonin related co-morbidities such as bulimia nervosa, post traumatic stress disorder and unipolar disorder.  Akt/PKB signaling is a key modulator of monoamine signaling and is activated by phosphorylation at 2 key residues: Ser473 by mTOR/RICTOR (mTOR complex 2) and Thr308 by PDK1 (phosphoinositide-dependant kinase 1). We have generated a fascinating model of impaired phosphorylation of Ser473 (and thereby Akt activation) by a neuronal genetic deletion of the RICTOR gene achieved with Cre-loxP technology.  This mouse model offers a unique opportunity to explore how insulin and Akt activity regulate SERT function and trafficking, and possibly in future studies, related serotoninergic behaviors.  This proposal will determine in an integrated fashion, changes in serotonin homeostasis resulting from changes in the regulation of Akt as a consequence of deletion of RICTOR/mTOR2 complex. We will define which key elements of the serotoninergic system have been modified by RICTOR deletion by coupling biochemical and electrophysiological techniques to achieve a comprehensive analysis of serotonin homeostasis.

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Jeremy Veenstra-VanderWeele, M.D.

Targeting the Serotonin Transporter Regulome to Understand Autism

Elevated platelet serotonin (5-HT), termed hyperserotemia, is the oldest biomarker in child psychiatry, but only recently has this almost 50-year-old observation begun to yield insights into pathophysiology. Building on this primary finding, recent neuroimaging studies have identified abnormalities in the brain 5-HT system as well. The serotonin transporter (SERT) has been a focus of attention in autism because of its role in taking up 5-HT into the platelet and in clearing 5- HT from the synapse. Multiple studies now implicate the SERT gene (SLC6A4) in autism, but it only accounts for a small fraction of the genetic regulation of platelet 5-HT. By mapping platelet 5- HT as a quantitative trait, we were able to identify a second gene, ITGB3, that also controls 5-HT homeostasis in the platelet. Recent studies in the Blakely lab demonstrate a physical and functional interaction between the encoded protein, integrin β3, and SERT to control uptake of 5- HT into the platelet. Remarkably, this interaction is preserved in midbrain synaptosomes, demonstrating conservation of a crucial mechanism in regulating SERT function. Confirming the importance of this interaction, three studies have now identified a gene-gene interaction between SLC6A4 and ITGB3 in association with autism. Our preliminary data in mice with decreased or absent expression of the integrin β3 gene reveals an impact on behavioral despair, response to SERT blockade, and preference for social novelty. Further studies using these mice are limited by their peripheral phenotype, which includes frequent hemorrhage and profound anemia. We propose to generate a floxed Itgb3 mouse that can be used to generate raphé-specific Itgb3 knockout by crossing with existing Pet1-Cre transgenic mice. Our initial behavioral studies will focus on findings from the somatic Itgb3 knockout mice. Subsequent work will allow us to probe 5- HT receptor sensitivity and explore the range of altered social behavior. Future studies will allow us to identify both regional and temporal specificity of these effects by crossing with tamoxifen- induced Pet1-Cre transgenic.

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Danny Winder, Ph.D.

Role of 5-HT in Amygdalar Synaptic Plasticity

Anxiety‐related disorders are a major burden on society. In addition to generalized anxiety, anxiety components emerge within other disorders including addiction and autism. 5HT plays a key role in anxiety, although the specific mechanisms involved remain elusive. Human studies have identified mutations and polymorphisms in SLC6A4 that are associated both with affective disorders as well as autism spectrum disorders. Mice with targeted deletions of SERT exhibit behavioral phenotypes consistent with this idea, and allow a window into beginning to assess mechanistic consequences of prolonged alterations in 5HT signaling. More recently a knockin mouse line has been produced by the Blakely lab, in which a polymorphism (G56A) associated with autism has been introduced into the mouse SERT locus. In contrast to the SERT knockout where 5HT clearance is impaired, studies from recombinant systems indicate that the G56A mutation produces enhanced catalytic activity of SERT as well as insensitivity to some forms of modulated transporter trafficking. Thus these two lines together allow us to assess how distinct forms of long‐term alterations in 5HT signaling affect key neuronal circuitries involved in anxiety behaviors. Actions of 5HT within the extended amygdala are thought to play important roles in constraining anxiety‐related behaviors. Indeed, in recent work, monkeys with a polymorphism in the SLC6A4 promoter were shown to exhibit enhanced activation of the bed nucleus of the stria terminalis (BNST), a component of the extended amygdala based on fMRI imaging in response to prolonged stressors. We will use a well‐characterized 5HT‐sensitive K+ current (GIRK current) in BNST neurons to assay the degree to which postsynaptic 5HT receptor sensitivity is altered in BNST neurons in these two mouse lines (see letter of support from Randy Blakely). Moreover, we will examine aspects of glutamatergic transmission in BNST neurons from these lines to begin to assess the long‐term consequences of disrupted 5HT dynamics on neuronal circuit activity.

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Bing Zhang, Ph.D.

Integrative Systems Biology Approaches to Elucidate Serotonin Network Modules and Their Regulators

Serotonin (5HT) is an important modulator of neural circuitry that controls a wide range of behavioral and physiological processes. Defective serotonergic signaling has been implicated in numerous human behavioral disorders such as impulsive violence, anxiety, and depression. In a recent study, combining genetic based tools with the microarray gene expression profiling technology, Dr. Deneris’ group has generated for the first time a comprehensive list of genes that are expressed in the 5HT neurons. However, it is not clear how these genes function and interact with one another to govern the generation of these neurons and how they are regulated. In this project, we propose that the gene expression data on 5HT neurons can be integrated with gene co-expression networks and protein interaction networks derived from publicly available datasets to identify network modules that are specifically activated in the 5HT systems. Because genes in a functional module are likely to be co-regulated, we will use computational approaches to further infer transcriptional regulatory mechanisms underlying the expression of the modules. Aim I: Identify network modules that are activated in the 5HT neurons. We will construct protein interaction networks and gene co-expression networks from publicly available data sets and identify network modules from these networks. Identified modules will be filtered through the 5HT gene expression data in rostral and caudal to screen for those that are activated in the 5HT neurons. Aim II: Identify transcriptional regulators of the 5HT-related functional modules. We hypothesize that genes in each 5HT-related module are coordinately regulated by transcriptional regulators, such as transcription factors. We will combine co-expression information, phylogenetic information, and transcription factor binding site information to identify these transcription factors. The project will not only improve our understanding of the intrinsic genetic program that governs vertebrate 5HT neuron generation, but also identify candidate regulators as potential therapeutic targets of psychiatric and neurological disorders.