Philip Washbourne - US grants

DESCRIPTION (provided by applicant): Chemical synapses are the primary means for transmitting information from one neuron to the next. Synapses are initially formed during development of the nervous system, and formation of appropriate synapses is crucial for establishment of neuronal circuits that underlie behavior and cognition. Minor irregularities during synapse formation can lead to developmental disorders such as autism, mental retardation and may contribute to psychological disorders. Most synapses in the vertebrate central nervous system (CNS) depend on the neurotransmitter glutamate, and thus glutamatergic synapses have been an important focus of study in trying to unravel these and other neurological disorders. A novel family of cell adhesion molecules (CAMs), the Synaptic Cell Adhesion Molecules (SynCAMs), has recently been proposed to mediate the formation of synapses. However, this work is based on experiments in neuronal culture, and knock-out mouse data so far does not corroborate this. Furthermore, it remains unknown how the SynCAMs, and CAMs in general, bring about the recruitment of synaptic elements to new adhesive contacts. We propose to test a model describing specific mechanisms through which SynCAM family members 1 and 2 can recruit both synaptic vesicles (SVs) to the presynaptic terminal and glutamate receptors to the postsynaptic specialization. We hypothesize that an interaction between SynCAM1 or 2 and CASK in axons can directly tether SV precursors to the site of SynCAM/SynCAM interaction. We also propose that binding of DAL-1 to SynCAM1 or 2 in dendrites results in formation of an actin/spectrin subsynaptic scaffold. These cytoskeletal elements then serve two functions: 1) strengthening of the adhesive nature of the synapse and morphological remodeling to generate a spine and 2) recruitment of NMDA type glutamate receptor transport packets via an actin-dependent transport mechanism. We propose to use a variety of techniques including biochemistry, immunolabeling, live-imaging, electrophysiology and behavioral tests, because a multidisciplinary approach will comprehensively test our model. We also propose to use various neuronal preparations for our experiments including cultured hippocampal neurons, cultured cerebellar granule cells (CGCs) and spinal cord neurons in zebrafish embryos in vivo. Testing our hypothesis in zebrafish will shed light on whether these proteins and their interactions are required for forming a specific circuit in a developing embryo that is required for a sensorimotor reflex. Our approach gives us the unprecedented opportunity to determine the mechanisms of glutamatergic synapse formation using behavioral, electrophysiological, genetic and biochemical approaches in both neuronal cultures and in a living vertebrate. PUBLIC HEALTH RELEVANCE: Synapses are sites at which nerve cells communicate with each other. Communication of nerve cells is absolutely necessary for nervous system function, ranging from simple reflexes to expressing philosophical thoughts. Errors during the formation of synapses are thought to be at the basis of nervous system disorders such as autism, mental retardation and schizophrenia. We propose to study two members of a family of genes, the Synaptic Cell Adhesion Molecules (SynCAMs) 1 and 2, which are thought to mediate the formation of synapses during development. Almost nothing is yet known about how SynCAMs make a contact site between two nerve cells become a place for active communication. We will investigate how these molecules carry out synapse formation by testing their function in rat nerve cells grown in culture and by testing their function in developing zebrafish embryos. These two systems allow us to determine the nature of the molecular interactions and to determine the importance of these interactions for the formation of synapses in a living organism, respectively. Our research will help understand the mechanisms by which synapses form and bring us closer to identifying the molecular deficits in individuals with autism and mental retardation.

DESCRIPTION (provided by applicant): Both genetic and environmental factors play significant roles in influencing and perhaps causing neurodevelopmental disorders such as autism, mental retardation, and schizophrenia. Recent advances in genetic analysis have revealed a number of genes linked to familial forms of these disorders. Very little is known, however, about how the environment may contribute to these disorders, or how the environment may interact with identified genetic causes. We propose that abnormal microbial colonization of the gut during development leads to aberrant expression levels of genes that are involved in brain development, specifically in synapse formation. We hypothesize that incorrect microbial colonization of the gut may exacerbate or drive behavioral deficits by promoting aberrant synapse formation that results in altered neuronal circuitry and function. We propose to examine interactions between early gut colonization by the microbiota and expression of synaptic cell adhesion molecules, such as Neuroligins, during development in zebrafish. This model provides an unparalleled opportunity to study the consequences of altered gut microbial colonization on synapse formation, development of neuronal circuitry, neuronal activity, and behavior, because we can manipulate both host genetics and microbial communities and follow development of defined neuronal populations in real time in living juveniles and adults. We also propose to determine whether there is a critical period during which colonization by a microbiota whose members express specific traits is required for normal synapse development, and to learn how the microbiota signals to the developing brain to promote the normal synapse formation required for normal behavior. Finally, we will test directly whether a dysbiotic, pro-inflammatory microbiota can alter synapse development, formation of neuronal circuitry, neuronal activity, and behavior, in both young and adult animals. Our proposed experiments will provide the first comprehensive view of how microbial signals affect development of neuronal anatomy and physiology and how this affects behavior at later stages of development and in adults. Revealing and characterizing an interaction between the microbiota and the genes that drive synapse formation would have a dramatic impact on current treatment approaches for individuals diagnosed with neurodevelopmental disorders.