GABAergic inhibition is essential for circuit development and function. In the developing visual system, GABAergic inhibition determines the onset and close of the critical period for ocular dominance plasticity. Furthermore, the development of GABAergic synaptic transmission is regulated by visual experience. These studies suggest that sensory input activity regulates the development of GABAergic neurons and their synaptic connections, and that, in turn, inhibitory GABAergic synaptic transmission regulates the development and plasticity of visual circuits. Despite this postulated pivotal role for inhibition in circuit development, the inability to visualize and perturb connectivity of GABAergic inhibitory neurons in an intact developing circuit has prevented a critical evaluation of this hypothesis. We have developed tools to address both of these limitations and now propose experiments to determine mechanisms controlling the development of GABAergic neurons and their synaptic connections in the intact Xenopus tadpole optic tectum and to determine how GABAergic inputs control the development of visual receptive fields and visually guided behavior. To visualize GABAergic neurons in the intact animal, we will use the VGAT promoter to drive expression of fluorescent proteins (FPs) and other genes of interest specifically in GABAergic neurons. We will decrease synaptic transmission onto GABAergic neurons by VGAT-driven expression of peptides which we have shown specifically inhibit GABAergic or glutamatergic synaptic transmission in a cell autonomous manner.
In Aim 1, we will collect time-lapse images of individual GABAergic tectal neurons expressing FP and FP-tagged synaptic proteins to identify mechanisms that control GABAergic neuronal development. We will test the role of glutamatergic and GABAergic synaptic transmission in GABAergic neuron development by expressing peptides to block synaptic transmission onto the imaged cell.
In Aim 2, we will combine in vivo time-lapse imaging, to identify stable and dynamic dendrites, and retrospective serial section electron microscopy (EM) to determine the synaptic rearrangements that occur during GABAergic neuronal development. Data from Aims 1 and 2 will demonstrate the activity-dependent mechanisms that control the morphological development and synaptic connectivity of GABAergic neurons in vivo.
In Aims 3 and 4 we will test the effect of blocking inhibitory and excitatory transmission on tectal visual receptive field properties, using cell-attached and whole recordings, and a visual avoidance assay, that will extend our studies into the behavioral arena. The aberrant development and function of inhibitory circuits is thought to underlie a variety of developmental neurological disorders, including autism, schizophrenia and depression, however the basic mechanisms governing the development of inhibitory circuits are relatively unknown. The experiments proposed here should illuminate activity-dependent mechanisms of GABAergic circuit development in vivo.

Public Health Relevance

This project will determine the mechanisms governing the development of GABAergic inhibitory neurons and their synaptic connections in the intact developing brain. We will determine the role of inhibitory neurons in perceiving visual information and in visually guided behavior. These studies will help us understanding developmental errors in brain circuit formation.

National Institute of Health (NIH)
National Eye Institute (NEI)
Research Project (R01)
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Central Visual Processing Study Section (CVP)
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Steinmetz, Michael A
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Scripps Research Institute
La Jolla
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Shen, Wanhua; Liu, Han-Hsuan; Schiapparelli, Lucio et al. (2014) Acute synthesis of CPEB is required for plasticity of visual avoidance behavior in Xenopus. Cell Rep 6:737-47
Hiramoto, Masaki; Cline, Hollis T (2014) Optic flow instructs retinotopic map formation through a spatial to temporal to spatial transformation of visual information. Proc Natl Acad Sci U S A 111:E5105-13
Bestman, Jennifer E; Cline, Hollis T (2014) Morpholino studies in Xenopus brain development. Methods Mol Biol 1082:155-71
McKeown, Caroline R; Sharma, Pranav; Sharipov, Heidi E et al. (2013) Neurogenesis is required for behavioral recovery after injury in the visual system of Xenopus laevis. J Comp Neurol 521:2262-78
Lee, Ping-Chang; He, Hai-Yan; Lin, Chih-Yang et al. (2013) Computer aided alignment and quantitative 4D structural plasticity analysis of neurons. Neuroinformatics 11:249-57
Sharma, Pranav; Schiapparelli, Lucio; Cline, Hollis T (2013) Exosomes function in cell-cell communication during brain circuit development. Curr Opin Neurobiol 23:997-1004
Bestman, Jennifer E; Lee-Osbourne, Jane; Cline, Hollis T (2012) In vivo time-lapse imaging of cell proliferation and differentiation in the optic tectum of Xenopus laevis tadpoles. J Comp Neurol 520:401-33
Miraucourt, Lois S; Silva, Jorge Santos da; Burgos, Kasandra et al. (2012) GABA expression and regulation by sensory experience in the developing visual system. PLoS One 7:e29086
Shen, Wanhua; McKeown, Caroline R; Demas, James A et al. (2011) Inhibition to excitation ratio regulates visual system responses and behavior in vivo. J Neurophysiol 106:2285-302
Li, Jianli; Erisir, Alev; Cline, Hollis (2011) In vivo time-lapse imaging and serial section electron microscopy reveal developmental synaptic rearrangements. Neuron 69:273-86

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