A goal of neurodevelopmental biology is to identify mechanisms by which normal paterning of the nervous system occurs to ultimately apply this knowledge to understanding neurodevelopmental disorders. Activity-dependent processes have been recognized as important mechanisms for early neurodevelopment and electrical synapses have been suggested to play an integral role in the establishment and refinement of neuronal networks. Electrical synapses are formed with gap junction communications between neuronal cells that allow for the direct transfer of current. The proteins responsible for most gap junction formations, connexins, have been implicated in a number of neurodevelopmental processes. However, increasing evidence indicates connexins play more than a pore-forming role, but may also participate in other intracellular and extracellular interactions that can have profound effects on development. The goal of the current study is to determine the role neuronal specific connexins play in neurodevelopment by exploring the many possible mechanisms by which these protein complexes may mediate developmental control. This study takes advantage of the Danio rerio (zebrafish) model to determine whether the neuronal specific connexin 35 (Cx35) plays a fundamental role in the development of the spinal cord. The two specific aims will test the hypotheses that (1) proper development of the zebrafish spinal cord requires developmentally regulated expression of Cx35 and (2) the mechanism by which Cx35 mediates neurodevelopment may be executed by pore function and/or intracellular interactions. The experimental approach utilizes a number of tools available in zebrafish, such as the availability of many transgenic lines, easy genetic manipulation, live- imaging, and pharmacological approaches.
Aim 1 will use in situ hybridization (ISH) and immunohistochemistry (IHC) to determine the cx35 mRNA expression profile and morpholino knockdown to determine if Cx35 is required for proper development of spinal cord cells in order to elucidate the cellular networks that require Cx35 for proper development.
Aim 2 wil use a pharmacological approach to uncouple gap junctions and the production of transgenic lines that express mutant forms of Cx35 to determine whether Cx35 developmental control is mediated by pore-dependent and/or pore-independent molecular processes. The role of connexin proteins in early development has not been fuly explored. Much of the current knowledge regarding connexins in the nervous system is on their role in fine-tuning already established networks. Investigating the role of connexins in early development may help deepen our understanding of the importance of these proteins in establishing complex neuronal networks and determining cellular development.

Public Health Relevance

Some human diseases with neurological symptoms have been linked to mutations in connexin genes, such as oculodentodigital dysplasia (ODDD) and a recessive hypomyelinating leukoencephalopathy called Pelizaeus- Merzbacher-like disease (PMLD). Additionally, electrical synapses are responsible for generating synchrony among coupled cells and atypical neural synchrony has been suggested to play a role in some developmental neurological diseases, such as Autism Spectrum Disorders. However, the mechanisms by which connexins mediate neurodevelopmental control is largely unknown, but investigations into the role connexins play in patterning the nervous system may provide insight into some neurological disease processes.

National Institute of Health (NIH)
National Institute of Neurological Disorders and Stroke (NINDS)
Predoctoral Individual National Research Service Award (F31)
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NST-2 Subcommittee (NST)
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Riddle, Robert D
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University of Colorado Denver
Schools of Medicine
United States
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Carlisle, Tara C; Ribera, Angeles B (2014) Connexin 35b expression in the spinal cord of Danio rerio embryos and larvae. J Comp Neurol 522:861-75