There are numerous clinical conditions in which altered sensory signaling from the face, head and/or viscera are manifested, e.g. peripheral neuropathies arising from diabetes, HIV/AIDS and herpes, trigeminal neuralgia, the various hereditary sensory neuropathies, neuropathies arising from trauma or surgical outcomes, and others. We have recently engineered a transgenic fish line that expresses an eGFP marker protein in neurons of the cranial sensory ganglia (CSG). The eGFP produced is soluble and fills the cell body together with its peripheral and central projections. This expression begins at the time these neurons are forming the ganglia and extending their axonal processes, and is maintained through at least three weeks of age. We propose using these transgenic fish in a ethylnitrosourea-based mutagenesis screen in an effort to identify genes involved in any of the following three processes: formation of the individual cranial ganglia, innervation of specific targets by different ganglia, and innervation of different targets by individual neurons within the same ganglia. There are two aims for the work proposed in this application: (1) to introduce ENU mutated genes into the TG(3.2::eGFPGR) line and use an F3 recessive screening paradigm to identify carrier families that produce embryos exhibiting defects in CSG development and connectivities and (2) to characterize the identified mutants. We have initiated a preliminary screen using a small number of carrier families (21). To date, we have already identified two mutants (out of 11 families screened) that have developmental abnormalities in their CSG by 96 hpf. Allele sl14 exhibits disrupted formation of the trigeminal ganglia while having normal epibranchial ganglia, whereas allele sl19 has improper targeting of the central and peripheral projections of the epibranchial ganglia while the trigeminal ganglia appear unaltered. This R21 proposal is designed to increase the size of our screen to 150-200 families (genomes) in order to generate a body of mutants that cover the wide spectrum of mechanisms responsible for CSG formation. A successful outcome of this proposal will also strengthen a nascent collaboration between the Voigt lab (SLU), which is focused on peripheral neurons and signaling issues and is adopting zebrafish as a model, and the Appel lab (Vanderbilt), an established zebrafish lab which is interested in mechanisms underlying the myelination of central and peripheral neurons.
Millions of people worldwide suffer degraded health and quality of life issues due to peripheral sensory neuropathies. Currently, treatment of such disorders is limited by the lack of clinically useful tools. The principal goal of this project is to provide a springboard for the development of new approaches to the treatment of these diseases by identifying genes, and thus potential therapeutic targets, involved in the normal development of the sensory nervous system.
