Molecular mechanisms of zebrafish motoneuron development
Hale, Laura Ann, 1978-
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Hale, Laura Ann, 1978-
This dissertation describes research to identify genes involved in specification, patterning and development of zebrafish primary motoneurons. We first examined the spatiotemporal expression patterns of retinoic acid and retinoid X receptor mRNAs to determine whether particular ones might be involved in motoneuron specification or patterning. Retinoic acid and retinoid X receptor mRNAs are expressed at the right time to pattern motoneurons, but the expression patterns did not suggest roles for particular receptors. In contrast, netrin mRNAs are expressed in specific motoneuron intermediate targets and knockdown experiments revealed an important role in development of VaP motoneurons. Two identified motoneurons, CaP and VaP, initially form an equivalence pair. CaPs extend long axons that innervate ventral muscle. VaPs extend short axons that stop at muscle fibers called muscle pioneers; VaPs later typically die. Previous work showed that during extension, CaP axons pause at several intermediate targets, including muscle pioneers, and that both CaP and muscle pioneers are required for VaP formation. We found that mRNAs for different Netrins are expressed in intermediate targets before CaP axon contact: netrin 1a in muscle pioneers, netrin 1b in hypochord, and netrin 2 in ventral somite. We show that Netrins are unnecessary to guide CaP axons but are necessary to prevent VaP axons from extending into ventral muscle. Netrin 1a is necessary to stop VaP axons at muscle pioneers, Netrin 1a and Netrin 2 together are necessary to stop VaP axons near the hypochord, and Netrin 1b appears dispensable for CaP and VaP development. We also identify Deleted in colorectal carcinoma as a Netrin receptor that mediates the ability of Netrin 1a to cause VaP axons to stop at muscle pioneers. Our results suggest Netrins refine axon morphology to ensure final cell-appropriate axon arborization. To learn whether Netrin proteins diffuse away from their sources of synthesis to function at a distance, we are developing Netrin antibodies. If successful, the antibodies will provide the research community at large with a new tool for understanding in vivo Netrin function. This dissertation includes both my previously published and unpublished coauthored material.