Date

May 2007

Document Type

Thesis

Degree Name

M.S.

Department

Dept. of Cell and Developmental Biology

Institution

Oregon Health & Science University

Abstract

Hair cells (HCs) are the sense receptors for the auditory and vestibular systems. I report here that asteroid, a gene identified in an ENU mutagenesis screen for deafness/vestibular dysfunction mutants, encodes vesicular glutamate transporter 3 (Vglut3). Hair cells in ast -1- fish show normal FM 1-43 uptake-indicative of normal mechanotransduction-and display no obvious morphological defects, suggesting that the ast defect is in neurotransmission. In situ hybridization shows that vglut3 is expressed in hair cells of the ear, whereas vglutl and -2 are not. A splicing defect that results in the exclusion of exon 2 from vglut3 mRNA correlates with the ast phenotype; however, the genomic lesion is not located in exonic sequence, suggesting that the lesion is likely in an intronic splicing regulatory element. Morpholinos directed against the vglut3 start codon and intron 2 splice sites delete ex on 2 in ~50% of vglut3 transcripts and cause a balance defect reminiscent of, though weaker than, the asteroid phenotype. While there is no apparent difference in synaptic ribbon number or ribbon localization in ast mutants, ultrastructural analysis reveals a decrease in the number of ribbon-associated synaptic vesicles, perhaps indicating a role for Vglut3 in synaptic vesicle biogenesis and/or trafficking. Thus Vglut3 may have roles beyond neurotransmitter transport. These results provide the first unambiguous identification of the hair cell vesicular glutamate transporter, and establish the asteroid mutation as a platform on which to conduct further investigation into hair cell synaptic biology.

Identifier

doi:10.6083/M43N21DV

School

School of Medicine

Share

COinS
 
 

To view the content in your browser, please download Adobe Reader or, alternately,
you may Download the file to your hard drive.

NOTE: The latest versions of Adobe Reader do not support viewing PDF files within Firefox on Mac OS and if you are using a modern (Intel) Mac, there is no official plugin for viewing PDF files within the browser window.