Identification of Genetic and Chemical Modulators of Zebrafish Mechanosensory Hair Cell Death

Abstract

Inner ear sensory hair cell death is observed in the majority of hearing and balance disorders, affecting the health of more than 600 million people worldwide. While normal aging is the single greatest contributor, exposure to environmental toxins and therapeutic drugs such as aminoglycoside antibiotics and antineoplastic agents are significant contributors. Genetic variation contributes markedly to differences in normal disease progression during aging and in susceptibility to ototoxic agents. Using the lateral line system of larval zebrafish, we developed an in vivo drug toxicity interaction screen to uncover genetic modulators of antibiotic-induced hair cell death and to identify compounds that confer protection. We have identified 5 mutations that modulate aminoglycoside susceptibility. Further characterization and identification of one protective mutant, sentinel (snl), revealed a novel conserved vertebrate gene. A similar screen identified a new class of drug-like small molecules, benzothiophene carboxamides, that prevent aminoglycoside-induced hair cell death in zebrafish and in mammals. Testing for interaction with the sentinel mutation suggests that the gene and compounds may operate in different pathways. The combination of chemical screening with traditional genetic approaches is a new strategy for identifying drugs and drug targets to attenuate hearing and balance disorders.

Superficial neuromasts facilitate non-visual feeding by larval striped bass (Morone saxatilis)

“…However, we confirmed these findings using immunocytochemical staining for S100, a calcium binding protein often found in sensory neurons (Abbate et al, 2002;Germanà et al, 2007;Heizmann, 2002). Moreover, confocal imaging showed that the reduced fluorescence of striped bass neuromasts was associated with deformed and sometimes completely missing hair cells following aminoglycoside treatment, similar to zebrafish neuromasts treated with neomycin (Harris et al, 2003;Owens et al, 2007;Owens et al, 2008). Together these results indicate that numerous structural and presumably biochemical changes occur in neuromasts exposed to neomycin and that these changes result in decreased function of the hair cells as reported previously in larval fish (Blaxter and Fuiman, 1989;Mukai, 2006).…”

Fluorescent aminoglycosides reveal intracellular trafficking routes in mechanosensory hair cells

“…We previously identified both drug treatments and genetic mutations that impede AG entry (42,82,83). Here we show that reducing the total amount of AGs entering the cell also results in a larger relative fraction of AGs being delivered to the lysosomal compartment.…”

“…Notably, not all early endocytic routes employ Rab5 (49), including routes that Using this technique, it is straightforward to quantitatively evaluate whether other compounds previously shown to protect HCs from AG exposure affect AG loading. Many such compounds have been identified in LL zebrafish screens (42)(43)(44). Here we show results for benzamil, raloxifene, ractopamine, and dynasore.…”

Modulation of dorsal root ganglion development by ErbB signaling and the scaffold protein Sorbs3

“…ENU mutagenesis and early pressure screens were carried out as described previously (Lister et al, 1999;Owens et al, 2008). Using bulk segregant analysis, the ouchless lesion was linked to chromosome 8, then fine resolution mapping of a total of 1304 individuals narrowed the region to between markers z53446 and z25210.…”