Neomycin-induced hair cell death and rapid regeneration in the lateral line of zebrafish (Danio rerio)

被引:383
作者
Harris, JA
Cheng, AG
Cunningham, LL
MacDonald, G
Raible, DW
Rubel, EW
机构
[1] Univ Washington, VMB Hearing Res Ctr, Seattle, WA 98195 USA
[2] Univ Washington, Dept Otolaryngol Head & Neck Surg, Seattle, WA 98195 USA
[3] Univ Washington, Grad Program Neurobiol & Behav, Seattle, WA 98195 USA
[4] Univ Washington, Dept Biol Struct, Seattle, WA 98195 USA
[5] Univ Washington, Core Commun Res, Seattle, WA 98195 USA
来源
JARO-JOURNAL OF THE ASSOCIATION FOR RESEARCH IN OTOLARYNGOLOGY | 2003年 / 4卷 / 02期
关键词
hair cell death; lateral line; aminoglycosides; hair cell regeneration; zebrafish; genetic screening;
D O I
10.1007/s10162-002-3022-x
中图分类号
Q189 [神经科学];
学科分类号
071006 ;
摘要
Mechanoreceptive hair cells are extremely sensitive to aminoglycoside antibiotics, including neomycin. Hair cell survival was assessed in larval wild-type zebrafish lateral line neuromasts 4 h after initial exposure to a range of neomycin concentrations for 1 h. Each of the lateral line neuromasts was scored in live fish for the presence or absence of hair cells using the fluorescent vital dye DASPEI to selectively label hair cells. All neuromasts were devoid of DASPEI-labeled hair cells 4 h after 500 muM neomycin exposure. Vital DASPEI staining was proportional to the number of hair cells per neuromast identified in fixed larvae rising immunocytochemistry for acetylated tubulin and phalloidin labeling. The time course of hair cell regeneration in the lateral line neuromasts was also analyzed following neomycin-induced damage. Regenerated hair cells were first observed using live DASPEI staining 12 and 24 h following neomycin treatment. The potential role of proliferation in regenerating hair cells was analyzed. A 1 h pulse-fix protocol using bromodeoxyuridine (BrdU) incorporation was used to identify S-phase cells in neuromasts. BrdU incorporation in neomycin-damaged neuromasts did not differ from control neuromasts 4 h after drug exposure but was dramatically upregulated after 12 h. The proliferative cells identified during a 1 h period at 12 h after neomycin treatment were able to give rise to new hair cells by 24-48 h after drug treatment. The results presented here provide a standardized preparation for studying and identifying genes that influence vertebrate hair cell death, survival, and regeneration following ototoxic insults.
引用
收藏
页码:219 / 234
页数:16
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