427
4.1 Hair Cell Death and Protection
Inner ear ototoxicity studies generally rely on daily systemic aminoglycoside injections spanning multiple days (e.g., Lombarte et al. 1993 ; Ramcharitar and Selckmann
2010 ). Intramuscular injection is simple to administer but often causes morbidity
due to nephrotoxic side effects. One group has recently employed direct intrasaccular gentamicin injection using X-ray-assisted needle placement in Atlantic cod
( Gadus morhua ) (Faucher et al. 2008a , 2009 ), an elegant approach that bypasses
systemic toxicity, although the systemic model more closely approximates human
clinical use. Using either treatment paradigm, most fi sh inner ear aminoglycoside
research is descriptive rather than mechanistic. Several studies in the oscar employed
SEM imaging of dissected sensory epithelia to demonstrate selective loss of striolar
hair cells in the utricle and lagena (Yan et al. 1991 ; Chang et al. 1992 ; Lombarte
et al. 1993 ), consistent with fi ndings of increased toxin sensitivity in mammalian
striolar hair cells (reviewed in Salvi et al. 2008 ). While these earlier studies did not
report evidence of saccular hair cell loss, Ramcharitar and Selckmann ( 2010 ) and
Uribe et al. ( 2013 ) demonstrated hair cell loss in the caudal region of the saccule in
goldfi sh and zebrafi sh, respectively, correlated with a signifi cant hearing threshold
shift in the low frequency range. These ototoxicity studies inform future biomedical
work using the fi sh inner ear as a model system.
In contrast to inner ear research, many lateral line biomedical studies are mechanistic or translational. This body of work primarily relies on a tractable genetic
model, the zebrafi sh. Over a decade ago, Williams and Holder ( 2000 ) and Harris
et al. ( 2003 ) demonstrated that hair cells of 5-day-old zebrafi sh larvae respond to
aminoglycoside damage similarly to mature mammalian hair cells, setting the stage
for a myriad of studies aimed at understanding the intracellular death cascade initiated by chemical ototoxins and at identifying novel therapeutics that protect hair
cells from drug damage.
Studies on zebrafi sh larvae demonstrate that mitochondrial swelling and a loss of
mitochondrial membrane potential occur shortly after treatment with the ototoxin
neomycin. These data are consistent with genetic and pharmacologic evidence for
the importance of Bcl-2 family proteins that regulate mitochondrial-dependent cell
death pathways (Owens et al. 2007 ; Coffi n et al. 2013a , b ). Mitochondria are a
major calcium store in many cell types, and an elegant series of experiments with
transgenic zebrafi sh expressing a genetically encoded calcium sensor clearly demonstrate that calcium dysregulation plays an important role in aminoglycosideinduced hair cell loss (Esterberg et al. 2013 , 2014 ). Neomycin treatment fi rst leads
to a decrease in calcium in the endoplasmic reticulum, following by an increase in
mitochondrial calcium and ending with a cytoplasmic calcium spike immediately
preceding hair cell death (Esterberg et al. 2013 , 2014 ). Nuclear condensation, a
hallmark of classical apoptosis, occurs in aminoglycoside-damaged hair cells,
although the role of caspases (cysteine-dependent proteases associated with apoptosis) has not been conclusively demonstrated (Williams and Holder 2000 ; Santos
et al. 2006 ; Ou et al. 2009 ; Coffi n et al. 2013a ). Collectively, these studies are conChemical Ototoxicity of the Fish Inner Ear and Lateral Line
4.1 Hair Cell Death and Protection
Inner ear ototoxicity studies generally rely on daily systemic aminoglycoside injections spanning multiple days (e.g., Lombarte et al. 1993 ; Ramcharitar and Selckmann
2010 ). Intramuscular injection is simple to administer but often causes morbidity
due to nephrotoxic side effects. One group has recently employed direct intrasaccular gentamicin injection using X-ray-assisted needle placement in Atlantic cod
( Gadus morhua ) (Faucher et al. 2008a , 2009 ), an elegant approach that bypasses
systemic toxicity, although the systemic model more closely approximates human
clinical use. Using either treatment paradigm, most fi sh inner ear aminoglycoside
research is descriptive rather than mechanistic. Several studies in the oscar employed
SEM imaging of dissected sensory epithelia to demonstrate selective loss of striolar
hair cells in the utricle and lagena (Yan et al. 1991 ; Chang et al. 1992 ; Lombarte
et al. 1993 ), consistent with fi ndings of increased toxin sensitivity in mammalian
striolar hair cells (reviewed in Salvi et al. 2008 ). While these earlier studies did not
report evidence of saccular hair cell loss, Ramcharitar and Selckmann ( 2010 ) and
Uribe et al. ( 2013 ) demonstrated hair cell loss in the caudal region of the saccule in
goldfi sh and zebrafi sh, respectively, correlated with a signifi cant hearing threshold
shift in the low frequency range. These ototoxicity studies inform future biomedical
work using the fi sh inner ear as a model system.
In contrast to inner ear research, many lateral line biomedical studies are mechanistic or translational. This body of work primarily relies on a tractable genetic
model, the zebrafi sh. Over a decade ago, Williams and Holder ( 2000 ) and Harris
et al. ( 2003 ) demonstrated that hair cells of 5-day-old zebrafi sh larvae respond to
aminoglycoside damage similarly to mature mammalian hair cells, setting the stage
for a myriad of studies aimed at understanding the intracellular death cascade initiated by chemical ototoxins and at identifying novel therapeutics that protect hair
cells from drug damage.
Studies on zebrafi sh larvae demonstrate that mitochondrial swelling and a loss of
mitochondrial membrane potential occur shortly after treatment with the ototoxin
neomycin. These data are consistent with genetic and pharmacologic evidence for
the importance of Bcl-2 family proteins that regulate mitochondrial-dependent cell
death pathways (Owens et al. 2007 ; Coffi n et al. 2013a , b ). Mitochondria are a
major calcium store in many cell types, and an elegant series of experiments with
transgenic zebrafi sh expressing a genetically encoded calcium sensor clearly demonstrate that calcium dysregulation plays an important role in aminoglycosideinduced hair cell loss (Esterberg et al. 2013 , 2014 ). Neomycin treatment fi rst leads
to a decrease in calcium in the endoplasmic reticulum, following by an increase in
mitochondrial calcium and ending with a cytoplasmic calcium spike immediately
preceding hair cell death (Esterberg et al. 2013 , 2014 ). Nuclear condensation, a
hallmark of classical apoptosis, occurs in aminoglycoside-damaged hair cells,
although the role of caspases (cysteine-dependent proteases associated with apoptosis) has not been conclusively demonstrated (Williams and Holder 2000 ; Santos
et al. 2006 ; Ou et al. 2009 ; Coffi n et al. 2013a ). Collectively, these studies are conChemical Ototoxicity of the Fish Inner Ear and Lateral Line