Extracellular Ca²⁺ and Mg²⁺ modulate aminoglycoside blockade of mechanotransducer channel-mediated Ca²⁺ entry in zebrafish hair cells: An in vivo study with the SIET

Zebrafish lateral-line hair cells are an in vivo model for studying hair cell development, function, and ototoxicity. However, the molecular identification and properties of the mechanotransducer (MET) channel in hair cells are still controversial. In this study, a noninvasive electrophysiological method, the scanning ion-electrode technique (SIET), was applied for the first time to investigate properties of MET channels in intact zebrafish embryos. With the use of a Ca²⁺-selective microelectrode to deflect hair bundles and simultaneously record the Ca²⁺ flux, the inward Ca²⁺ flux was detected at stereo cilia of hair cells in 2- to ~4-day postfer-tilization embryos. Ca²⁺ influx was blocked by MET channel blockers (BAPTA, La³⁺, Gd³⁺, and curare). In addition, 10 μM amino glycoside antibiotics (neomycin and gentamicin) were found to effectively block Ca²⁺ influx within 10 min. Elevating the external Ca²⁺ level (0.2-2 mM) neutralized the effects of neomycin and gentamicin. However, elevating the Mg²⁺ level up to 5 mM neutralized blockade by gentamicin but not by neomycin. This study demonstrated MET channel-mediated Ca²⁺ entry at hair cells and showed that the SIET to be a sensitive approach for functionally assaying MET channels in zebrafish.