Dimethyl chlorosilane

To construct ion-selective microelectrodes, glass capillary tubes (no. TW 150–4; World Precision Instruments, Sarasota, FL, USA) were pulled on a Sutter P-97 Flaming Brown pipette puller (Sutter Instruments, San Rafael, CA, USA) into micropipettes with tip diameters of 3~4 μm. The micropipettes were then baked at 120°C overnight and coated with dimethyl chlorosilane (Sigma-Aldrich) for 30 min. The micropipettes were backfilled with a 1-cm column of 100 mM CaCl₂ for the Fluka Ca²⁺-selective microelectrode. The microelectrode was then frontloaded with a 20~30-μm column of Ca²⁺ ionophore I cocktail A (Sigma-Aldrich) to create a Ca²⁺-selective microelectrode.

SIET was used to measure Na⁺ flux activity at the epithelium surface of medaka larva. Glass capillary tubes (no. TW 150 – 4; World Precision Instruments, Sarasota, FL) were pulled on a Sutter P-97 Flaming Brown pipette puller (Sutter Instruments, San Rafael, CA) into micropipettes with tip diameters of 3–4 μm. The micropipettes were then baked at 120 °C overnight and coated by incubation with dimethyl chlorosilane (Sigma-Aldrich) for 30 min. The micropipettes were backfilled with a 1-cm column of electrolytes and frontloaded with a 20–30 μm column of liquid ion-exchange cocktail (Sigma-Aldrich) to create an ion-selective microelectrode (probe). The ionophore cocktail (and electrolytes) was Na⁺ ionophore II cocktail A (100 mM NaCl). To calibrate the ion-selective probe, the Nernstian response of each microelectrode was evaluated by placing it in a series of standard solutions (0.1, 1, and 10 mM NaCl dissolved in distilled water). By plotting the voltage output of the probe against log [Na⁺] value, linear regression yielded a Nernstian slope of 56.7 ± 0.5 (N = 10). In preliminary tests, the selectivity of the Fluka Na⁺ ionophore II cocktail A was 10–16 times more selective to Na⁺ than to NH₄⁺ (measured in 1–10 mM Na⁺ solution).