Calcium Indicator Solution
Dissolve 50 µg of Fluo-4 AM dye (Thermo Fisher F14201) in 5 µL of dimethyl sulfoxide (DMSO; Sigma-Aldrich D8418) before adding 10 µL of Pluronic F-127 (Biotium 59005) and 35 µL of frog ringer to the solution. Prepare this solution on the day of the experiment with all ingredients at room temperature; store for the day at room temperature.
Centrifuge solution at 16,100g for 60 sec at room temperature. Transfer the supernatant to a new tube and discard the pellet. Add 0.3 µL of the fluorescent tracer cascade-blue dextran (Thermo Fisher D1976; 3 mm in frog ringer) to the solution to facilitate the observation of dye extrusion into the tissue.
(Optional) Add 3 µL of MK571 (50 µm in frog ringer; Alexis Biochemicals) to the calcium indicator solution.
