Figure. 2. Oscillations in free Ca²⁺ and Oxygen consumption in permeabilized HIT cells induced by an oscillating glycolytic muscle extract.

The reaction mixture contained 2 μmol/l fura 2 (free acid), 2 mmol/l sodium ATP, 3 mmol/l MgCl₂, 20 mmol/l HEPES adjusted to pH 7.1 with KOH, 97 mmol/l KCl, 5 mmol/l KHCO₃, 6.5 mmol/l potassium phosphate, 0.3 mmol/l GTP, 4 mmol/l sodium aspartate, 10 mmol/l glucose, 30 μmol/l NAD, 0.2 U/ml crystalline yeast hexokinase (gel filtered in 20 mmol/l HEPES), rat muscle extract equivalent to 1 mg of protein per milliliter, ~80 mg/ml saponin, and HIT cells equivalent to ~1 mg protein per milliliter. The reaction was started by addition of muscle extract, saponin, and hexokinase in rapid succession. Free Ca²⁺ was measured using the fluorescent signals of fura 2 and O₂ consumption using an O₂ electrode. Note that air was blown over the surface of the solution to balance the average rate of O₂ consumption; thus, a rise in the trace indicates a decreased rate of O₂ consumption, and a fall in the trace indicates an increased rate of O₂ consumption, compared with the average. These traces are representative of experiments that were repeated at least three times.