Figure 2. Vesicle acidification is strongly temperature dependent.
A–C, average fluorescence traces at 25°C (n = 91), 30°C (n = 83) and 35°C (n = 56). The extracellular buffer is changed twice from pH 7.4 to 5.3 and back, 12.5 s before and 5 s after 80 APs at 40 Hz (arrows). The first buffer change gives the baseline when the sypHy at the surface is quenched (hatched line). The second buffer change traps vesicles that have been endocytosed and are in the process of reacidifying. Exponential fits to the fluorescence decay of these trapped vesicles give the rate that the vesicular proton pumps restore the pH at the different temperatures (black lines). D, plotting time constants of reacidification over a temperature range from 25°C to 35°C. continuous black line is best-fit linear regression with slope of –0.45 (Pearson's r2= 0.94).