Effect of PMR1 silencing on glucose-stimulated insulin secretion. A: MIN6 cells were cultured for 16 h at 3 mmol/l glucose and then incubated with KRBB supplemented with 3 mmol/l glucose for
15 min. Cells were then incubated for a further 40 min in KRBB supplemented with 3, 11, or 30 mmol/l glucose, or with 11 mmol/l
glucose plus 10 mmol/l TEA. Released and total insulin were measured by radioimmunoassay. Data are the means of at least three
separate experiments. Basal insulin secretion (at 3 mmol/l glucose) was not significantly different in control and PMR1-depleted
cells (0.12 and 0.11% of total insulin per 40 min, respectively; means of two separate experiments). B: After culturing for 16 h at 3 mmol/l glucose, MIN6 cells were loaded with fluo-3-AM for 40 min in KRBB containing 3 mmol/l
glucose. Cells were then challenged with KRBB supplemented with 11 mmol/l glucose plus 10 mmol/l TEA. Ca2+ oscillations were monitored on a Olympus IX-70 inverted microscope (excitation 480 nm, emission 515 nm). C: Average [Ca2+]cyt calculated by integrating [Ca2+] over the final 5 min of stimulation with 11 mmol/l glucose and 10 mmol/l TEA for control and PMR1 siRNA-treated cells. D–F: Amplitude (D), duration (measured as indicated in B) (E), and decay rate (F) of slow [Ca2+]cyt oscillations during the final 5 min of stimulation with 11 mmol/l glucose and 10 mmol/l TEA for control and PMR1 siRNA-treated
cells. C–F: Data are the means of at least 10 cells in each case. □, control; ▪, PMR1−.