Fig. 1

Quinacrine fluorescence and VNUT staining of GLP-1 secreting cells. a GLUTag cell under brightfield illumination and b 480 nm fluorescence excitation after incubation with 5 µM quinacrine. c RFP fluorescence of an L-cell from a Glu-Cre x Rosa26tdRFP reporter mouse and d 480 nm fluorescence of the cells shown in c after incubation with 5 µM quinacrine. e Example image from total internal reflection fluorescence (TIRF) microscopy of GLUTag cells after incubation with quinacrine (5 µM). Images were collected every 50 ms for 30 s. Inset shows zoomed view of vesicle in the white box, the yellow circle is the mask within which intensities were measured. f Representative profiles of quinacrine intensity for the image in e during transient fluorescence increase events (black) and failed secretion (grey). Transient increases occurred in 21 out of 35 cells examined with a median frequency of 14.9 spikes mm⁻² s⁻¹ in responsive cells (IQR: 5.0–27.3 spikes mm⁻² s⁻¹, n = 24 movies from seven independent experiments). g Intensity profile of the vesicle labelled * in e and f, with images showing vesicle signal at various time points during the spike: i = prior to spike, ii = peak of the spike, iii = during fluorescence dissipation, iv = return to initial signal. h GLUTag cells, i mouse ileum (left) or colon (right) and j human colonic cultures immunostained for the vesicular nucleotide transporter (VNUT, green, top panel) and GLP-1 or PYY (magenta, middle panel). Bottom panel represents merged images. Scale bars represent 10 μm, apart from g (3 μm)