Fusion of secretory vesicles visualized by IRM. (A) An omega shape in a chromaffin cell from hamster adrenal medulla (figure modified from Benedeczky, 1983). (B) Schematic diagram of the experimental configuration showing an omega shape in the footprint. The image obtained by IRM was determined by reflection of light at the interface between the coverslip and medium (R₁) and the medium and cell surface (R₂). If the distance (d) between these two interfaces is of the order of the wavelength of light, then R₁ and R₂ interfere. (C) Difference images from the footprint of a chromaffin cell visualized by IRM. Average of three frames at rest (a), during stimulation (b), immediately after stimulation (c), and 13 s later (d). The stimulus was a train of 24 depolarizing steps 105 ms long, delivered at 5 Hz. (D) The relationship between the cumulative number of events counted in the footprint (red) and the increase in capacitance of the whole cell (black), from the experiment shown in C. Letters a–d show the timing of the corresponding frames in C. The lower trace showing the calcium current also indicates the timing of the stimulus. The blue line describes a single exponential fit (τ = 11.8 s) to the endocytosis phase. Intensity change as a function of time for the synchronous (white arrow) and asynchronous (red arrow) fusion events indicated in Fig. 1 C. (E) The change in area of the whole cell measured by capacitance (black) compared with the area of the footprint measured by IRM (red). Averaged results from 14 cells. Footprint area did not change significantly when vesicles fused, indicating that they did not collapse.