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- W2122326359 abstract "The morphological dynamics of the plasma membrane were visualized in bovine adrenal chromaffin cells using polarized total internal reflection fluorescence microscopy (TIRFM). This method is based on monitoring the fluorescence of an oriented membrane probe (the carbocyanine dye, DiI) excited by a polarized evanescent field created by TIR illumination. DiI has been shown to embed in the membrane with its transition dipole moments nearly in the plane of the membrane. Thus, by monitoring the pixel-by-pixel ratio of the membrane-embedded DiI fluorescence excited by the two polarizations (p - perpendicular to substrate; s - parallel to substrate) over time, regions of membrane curvature are vividly highlighted. To relate the orientation of the membrane with exocytosis, granules were labeled with the marker neuropeptide (NPY) - cerulean. In response to high KCl depolarization, fusion of granules coincided with 15-20% increases in DiI-membrane p/s values at locations of NPY-Cer release. The p/s values then often declined over several seconds to approximately pre-fusion levels. In other instances, the p/s values declined more slowly providing evidence of longer-lasting membrane curvature. Some granules were associated with areas of the membrane with increased curvature (larger p/s values) prior to stimulation. These granules were significantly more likely to undergo exocytosis than those without such associations. When endocytosis was inhibited with Dynasore (inhibitor of dynamin 1 and 2), membrane p/s changes following fusion were long-lived (10s of seconds). This is consistent with endocytosis occuring at the site of exocytosis. These experiments are the first to directly visualize plasma membrane changes occuring with secretion. They reveal an association of the granule and plasma membrane for seconds before exocytosis and provide direct evidence for maintained curvature of the plasma membrane after exocytosis." @default.
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- W2122326359 date "2009-02-01" @default.
- W2122326359 modified "2023-09-29" @default.
- W2122326359 title "The Dynamics Of Secretion-associated Plasma Membrane Changes Visualized With Polarized TIRFM" @default.
- W2122326359 doi "https://doi.org/10.1016/j.bpj.2008.12.3737" @default.
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