STUDYING NEURONAL PEPTIDE RELEASE AND SECRETORY GRANULE DYNAMICS WITHGREEN FLUORESCENT PROTEIN

Neurons and endocrine cells release peptides stored in a limited pool of secretory granules. Although it has been possible to measure secret ion or exocytosis, studying events within cells that influence the siz e and speed of secretory responses has been difficult. Here we describ e how green fluorescent protein (GFP)-tagged hormones can be used to m easure release and granule mobility. Epifluorescence allows one to mea sure total peptide content and granule trajectories within a plane of focus. Confocal microscopy facilitates measurements within a better de fined optical section. Granule mobility can also be measured with fluo rescence recovery after photobleaching (FRAP). Application of these op tical approaches to the study of neuronal peptide secretion has reveal ed that the releasable pool is not spatially defined and that sustaine d peptide release is limited by the availability of mobile secretory g ranules. granules. (C) 1998 Academic Press.