Measuring peptide secretion using the FMRFamide tagging technique.
Peptides are transmitters produced by a wide variety of neurons and neuroendocrine cells. They mediate a remarkable range of physiological processes. To better understand the roles played by peptides, a number of methods have been developed that can monitor their secretion. Although each has particular strengths, they cannot rapidly detect the secretion of chemically defined peptides. However, a recently developed approach termed "FMRFamide-tagging" may be useful in this regard. A genetically encoded electrophysiological tag is attached to the peptide prohormone of interest. The "tagged" prohormone together with an ionotropic receptor that binds the tag are expressed in the cell type under investigation. Secretion of the tag (and the co-secreted peptide of interest) are revealed by rapid inward membrane currents that are due to the activation of the tag receptor. In this manner, peptide secretion can be followed on a millisecond time scale. This protocol gives the details of the approach and its potential application to a range of cell types.
Whim
Matthew D
MD
Department of Pharmacology, University College London, Gower Street, London WC1E 6BT, UK. m.whim@ucl.ac.uk
D000818:Animals; D002118:Calcium / Q000378:metabolism; D002478:Cells, Cultured; D019439:Chromaffin Cells / Q000737:chemistry / Q000166:cytology / Q000378:metabolism; D005110:Extracellular Space / Q000737:chemistry; D019835:FMRFamide / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D049452:Green Fluorescent Proteins; D007150:Immunohistochemistry / Q000295:instrumentation / Q000379:methods; D008164:Luminescent Proteins / Q000096:biosynthesis / Q000235:genetics; D008564:Membrane Potentials; D051379:Mice; D009490:Neurosecretory Systems / Q000737:chemistry / Q000166:cytology / Q000378:metabolism; D018408:Patch-Clamp Techniques / Q000295:instrumentation / Q000379:methods; D010455:Peptides / Q000235:genetics / Q000378:metabolism*; D010957:Plasmids / Q000096:biosynthesis / Q000235:genetics; D051381:Rats; D018010:Receptors, Invertebrate Peptide / Q000096:biosynthesis / Q000235:genetics; D022142:Secretory Vesicles / Q000737:chemistry / Q000378:metabolism; D014162:Transfection / Q000295:instrumentation / Q000379:methods
C055209:FMRFamide receptor; D008164:Luminescent Proteins; D010455:Peptides; D018010:Receptors, Invertebrate Peptide; D049452:Green Fluorescent Proteins; D019835:FMRFamide; D002118:Calcium