High resolution electrophysiological techniques for the study of calcium-activated exocytosis.
BACKGROUND
Neurotransmitters, neuropeptides and hormones are released from secretory vesicles of nerve terminals and neuroendocrine cells by calcium-activated exocytosis. A key step in this process is the formation of a fusion pore between the vesicle membrane and the plasma membrane. Exocytotic fusion leads to an increase in plasma membrane area that can be measured as a proportional increase in plasma membrane capacitance.


METHODS
High resolution capacitance measurements in single cells, nerve terminals and small membrane patches have become possible with the development of the patch clamp technique. This review discusses the methods of whole cell patch clamp capacitance measurements and their use in conjunction with voltage clamp pulse stimulation and with stimulation by photorelease of caged calcium. It also discusses patch capacitance measurements for the study of single exocytotic events and fusion pore properties in neuroendocrine cells and nerve terminals.


CONCLUSIONS
Capacitance measurements provide high resolution information on the extent and time course of fusion for the characterization of vesicle pools and the kinetics of exocytosis. They allow the characterization of the mode of fusion including distinction of single vesicle full fusion, transient kiss-and-run fusion or multivesicular compound exocytosis. Furthermore, measurement of fusion pore conductances and their dynamic behavior has enabled the characterization of fusion pore properties in a way that resembles the characterization of ion channel function through single channel recordings.


CONCLUSIONS
The combination of patch clamp capacitance measurements with pharmacological and molecular manipulations of exocytosis is emerging as a powerful approach to investigate the molecular mechanisms of calcium-activated exocytotic fusion pore formation. This article is part of a Special Issue entitled Biochemical, biophysical and genetic approaches to intracellular calcium signalling.
School of Applied and Engineering Physics, Cornell University, Ithaca, NY 14853, USA. ml95@cornell.edu
Manfred
M
Lindau
D000465:Algorithms; D000818:Animals; D020013:Calcium Signaling; D025802:Electric Capacitance; D005089:Exocytosis; D006801:Humans; D008564:Membrane Potentials; D018408:Patch-Clamp Techniques; D022142:Secretory Vesicles / Q000378:metabolism; D059010:Single-Cell Analysis