Visualizing secretion and synaptic transmission with pH-sensitive green fluorescent proteins

Gero Miesenböck, Dino A. De Angelis and James E. Rothman
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Gero Miesenböck: Cellular Biochemistry and Biophysics Program, Memorial Sloan-Kettering Cancer Center
Dino A. De Angelis: Cellular Biochemistry and Biophysics Program, Memorial Sloan-Kettering Cancer Center
James E. Rothman: Cellular Biochemistry and Biophysics Program, Memorial Sloan-Kettering Cancer Center

Nature, 1998, vol. 394, issue 6689, 192-195

Abstract: Abstract In neural systems, information is often carried by ensembles of cells rather than by individual units. Optical indicators1 provide a powerful means to reveal such distributed activity, particularly when protein-based and encodable in DNA2,3,4: encodable probes can be introduced into cells, tissues, or transgenic organisms by genetic manipulation, selectively expressed in anatomically or functionally defined groups of cells, and, ideally, recorded in situ, without a requirement for exogenous cofactors. Here we describe sensors for secretion and neurotransmission that fulfil these criteria. We have developed pH-sensitive mutants of green fluorescent protein (‘pHluorins’) by structure-directed combinatorial mutagenesis, with the aim of exploiting the acidic pH inside secretory vesicles5,6 to monitor vesicle exocytosis and recycling. When linked to a vesicle membrane protein, pHluorins were sorted to secretory and synaptic vesicles and reported transmission at individual synaptic boutons, as well as secretion and fusion pore ‘flicker’ of single secretory granules.

Date: 1998
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DOI: 10.1038/28190

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