Synapto-pHluorin

Synapto-pHluorin is a genetically encoded optical indicator of vesicle release and recycling. It is used in neuroscience to study transmitter release. It consists of a pH-sensitive form of green fluorescent protein (GFP) fused to the luminal side of a vesicle-associated membrane protein (VAMP). At the acidic pH inside transmitter vesicles, synapto-pHluorin is non-fluorescent (quenched). When vesicles get released, synapto-pHluorin is exposed to the neutral extracellular space and the presynaptic terminal becomes brightly fluorescent. Following endocytosis, vesicles become re-acidified and the cycle can start again. Chemical alkalinization of all vesicles is often used for normalization of the synapto-pHluorin signals. Synapto-pHluorin sometimes consists of yellow fluorescent protein (YFP) to monitor the cytoplasm because its pKa is higher than GFP (7.1 versus 6.0).[1]

History

Synapto-pHluorin was invented by Gero Miesenböck in 1998.[2] In 2006, an improved version was published, using synaptophysin to target the GFP to vesicles.[3] In 2013, a two-color release sensor (ratio-sypHy) was introduced to determine the size of the recycling pool at individual synapses.[4]

Applications

Synapto-pHluorin is mainly used by neurobiologists to study transmitter release and recycling at presynaptic terminals.[4] It has also been applied to the study of insulin secretion in beta cells of the pancreas.[5]

References
  1. Ashby MC, Ibaraki K, Henley JM (2004) It's green outside: tracking cell surface proteins with pH-sensitive GFP. TRENDS in Neuroscience 27(5):257-61
  2. Miesenböck G, De Angelis DA, Rothman JE (1998) Visualizing secretion and synaptic transmission with pH-sensitive green fluorescent proteins. Nature 394(6689):192-5.
  3. Granseth B, Odermatt B, Royle SJ, Lagnado L. (2006) Clathrin-mediated endocytosis is the dominant mechanism of vesicle retrieval at hippocampal synapses. Neuron 51(6):773-86
  4. Rose, Tobias; Schoenenberger, Philipp; Jezek, Karel; Oertner, Thomas G. (2013). "Developmental Refinement of Vesicle Cycling at Schaffer Collateral Synapses". Neuron. 77 (6): 1109–1121. doi:10.1016/j.neuron.2013.01.021. PMID 23522046.
  5. Tsuboi T, Rutter GA (2003) Multiple forms of "kiss-and-run" exocytosis revealed by evanescent wave microscopy. Curr Biol. 13(7):563-7.
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