Synapto-pHluorin

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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]

Contents

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]

Related Research Articles

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<span class="mw-page-title-main">Chemical synapse</span> Biological junctions through which neurons signals can be sent

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<span class="mw-page-title-main">Exocytosis</span> Active transport and bulk transport in which a cell transports molecules out of the cell

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<span class="mw-page-title-main">AMPA receptor</span> Transmembrane protein family

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<span class="mw-page-title-main">Gero Miesenböck</span>

Gero Andreas Miesenböck is an Austrian scientist. He is currently Waynflete Professor of Physiology and Director of the Centre for Neural Circuits and Behaviour (CNCB) at the University of Oxford and a fellow of Magdalen College, Oxford.

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<span class="mw-page-title-main">Axon terminal</span>

Axon terminals are distal terminations of the telodendria (branches) of an axon. An axon, also called a nerve fiber, is a long, slender projection of a nerve cell, or neuron, that conducts electrical impulses called action potentials away from the neuron's cell body, or soma, in order to transmit those impulses to other neurons, muscle cells or glands.

The ribbon synapse is a type of neuronal synapse characterized by the presence of an electron-dense structure, the synaptic ribbon, that holds vesicles close to the active zone. It is characterized by a tight vesicle-calcium channel coupling that promotes rapid neurotransmitter release and sustained signal transmission. Ribbon synapses undergo a cycle of exocytosis and endocytosis in response to graded changes of membrane potential. It has been proposed that most ribbon synapses undergo a special type of exocytosis based on coordinated multivesicular release. This interpretation has recently been questioned at the inner hair cell ribbon synapse, where it has been instead proposed that exocytosis is described by uniquantal release shaped by a flickering vesicle fusion pore.

<span class="mw-page-title-main">Active zone</span>

The active zone or synaptic active zone is a term first used by Couteaux and Pecot-Dechavassinein in 1970 to define the site of neurotransmitter release. Two neurons make near contact through structures called synapses allowing them to communicate with each other. As shown in the adjacent diagram, a synapse consists of the presynaptic bouton of one neuron which stores vesicles containing neurotransmitter, and a second, postsynaptic neuron which bears receptors for the neurotransmitter, together with a gap between the two called the synaptic cleft. When an action potential reaches the presynaptic bouton, the contents of the vesicles are released into the synaptic cleft and the released neurotransmitter travels across the cleft to the postsynaptic neuron and activates the receptors on the postsynaptic membrane.

<span class="mw-page-title-main">Synaptic fatigue</span>

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Kiss-and-run fusion is a type of synaptic vesicle release where the vesicle opens and closes transiently. In this form of exocytosis, the vesicle docks and transiently fuses at the presynaptic membrane and releases its neurotransmitters across the synapse, after which the vesicle can then be reused.

Neurotransmitters are released into a synapse in packaged vesicles called quanta. One quantum generates what is known as a miniature end plate potential (MEPP) which is the smallest amount of stimulation that one neuron can send to another neuron. Quantal release is the mechanism by which most traditional endogenous neurotransmitters are transmitted throughout the body. The aggregate sum of many MEPPs is known as an end plate potential (EPP). A normal end plate potential usually causes the postsynaptic neuron to reach its threshold of excitation and elicit an action potential. Electrical synapses do not use quantal neurotransmitter release and instead use gap junctions between neurons to send current flows between neurons. The goal of any synapse is to produce either an excitatory postsynaptic potential (EPSP) or an inhibitory postsynaptic potential (IPSP), which generate or repress the expression, respectively, of an action potential in the postsynaptic neuron. It is estimated that an action potential will trigger the release of approximately 20% of an axon terminal's neurotransmitter load.

References

  1. Ashby, Michael C.; Ibaraki, Kyoko; Henley, Jeremy M. (May 2004). "It's green outside: tracking cell surface proteins with pH-sensitive GFP". Trends in Neurosciences. 27 (5): 257–261. doi:10.1016/j.tins.2004.03.010. PMID   15111007. S2CID   23124974.
  2. Miesenböck, Gero; De Angelis, Dino A.; Rothman, James E. (July 1998). "Visualizing secretion and synaptic transmission with pH-sensitive green fluorescent proteins". Nature. 394 (6689): 192–195. doi:10.1038/28190. ISSN   1476-4687. PMID   9671304. S2CID   4320849.
  3. Granseth, Björn; Odermatt, Benjamin; Royle, Stephen J.; Lagnado, Leon (September 2006). "Clathrin-Mediated Endocytosis Is the Dominant Mechanism of Vesicle Retrieval at Hippocampal Synapses". Neuron. 51 (6): 773–786. doi: 10.1016/j.neuron.2006.08.029 . PMID   16982422.
  4. 1 2 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, Takashi; Rutter, Guy A. (April 2003). "Multiple Forms of "Kiss-and-Run" Exocytosis Revealed by Evanescent Wave Microscopy". Current Biology. 13 (7): 563–567. doi: 10.1016/S0960-9822(03)00176-3 . PMID   12676086.