Abstract
Neurotransmitter release is triggered by an increase in the cytosolic Ca2+ concentration ([Ca2+]i), but it is unknown whether the Ca2+-sensitivity of vesicle fusion is modulated during synaptic plasticity. We investigated whether the potentiation of neurotransmitter release by phorbol esters1,2,3, which target presynaptic protein kinase C (PKC)/munc-13 signalling cascades4,5,6, exerts a direct effect on the Ca2+-sensitivity of vesicle fusion. Using direct presynaptic Ca2+-manipulation and Ca2+ uncaging at a giant presynaptic terminal, the calyx of Held, we show that phorbol esters potentiate transmitter release by increasing the apparent Ca2+-sensitivity of vesicle fusion. Phorbol esters potentiate Ca2+-evoked release as well as the spontaneous release rate. We explain both effects by an increased fusion ‘willingness’ in a new allosteric model of Ca2+-activation of vesicle fusion. In agreement with an allosteric mechanism, we observe that the classically high Ca2+ cooperativity in triggering vesicle fusion (∼ 4) is gradually reduced below 3 µM [Ca2+]i, reaching a value of <1 at basal [Ca2+]i. Our data indicate that spontaneous transmitter release close to resting [Ca2+]i is a consequence of an intrinsic property of the molecular machinery7,8 that mediates synaptic vesicle fusion.
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Acknowledgements
We thank E. Neher for discussions, and P. Ascher, F. Felmy, E. Neher, T. Sakaba and M. Wölfel for comments on the manuscript. This work was supported by the Deutsche Forschungsgemeinschaft (DFG). R.S. is a Heisenberg fellow of the DFG.
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This file contains Supplementary Figures S1 and S2 and accompanying legends. Supplementary Figure S1 shows that EPSCs evoked by Ca2+-uncaging, and by brief presynaptic depolarizations are potentiated with a similar time-course by 1 µM PDBu, but the potentiation is transient under conditions of presynaptic whole-cell recording. Supplementary Figure S2 shows that EPSCs evoked by brief, and by prolonged presynaptic depolarizations are potentiated differentially by 1 µM PDBu, suggesting that the size of a readily-releasable vesicle pool is unchanged by PDBu. Furthermore, Ca2+-currents evoked by brief presynaptic depolarizations are slightly (∼15 %) increased by 1 µM PDBu. (DOC 1144 kb)
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Lou, X., Scheuss, V. & Schneggenburger, R. Allosteric modulation of the presynaptic Ca2+ sensor for vesicle fusion. Nature 435, 497–501 (2005). https://doi.org/10.1038/nature03568
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DOI: https://doi.org/10.1038/nature03568
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