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Fast vesicle replenishment allows indefatigable signalling at the first auditory synapse


Ribbon-type synapses in inner hair cells of the mammalian cochlea encode the complexity of auditory signals by fast and tonic release through fusion of neurotransmitter-containing vesicles. At any instant, only about 100 vesicles are tethered to the synaptic ribbon, and about 14 of these are docked to the plasma membrane1,2, constituting the readily releasable pool3. Although this pool contains about the same number of vesicles as that of conventional synapses4,5, ribbon release sites operate at rates of about two orders of magnitude higher3,6,7 and with submillisecond precision8,9,10,11. How these sites replenish their vesicles so efficiently remains unclear3,12,13. We show here, using two-photon imaging of single release sites in the intact cochlea, that preformed vesicles derived from cytoplasmic vesicle-generating compartments14 participate in fast release and replenishment. Vesicles were released at a maximal initial rate of 3 per millisecond during a depolarizing pulse, and were replenished at a rate of 1.9 per millisecond. We propose that such rapid resupply of vesicles enables temporally precise and sustained release rates. This may explain how the first auditory synapse can encode with indefatigable precision without having to rely on the slow, local endocytic vesicle cycle7.

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Figure 1: Experimental design.
Figure 2: Imaging individual ribbon release sites.
Figure 3: Kinetics of release and recovery at individual release sites.
Figure 4: Quantification of vesicular release and recovery.


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This work was supported by the MRC, the Wellcome Trust and a BBSRC Imaging grant to C.D.R. We thank U. Egert for his help with part of the image analysis.

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Correspondence to Claudius B. Griesinger or Jonathan F. Ashmore.

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Supplementary information

Supplementary Figure S1

Pharmacology of the calcium current in adult IHCs. The currents showed the properties of a Cav1.3 current with the α1D subunit, that is, non-inactivating L-type current. L-type blockers abolished or reduced stimulus-induced destaining of synaptic ribbons in IHCs in situ indicating that destaining represents vesicular release. (PDF 371 kb)

Supplementary Figure S2

Additional examples of synaptic release from ribbon release sites and accretion of labelled vesicles from cytoplasmic pools at release sites after stimulation. (PDF 1334 kb)

Supplementary Video S1

The video sequence shows increase of fluorescence derived from cytoplasm at a single release site following stimulation of the cell with a 50 s long stimulus train at 40 Hz. The fluorescence increase is interpreted as vesicle accretion from preformed labelled cytoplasmic pools. (AVI 17761 kb)

Supplementary Legends

Supplementary Legends to accompany the Supplementary Figures S1 and S2, and Supplementary Video S1 (DOC 23 kb)


Erratum regarding incomplete Supplementary Information uploading to Nature's website for this paper. This correction was made on 03 June 2005. (DOC 19 kb)

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Griesinger, C., Richards, C. & Ashmore, J. Fast vesicle replenishment allows indefatigable signalling at the first auditory synapse. Nature 435, 212–215 (2005).

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