Chemical synapses without synaptic vesicles: Purinergic neurotransmission through a CALHM1 channel-mitochondrial signaling complex
Tasting with ATP
The savory taste of umami, the sweetness of sugar, and the bitterness of quinine are transduced by type II taste cells. Unlike most other receptor cells, type II taste cells release their neurotransmitter, ATP, through voltage-gated CALHM1 channels instead of neurotransmitter-containing vesicles. Romanov et al. found that the source of ATP was unusual, large mitochondria, closely opposed to clusters of CALHM1 channels within the plasma membrane of type II taste cells. This arrangement enables an alternate method of chemical neurotransmission that does not rely on vesicles.
Abstract
Conventional chemical synapses in the nervous system involve a presynaptic accumulation of neurotransmitter-containing vesicles, which fuse with the plasma membrane to release neurotransmitters that activate postsynaptic receptors. In taste buds, type II receptor cells do not have conventional synaptic features but nonetheless show regulated release of their afferent neurotransmitter, ATP, through a large-pore, voltage-gated channel, CALHM1. Immunohistochemistry revealed that CALHM1 was localized to points of contact between the receptor cells and sensory nerve fibers. Ultrastructural and super-resolution light microscopy showed that the CALHM1 channels were consistently associated with distinctive, large (1- to 2-μm) mitochondria spaced 20 to 40 nm from the presynaptic membrane. Pharmacological disruption of the mitochondrial respiratory chain limited the ability of taste cells to release ATP, suggesting that the immediate source of released ATP was the mitochondrion rather than a cytoplasmic pool of ATP. These large mitochondria may serve as both a reservoir of releasable ATP and the site of synthesis. The juxtaposition of the large mitochondria to areas of membrane displaying CALHM1 also defines a restricted compartment that limits the influx of Ca2+ upon opening of the nonselective CALHM1 channels. These findings reveal a distinctive organelle signature and functional organization for regulated, focal release of purinergic signals in the absence of synaptic vesicles.
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Supplementary Material
Summary
Fig. S1. Validation of the CALHM1 antibody.
Fig. S2. Localization of CALHM1 immunoreactivity in taste buds.
Fig. S3. Calcium responses in sensor cells.
Fig. S4. Effect of CBX on Mito-ID fluorescence in living taste cells.
Table S1. Amplitudes of calcium responses in type II taste cells from Fig. 3B.
Movie S1. 3D visualization of the relationship between a type II taste cell and the innervating nerve fiber shown in Fig. 1A.
Movie S2. 3D visualization of mitochondria and taste cell shown in Fig. 1B.
Movie S3. Relationship of atypical and typical mitochondria.
Movie S4. Tubular cristae in atypical mitochondria.
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Published In
Science Signaling
Volume 11 | Issue 529
May 2018
May 2018
Copyright
Copyright © 2018 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.
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Submission history
Received: 23 June 2017
Accepted: 12 April 2018
Acknowledgments
We thank N. Shultz and M. Li for assistance with immunohistochemistry, R. Russell and R. K. Johnson for segmentations and reconstructions from sbfSEM data, and S. Fore (Carl Zeiss Microscopy, LLC) for assistance in acquiring the Airyscan images. We also appreciate the following for review of this manuscript at various stages in its preparation and finalization: F. A. Chaudhry (University of Oslo, Oslo, Norway), K. Beam and S. C. Kinnamon (University of Colorado School of Medicine), and E. Liman (University of Southern California). Funding: S.S.K. was supported by the Russian Academy of Sciences (Program #7) and the Russian Foundation for Basic Research (grant no. 13-04-40082). I.A. was supported by the Swedish Research Council, the Bertil Hallsten Research Foundation, the Jeassons Foundation, the Strategic Regeneration Foundation, and the Knut and Alice Wallenberg Foundation (CLICK). T.H. was funded by the Swedish Research Council, the Swedish Brain Foundation, the Novo Nordisk Foundation, the Petrus and Augusta Hedlunds Foundation, a European Research Council (ERC) Advanced Grant (“Secret-Cells,” ERC-AdG-2015-695136), and the European Commission Integrated Project “PAINCAGE.” R.A.R. was a European Molecular Biology Organization (EMBO) long-term research fellow (ALTF 596-2014) co-funded by the European Commission FP7 (Marie Curie Actions, EMBOCOFUND2012, GA-2012-600394). R.A.R. was also supported by the Ministry of Education and Science of the Russian Federation (agreement no. 14.575.21.0074 with Immanuel Kant Baltic Federal University; project leader, V. Kasymov). This study was supported by grants from the National Institute for Deafness and Communicative Disorders of the NIH (USA) (to T.E.F., R.S.L., B.H., R.Y., G.J.K., and J.C.K.), grants 1R21DC013186 and R01DC014728 (to T.E.F.), and grant P30DC004657 to D. Restrepo (University of Colorado School of Medicine). P.M. was supported by a grant from the National Institute on Aging of the NIH (R01AG042508). Author contributions: S.S.K., T.E.F., and R.A.R. designed the study. R.A.R., T.E.F., and R.Y. performed immunostaining. R.A.R. performed calcium imaging and electrophysiological experiments. R.A.R., O.A.R., and M.F.B. performed experiments with cellular ATP and ATP/ADP sensors. H.Z. and P.M. developed and characterized the antibody against CALHM1. V.V.R., T.E.F., B.H., R.S.L., R.Y., L.E.S., G.J.K., A.L., and J.C.K. performed EM experiments and made 3D reconstructions. G.D.C. and O.A.R. performed experiments with effects of CBX. R.A.R., T.E.F., I.A., T.H., P.M., and S.S.K. analyzed data and contributed to writing the paper. All authors commented on the manuscript and approved its submission. Competing interests: The authors declare that they have no competing interests. Data and materials availability: All the data required to evaluate the conclusions in the paper are present in the paper or the Supplementary Materials. The Calmh1 knockout mice require a material transfer agreement from P. Marambaud (The Feinstein Institute for Medical Research).
Authors
Funding Information
National Institute on Aging: R01AG042508
Russian Foundation for Basic Research: 13-04-40082
European Research Council: ERC-AdG-2015-695136
Swedish Brain Foundation
EMBO: ALTF 596-2014
Ministry of Education and Science of the Russian Federation: 14.575.21.0074
Swedish Brain Foundation
Knut and Alice Wallenberg foundation
Bertil Hallsten Research Foundation
Jeassons Foundation
Petrus and Augusta Hedlunds Foundation
European Commission Integrated Project: GA-2012-600394
Novo Nordisk Foundation
Novo Nordisk Foundation
European Commission Integrated Project
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