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PMID: 20837642 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't

Calcium concentration jumps reveal dynamic ion selectivity of calcium-activated chloride currents in mouse olfactory sensory neurons and TMEM16b-transfected HEK 293T cells.

The Journal of physiology ·Vol. 588 ·No. Pt 21 ·2010-11-01 ·Pages 4189-204

Sagheddu C, Boccaccio A, Dibattista M, Montani G, Tirindelli R, Menini A

Abstract

Ca(2+)-activated Cl(-) channels play relevant roles in several physiological processes, including olfactory transduction, but their molecular identity is still unclear. Recent evidence suggests that members of the transmembrane 16 (TMEM16, also named anoctamin) family form Ca(2+)-activated Cl(-) channels in several cell types. In vertebrate olfactory transduction, TMEM16b/anoctamin2 has been proposed as the major molecular component of Ca(2+)-activated Cl(-) channels. However, a comparison of the functional properties in the whole-cell configuration between the native and the candidate channel has not yet been performed. In this study, we have used the whole-cell voltage-clamp technique to measure functional properties of the native channel in mouse isolated olfactory sensory neurons and compare them with those of mouse TMEM16b/anoctamin2 expressed in HEK 293T cells. We directly activated channels by rapid and reproducible intracellular Ca(2+) concentration jumps obtained from photorelease of caged Ca(2+) and determined extracellular blocking properties and anion selectivity of the channels. We found that the Cl(-) channel blockers niflumic acid, 5-nitro-2-(3-phenylpropylamino)benzoic acid (NPPB) and DIDS applied at the extracellular side of the membrane caused a similar inhibition of the two currents. Anion selectivity measured exchanging external ions and revealed that, in both types of currents, the reversal potential for some anions was time dependent. Furthermore, we confirmed by immunohistochemistry that TMEM16b/anoctamin2 largely co-localized with adenylyl cyclase III at the surface of the olfactory epithelium. Therefore, we conclude that the measured electrophysiological properties in the whole-cell configuration are largely similar, and further indicate that TMEM16b/anoctamin2 is likely to be a major subunit of the native olfactory Ca(2+)-activated Cl(-) current.

MeSH Terms
4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid/pharmacology Adenylyl Cyclases/metabolism Animals Anoctamins Calcium/metabolism Chloride Channels/antagonists & inhibitors,drug effects,genetics,metabolism HEK293 Cells Humans Kidney/cytology,metabolism Mice Mice, Inbred Strains Models, Animal Niflumic Acid/pharmacology Nitrobenzoates/pharmacology Olfactory Nerve/cytology,metabolism Patch-Clamp Techniques Sensory Receptor Cells/cytology,metabolism Transfection
Chemicals
ANO2 protein, mouse Anoctamins Chloride Channels Nitrobenzoates 5-nitro-2-(3-phenylpropylamino)benzoic acid Niflumic Acid Adenylyl Cyclases adenylate cyclase 3 4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid Calcium
Authors & Affiliations
6 authors, click to expand affiliations / ORCID
Sagheddu Claudia
International School for Advanced Studies, Scuola Internazionale Superiore di Studi Avanzati, SISSA, Trieste, Italy.
Boccaccio Anna
Dibattista Michele
Montani Giorgia
Tirindelli Roberto
Menini Anna
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Article Info
Journal
The Journal of physiology
Abbr.
J Physiol
ISSN
1469-7793
DOI
10.1113/jphysiol.2010.194407
Published
2010-11-01
Epub
2010-00-13
Pages
4189-204
Language
English
Region
England
NLM ID
0266262
PMCID
PMC3002450
Subset
IM
External Links
PubMed source DOI: 10.1113/jphysiol.2010.194407 PMC Full Text View journal details
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