title: Conditional knockout of TMEM16A/anoctamin1 abolishes the calcium-activated chloride current in mouse vomeronasal sensory neurons.
abstract: Pheromones are substances released from animals that, when detected by the vomeronasal organ of other individuals of the same species, affect their physiology and behavior. Pheromone binding to receptors on microvilli on the dendritic knobs of vomeronasal sensory neurons activates a second messenger cascade to produce an increase in intracellular Ca(2+) concentration. Here, we used whole-cell and inside-out patch-clamp analysis to provide a functional characterization of currents activated by Ca(2+) in isolated mouse vomeronasal sensory neurons in the absence of intracellular K(+). In whole-cell recordings, the average current in 1.5 µM Ca(2+) and symmetrical Cl(-) was -382 pA at -100 mV. Ion substitution experiments and partial blockade by commonly used Cl(-) channel blockers indicated that Ca(2+) activates mainly anionic currents in these neurons. Recordings from inside-out patches from dendritic knobs of mouse vomeronasal sensory neurons confirmed the presence of Ca(2+)-activated Cl(-) channels in the knobs and/or microvilli. We compared the electrophysiological properties of the native currents with those mediated by heterologously expressed TMEM16A/anoctamin1 or TMEM16B/anoctamin2 Ca(2+)-activated Cl(-) channels, which are coexpressed in microvilli of mouse vomeronasal sensory neurons, and found a closer resemblance to those of TMEM16A. We used the Cre-loxP system to selectively knock out TMEM16A in cells expressing the olfactory marker protein, which is found in mature vomeronasal sensory neurons. Immunohistochemistry confirmed the specific ablation of TMEM16A in vomeronasal neurons. Ca(2+)-activated currents were abolished in vomeronasal sensory neurons of TMEM16A conditional knockout mice, demonstrating that TMEM16A is an essential component of Ca(2+)-activated Cl(-) currents in mouse vomeronasal sensory neurons.
authors.affiliation: Neurobiology Group, SISSA, Scuola Internazionale Superiore di Studi Avanzati, 34136 Trieste, Italy.
authors.forename: Asma
authors.initials: A
authors.lastname: Amjad
authors.affiliation: Neurobiology Group, SISSA, Scuola Internazionale Superiore di Studi Avanzati, 34136 Trieste, Italy.
authors.forename: Andres
authors.initials: A
authors.lastname: Hernandez-Clavijo
authors.affiliation: Neurobiology Group, SISSA, Scuola Internazionale Superiore di Studi Avanzati, 34136 Trieste, Italy.
authors.forename: Simone
authors.initials: S
authors.lastname: Pifferi
authors.affiliation: Neurobiology Group, SISSA, Scuola Internazionale Superiore di Studi Avanzati, 34136 Trieste, Italy.
authors.forename: Devendra Kumar
authors.initials: DK
authors.lastname: Maurya
authors.affiliation: Istituto di Biofisica, National Research Council, 16149 Genova, Italy.
authors.forename: Anna
authors.initials: A
authors.lastname: Boccaccio
authors.affiliation: Neurobiology Group, SISSA, Scuola Internazionale Superiore di Studi Avanzati, 34136 Trieste, Italy.
authors.forename: Jessica
authors.initials: J
authors.lastname: Franzot
authors.affiliation: Department of Anatomy, University of California, San Francisco, School of Medicine, San Francisco, CA 94143.
authors.forename: Jason
authors.initials: J
authors.lastname: Rock
authors.affiliation: Neurobiology Group, SISSA, Scuola Internazionale Superiore di Studi Avanzati, 34136 Trieste, Italy menini@sissa.it.
authors.forename: Anna
authors.initials: A
authors.lastname: Menini
mesh_terms: D000200:Action Potentials; D000818:Animals; D000075369:Anoctamin-1; D002118:Calcium / Q000378:metabolism; D002478:Cells, Cultured; D018118:Chloride Channels / Q000235:genetics / Q000378:metabolism*; D017353:Gene Deletion; D051379:Mice; D011984:Sensory Receptor Cells / Q000378:metabolism* / Q000502:physiology; D019147:Vomeronasal Organ / Q000166:cytology / Q000378:metabolism*
chemical_list: C531181:ANO1 protein, mouse; D000075369:Anoctamin-1; D018118:Chloride Channels; D002118:Calcium