A cyclic nucleotide-gated conductance in olfactory receptor cilia

Nakamura, Tadashi; Gold, Geoffrey H.

doi:10.1038/325442a0

Cited by 1,079 publications

(607 citation statements)

References 26 publications

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“…These observations suggest that in type I cells 02 interacts with the K+ channels either directly or through a site closely associated with them. In this respect type I cells differ from olfactory or taste chemoreceptor cells in which natural stimuli modulate ion channels through the action of intracellular cyclic nucleotides (15)(16)(17) .r. T"r----.…”

Section: Resultsmentioning

confidence: 99%

Single K+ channels in membrane patches of arterial chemoreceptor cells are modulated by O2 tension.

Ganfornina

López‐Barneo

1991

Proc. Natl. Acad. Sci. U.S.A.

149

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Type I cells of the carotid body are known to participate in the detection of 02 tension in arterial blood but the primary chemotransduction mechanisms are not well understood. Here we report the existence in excised membrane patches of type I cells of a single K+ channel type modulated by changes in P02. Open probability of the O2-sensitive K+ channel reversibly decreased by at least 50% on exposure to hypoxia but single-channel conductance (-20 pS) was unaltered. In the range between 70 and 150 mmHg (1 mmHg = 133 Pa) the decrease of single-channel open probability was proportional to the Po2 measured in the vicinity of the membrane patch. The inhibition of K+ channel activity by low P02 was independent of the presence of non-hydrolyzable guanine triphosphate analogues at the internal face of the membrane. The results indicate that the 02 sensor of type I cells is in the plasma membrane and suggest that environmental 02 interacts directly with the K+ channels.Type I, or glomus, cells of the carotid body have been considered for decades to be responsible for the detection of oxygen tension (Po2) in the arterial blood but the mechanisms involved in the process of chemotransduction have remained obscure (1, 2). However, it has been recently discovered that type I cells from adult rabbits can generate action potentials and that they have a voltage-dependent K+ current selectively and reversibly attenuated by lowering Po2 (3)(4)(5)(6)(7)(8). Inhibition of this K+ current under hypoxic conditions could produce an increase in the firing frequency ofchemoreceptor cells, leading to Ca2' influx, enhanced transmitter release, and activation of the afferent fibers of the sinus nerve (1, 2, 6). The primary site for 02 detection is, however, unknown. Exposure to cyanide or to extreme hypoxia (<40 mmHg; 1 mmHg = 133 Pa) induces an increase of [Ca2+]j in type I cells, possibly due to Ca2' release from mitochondria, and a subsequent activation of a Ca2+-dependent K+ current (9, 10). Therefore it has been argued that the modification ofthe K+ current by lowering Po2 might be a secondary phenomenon rather than an initial step in the process of chemotransduction. Now we report the identification of a single K+ channel type that fully accounts for the properties ofthe macroscopic 02-sensitive K+ current.Furthermore, we show that in excised membrane patches the activity of these K+ channels is reversibly modulated by P02. Our results support the view that the K+ channels are directly regulated by 02 and strongly suggest that the 02 sensor of chemoreceptor cells is in, or associated with, the plasma membrane. This type of Ke channel regulation found in the carotid body may have an even broader functional interest because it could also be involved in physiological responses to hypoxia in other tissues. METHODS Experiments were performed on enzymatically dispersed type I cells isolated from rabbit carotid bodies. The methods followed in cell dissociation and culture were the same as previously described (3, 4). Cells were plate...

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Section: Resultsmentioning

confidence: 99%

Single K+ channels in membrane patches of arterial chemoreceptor cells are modulated by O2 tension.

Ganfornina

López‐Barneo

1991

Proc. Natl. Acad. Sci. U.S.A.

149

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“…Among others, the cNMP superfamily includes a variety of bacterial regulators, such as the catabolite gene activator protein (CAP) (Eron et al 1971;Weber and Steitz 1987), cyclic nucleotide-gated ion channels (Nakamura and Gold 1987;Ludwig et al 1990), guanine nucleotide exchange factors (Kawasaki et al 1998), and the regulators of PKA and PKG. The distinctive feature of the members of the superfamily is the presence of a common structural domain of about 120 residues (Shabb and Corbin 1992) capable of binding cyclic nucleotides.…”

Section: Introductionmentioning

confidence: 99%

Classification and Phylogenetic Analysis of the cAMP-Dependent Protein Kinase Regulatory Subunit Family

2002

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Abstract. The members of the PKA regulatory subunit family (PKA-R family) were analyzed by multiple sequence alignment and clustering based on phylogenetic tree construction. According to the phylogenetic trees generated from multiple sequence alignment of the complete sequences, the PKA-R family was divided into four subfamilies (types I to IV). Members of each subfamily were exclusively from animals (types I and II), fungi (type III), and alveolates (type IV). Application of the same methodology to the cAMP-binding domains, and subsequently to the region delimited by ␤-strands 6 and 7 of the crystal structures of bovine RI␣ and rat RII␤ (the phosphate-binding cassette; PBC), proved that this highly conserved region was enough to classify unequivocally the members of the PKA-R family. A single signature sequence, F-G-E-[LIV]-A-L-[LIMV]-x(3)-[PV]-R-[ANQV]-A, corresponding to the PBC was identified which is characteristic of the PKA-R family and is sufficient to distinguish it from other members of the cyclic nucleotide-binding protein superfamily. Specific determinants for the A and B domains of each Rsubunit type were also identified. Conserved residues defining the signature motif are important for interaction with cAMP or for positioning the residues that directly interact with cAMP. Conversely, residues that define subfamilies or domain types are not conserved and are mostly located on the loop that connects ␣-helix BЈ and ␤ strand 7.

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“…In the MOE, it colocalized with the cilia marker protein 4 acetylated tubulin (Fig. 2b) and the Cnga2 subunit of the olfactory CNG channel 3,25,26 ( Fig. 2c).…”

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confidence: 96%

Ca2+-activated Cl− currents are dispensable for olfaction

et al. 2011

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Graduate student at the Freie Universität Berlin Olfaction in mammals involves the binding of odorants to a subset of the large number of different G protein-coupled receptors that are present in the cilia of olfactory sensory neurons (OSNs) 1 . These neurons are predominantly found in the MOE, which senses and discriminates myriad volatile compounds. In the mouse, the nose contains additional, apparently more specialized olfactory organs, such the septal organ of Masera and the VNO 2 . Sensory neurons in the VNO are morphologically distinct and use odorant receptors and signal transduction cascades that differ from those found in the MOE.In the canonical OSN signal transduction pathway, odorant binding to the receptor locally increases cytosolic cAMP through activation of the olfactory G protein G olf and adenylate cyclase type III 1,2 . cAMP then opens heteromeric cyclic nucleotide-gated (CNG) cation channels 3 . The resulting influx of Na + and Ca 2+ depolarizes the plasma membrane and raises the cytosolic Ca 2+ concentration ([Ca 2+ ] i ), thereby activating Ca 2+ -activated Cl -channels 1,4-10 . All these components of the signal transduction cascade are localized to sensory cilia, which are embedded in the mucus covering the MOE. These cilia provide a large interaction surface for odorants, and the cilia's small diameters facilitate large local increases in cytosolic cAMP and [Ca 2+ ].There has been broad consensus that Ca 2+ -activated Cl -channels powerfully amplify olfactory signal transduction 1,4-10 . These channels are thought to mediate an outward flow of Cl -, which generates a depolarizing current that, in rodents, is five-to tenfold larger than currents through CNG channels 8,11,12 . A prerequisite for Cl -efflux 3 is an inside-out electrochemical Cl --gradient. It is believed that cytosolic chloride concentration of OSNs is raised by the Na + K + 2Cl -co-transporter Nkcc1 9,10,13 and that [Cl -] is low in the mucus surrounding the cilia 14,15 . However, mucosal ion concentrations are difficult to measure in vivo, and Nkcc1 knockout mice display attenuated electro-olfactograms (EOGs) 11,16 , but normal olfactory sensitivity 17 .To investigate the role of Ca 2+ -activated Cl -channels in olfaction, we disrupted Ano2 in mice. Ano2 is a member of the Anoctamin (Tmem16) gene family, which encodes several Ca 2+ -activated Cl -channels [18][19][20] . Agreeing with recent results 13,21-23 , our knockout-controlled immunolabeling showed Ano2 expression in cilia of OSNs in the MOE, in microvilli of VNO sensory neurons and in synapses of photoreceptors. Additionally, we found Ano2 in the olfactory bulb. Ano1 expression overlapped with Ano2 in the VNO and the retina, but not in the MOE. Patch-clamp analysis showed that Ca 2+ -activated Cl -currents were undetectable in Ano2 -/-OSNs.Unexpectedly, however, EOGs of Ano2 -/-mice were reduced by only up to ~40%, and Ano2 -/-mice were able to smell normally. Our work calls for a revision of the current view that Ca 2+ -activated Cl -channels have a crucial role...

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A cyclic nucleotide-gated conductance in olfactory receptor cilia

Cited by 1,079 publications

References 26 publications

Single K+ channels in membrane patches of arterial chemoreceptor cells are modulated by O2 tension.

Single K+ channels in membrane patches of arterial chemoreceptor cells are modulated by O2 tension.

Classification and Phylogenetic Analysis of the cAMP-Dependent Protein Kinase Regulatory Subunit Family

Ca2+-activated Cl− currents are dispensable for olfaction

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