Ion Channels

Abstract

Alexander SPH, Mathie A, Peters JA (2008). Guide to Receptors and Channels (GRAC), 3rd edn. Br J Pharmacol 153 (Suppl. 2): S1–S209. Overview: Acid-sensing ion channels (ASICs, provisional nomenclature; see Wemmie et al., 2006; Lingueglia et al., 2007) are members of a Na+ channel superfamily that includes the epithelial Na channel (ENaC), the FMRF-amide activated channel (FaNaC) of invertebrates, the degenerins (DEG) of Caenorhabitis elegans, channels in Drosophila melanogaster and ‘orphan’ channels that include BLINaC (Sakai et al., 1999) and INaC (Schaefer et al. (2000). ASIC subunits contain two putative TM domains and assemble as homo- or hetero-trimers (Jasti et al., 2007) to form proton-gated, voltage-insensitive, Na+ permeable, channels. Splice variants of ASIC1 [provisionally termed ASIC1a (ASIC, ASICα, BNaC2α) (Waldmann et al. 1997a), ASIC1b (ASICβ, BNaC2β) (Chen et al., 1998) and ASIC1b2 (ASICβ2) (Ugawa et al., 2001); note that ASIC1a is also permeable to Ca2+] and ASIC2 [provisionally termed ASIC2a (MDEG1, BNaC1α, BNC1a) (Price et al., 1996; Waldmann et al., 1996; Garcia-Anoveros et al., 1997) and ASIC2b (MDEG2, BNaC1β) (Lingueglia et al., 1997)] have been cloned. Unlike ASIC2a (listed in table), heterologous expression of ASIC2b alone does not support H+-gated currents. A third member, ASIC3 (DRASIC, TNaC1) (Waldmann et al., 1997b), has been identified. Transcripts encoding a fourth mammalian member of the acid-sensing ion channel family (ASIC4/SPASIC) do not produce a proton-gated channel in heterologous expression systems (Akopian et al., 2000; Grunder et al., 2000), whereas one zebrafish orthologue (zASIC4.1) is functional as a homomer (Paukert et al., 2004) but a second (zASIC4.2) is not (Chen et al., 2007). ASIC channels are expressed in central and peripheral neurons including nociceptors where they participate in neuronal sensitivity to acidosis. The activation of ASIC1a within the brain contributes to neuronal injury caused by focal ischemia (Xiong et al., 2007). Further proposed roles for centrally and peripherally located ASICs are reviewed in Wemmie et al. (2006) and Lingueglia (2007). The relationship of the cloned ASICs to endogenously expressed proton-gated ion channels is becoming established (Escoubas et al., 2000; Sutherland et al., 2001; Wemmie et al., 2002, 2003, 2006; Lingueglia et al., 2006; 2007, Diochot et al., 2004, 2007). Heterologously expressed heteromultimers form ion channels with altered kinetics, ion selectivity, pH- sensitivity and sensitivity to blockers (Lingueglia et al., 1997; Babinski et al., 2000, Escoubas et al., 2000) that resemble some of the native proton activated currents recorded from neurones. Psalmotoxin 1 (PcTx1) inhibits ASIC1a by modifying activation and desensitization by H+, but has little effect upon ASIC1b, ASIC2a, ASIC3, or ASIC1a expressed as a heteromultimer with either ASIC2a, or ASIC3 (Escoubas et al., 2000; Diochot et al., 2007). Blockade of ASIC1a by PcTx1 results in the activation of the endogenous enkephalin pathway and has very potent analgesic effects in rodents (Mazzuca et al., 2007). APETx2 most potently blocks homomeric ASIC3 channels, but also ASIC2b + ASIC3, ASIC1b + ASIC3, and ASIC1a + ASIC3 heteromeric channels with IC50 values of 117 nM, 900 nM and 2 μM, respectively. APETx2 has no effect on ASIC1a, ASIC1b, ASIC2a, or ASIC2a + ASIC3 (Diochot et al., 2004, 2007). A-317567 blocks ASIC channels native to dorsal root ganglion neurones with an IC50 within the range 2-30 μM (Dube et al., 2005). The pEC50 values for proton activation of ASIC channels are influenced by numerous factors including extracellular di- and poly-valent ions, Zn2+, protein kinase C and serine proteases (Lingueglia et al., 2006). Rapid acidification is required for activation of ASIC1 and ASIC3 due to fast inactivation/desensitization. pEC50 values for H+-activation of either transient, or sustained, currents mediated by ASIC3 vary in the literature and may reflect species and/or methodological differences (Waldmann et al., 1997b; de Weille et al., 1998; Babinski et al., 1999). The transient and sustained current components mediated by rASIC3 are selective for Na+ (Waldmann et al., 1997b); for hASIC3 the transient component is Na+ selective (PNa/PK>10) whereas the sustained current appears non-selective (PNa/PK = 1.6) (de Weille et al., 1998; Babinski et al., 1999). Nonsteroidal anti-inflammatory drugs (NSAIDs) are direct blockers of ASIC currents within the therapeutic range of concentrations (reviewed by Voilley, 2004). ASIC1a is blocked by flurbiprofen and ibuprofen and currents mediated by ASIC3 are inhibited by salicylic acid, aspirin and diclofenac. Extracellular Zn2+ potentiates proton activation of homomeric and heteromeric channels incorporating ASIC2a, but not homomeric ASIC1a or ASIC3 channels (Baron et al., 2001). However, removal of contaminating Zn2+ by chealation reveals a high affinity block of homomeric ASIC1a and heteromeric ASIC1a + ASIC2 channels by Zn2+ indicating complex biphasic actions of the divalent (Chu et al., 2004). Ammonium activates ASIC channels (most likely ASIC1a) in midbrain dopaminergic neurones which may be relevant to neuronal disorders that are associated with hyperammonemia (Pidoplichko and Dani, 2006). The positive modulation of homomeric, heteromeric and native ASIC channels by the peptide FMRFamide and related substances, such as neuropeptides FF and SF, is reviewed in detail by Lingueglia et al. (2006). Inflammatory conditions and particular pro-inflammatory mediators induce overexpression of ASIC-encoding genes, enhance ASIC currents (Mamet et al., 2002), and in the case of arachidonic acid directly activate the channel (Smith et al., 2007). Abbreviations: A-317567, C-{6-[2-(1-Isopropyl-2-methyl-1,2,3,4-tetrahydro-isoquinolin-7-yl)-cyclopropyl]-naphthalen-2-yl}-methanediamine; EIPA, ethylisopropylamiloride; FMRFamide, Phe-Met-Arg-Phe-amide; Neuropeptide FF, Phe-Leu-Phe-Gln-Pro-Gln-Arg-Phe-amide; Neuropeptide SF, Ser-Leu-Ala-Pro-Gln-Arg-Phe-amide Diochot S, Salinas M, Baron A, Escoubas P, Lazdunski M (2007). Peptides inhibitors of acid-sensing ion channels. Toxicon 49: 271–284. Kress M, Waldmann R (2006). Acid sensing ionic channels. Curr Top Membr 57: 241–276. Krishtal O (2003). The ASICs: Signaling molecules? Modulators? Trends Neurosci 26: 477–483. Lingueglia E (2007). Acid-sensing ion channels in sensory perception. J Biol Chem 282: 17325–17329. Lingueglia E, Deval E, Lazdunski M (2006). FMRFamide-gated sodium channel and ASIC channels: a new class of ionotropic receptors for FMRFamide and related peptides. Peptides 27: 1138–1152. Reeh PW, Kress M (2001). Molecular physiology of proton transduction in nociceptors. Curr Opin Pharmacol 1: 45–51. Sutherland SP, Cook SP, McCleskey EW (2000). Chemical mediators of pain due to tissue damage and ischemia. Prog Brain Res 129: 21–38. Voilley N (2004). Acid-sensing ion channels (ASICs): new targets for the analgesic effects of non-steroid anti-inflammatory drugs (NSAIDs). Curr Drug Targets Inflamm Allerg 3: 71–79. Waldmann R (2001). Proton-gated cation channels-neuronal acid sensors in the central and peripheral nervous system. Adv Exp Med Biol 502: 293–304. Waldmann R, Lazdunski M (1998). H+-gated cation channels: neuronal acid sensors in the NaC/DEG family of ion channels. Curr Opin Neurobiol 8: 418–424. Wemmie JA, Price MP, Welsh MJ (2006). Acid-sensing ion channels: advances, questions and therapeutic opportunities. Trends Neurosci 29: 578–586. Xiong ZG, Chu XP, Simon RP (2007). Acid sensing ion channels-novel therapeutic targets for ischemic brain injury. Front Biosci 12: 1376–1386. Xiong ZG, Pignataro G, Li M, Chang SY, Simon RP (2007). Acid-sensing ion channels as pharmacological targets for neurodegenerative diseases. Curr Opin Pharmacol Oct 16 [E-pub ahead of print]. Xu TL, Xiong ZG (2007). Dynamic regulation of acid-sensing ion channels by extracellular and intracellular modulators. Curr Med Chem 14: 1753–1763. Akopian AN et al. (2000). Neuroreport 11: 2217–2222. Babinski K et al. (1999). J Neurochem 72: 51–57. Babinski K et al. (2000). J Biol Chem 37: 28519–28525. Baron A et al. (2001). J Biol Chem 276: 35361–35367. Chen C-C et al. (1998). Proc Natl Acad Sci USA 95: 10240–10245. Chen X et al. (2007). J Biol Chem 282: 30406–30416. Chu X-P et al. (2004). J Neurosci 24: 8678–8689. De Weille JR et al. (1998). FEBS Lett 433: 257–260. Diochot S et al. (2004). EMBO J 23: 1516–1525. Dube GR et al. (2005). Pain 117: 88–96. Escoubas P et al. (2000). J Biol Chem 275: 25116–25121. Garcia-Anoveros J et al. (1997). Proc Natl Acad Sci USA 94: 1459–1464. Grunder S et al. (2000). Neuroreport 11: 1607–1611. Jasti J et al. (2007). Nature 449: 316–323. Lingueglia E et al. (1997). J Biol Chem 272: 29778–29783. Mamet J et al. (2002). J Neurosci 22: 10662–10670. Mazzuca M et al. (2007). Nat Neurosci 10: 943–945. Paukert M et al. (2004). J Biol Chem 279: 18733–18791. Pidoplichko VI, Dani JA (2006). Proc Natl Acad Sci USA 103: 11376–11380. Price MP et al. (1996). J Biol Chem 271: 7879–7882. Sakai H et al. (1999). J Physiol (Lond) 519: 323–333. Schaefer L et al. (2000). FEBS Lett 471: 205–210. Smith ES et al. (2007). Neuroscience 145: 686–698. Sutherland SP et al. (2001). Proc Natl Acad Sci USA 98: 711–716. Ugawa S et al. (2001). Neuroreport 12: 2865–2869. Waldmann R et al. (1996). J Biol Chem 271: 10433–10436. Waldmann R et al. (1997a). Nature 386: 173–177. Waldmann R et al. (1997b). J Biol Chem 272: 20975–20978. Wemmie JA et al. (2002). Neuron 34: 463–477. Wemmie JA et al. (2003). J Neurosci 23: 5496–5502. We recommend that any citations to information in the Guide are presented in the following format: Alexander SPH, Mathie A, Peters JA (2008). Guide to Receptors and Channels (GRAC), 3rd edn. Br J Pharmacol 153 (Suppl. 2): S1-S209. Overview: Aquaporins and aquaglyceroporins are membrane channels that allow the permeation of water and certain other small solutes across the cell membrane. Since the isolation and cloning of the first aquaporin (AQP1) (Preston et al., 1992), 12 additional members of the family have been identified, although little is known about the functional properties of two of these (AQP11 (ENSG00000178301) and AQP12 (ENSG00000184945)). The other 11 aquaporins can be divided into two families (aquaporins and aquaglyceroporins) depending on whether they are permeable to glycerol (King et al., 2004). One or more members of this family of proteins have been found to be expressed in almost all tissues of the body. Individual AQP subunits have six transmembrane domains with an inverted symmetry between the first three and last three domains (Castle, 2005). Functional AQPs exist as tetramers but, unusually, each subunit contains a separate pore, so each channel has four pores. AQP6 is an intracellular channel permeable to anions as well as water (Yasui et al., 1999). Agre P (2006). The aquaporin water channels. Proc Am Thoroc Soc 3: 5–13. Agre P, King LS, Yasui M, Guggino WB, Ottersen OP, Fujiyoshi Y et al. (2002). Aquaporin water channels: from atomic structure to clinical medicine. J Physiol 542: 3–16. Amiry-Moghaddam M, Ottersen OP (2003). The molecular basis of water transport in the brain. Nat Rev Neurosci 4: 991–1001. Castle NA (2005). Aquaporins as targets for drug discovery. Drug Discov Today 10: 485–493. De Groot BL, Grubmuller H (2005). The dynamics and energetics of water permeation and proton exclusion in aquaporins. Curr Opin Struct Biol 15: 176–183. Frigeri A, Nicchia GP, Svelto M (2007). Aquaporins as targets for drug discovery. Curr Pharm Des 13: 2421–2427. Jeyaseelan K, Sepramaniam S, Armugam A, Wintour EM (2006). Aquaporins: a promising target for drug development. Expert Opin Ther Targets 10: 889–909. Kimelberg HK (2004). Water homeostasis in the brain: basic concepts. Neuroscience 129: 851–860. King KL, Kozono D, Agre P (2004). From structure to disease: the evolving tale of aquaporin biology. Nat Rev Mol Cell Biol 5: 687–698. Wang F, Feng XC, Li YM, Yang H, Ma TH (2006). Aquaporins as potential drug targets. Acta Pharmacol Sin 27: 395–401. Preston GM et al. (1992). Science 256: 385–387. Yasui M et al. (1999). Nature 402: 184–187. We recommend that any citations to information in the Guide are presented in the following format: Alexander SPH, Mathie A, Peters JA (2008). Guide to Receptors and Channels (GRAC), 3rd edn. Br J Pharmacol 153 (Suppl. 2): S1-S209. Overview: Calcium (Ca2+) channels are voltage-gated ion channels present in the membrane of most excitable cells. The nomenclature for Ca2+ channels was proposed by Ertel et al. (2000) and approved by the NC-IUPHAR subcommittee on Ca2+ channels (Catterall et al., 2005). Ca2+ channels form hetero-oligomeric complexes. The α1 subunit is pore-forming and provides the extracellular binding site(s) for practically all agonists and antagonists. The 10 cloned α-subunits can be grouped into three families: (1) the high-voltage activated dihydropyridine-sensitive (L-type, Cav1.x) channels; (2) the high-voltage activated dihydropyridine-insensitive (Cav2.x) channels and (3) the low-voltage-activated (T-type, Cav3.x) channels. Each α1 subunit has four homologous repeats (I-IV), each repeat having six transmembrane domains and a pore-forming region between transmembrane domains S5 and S6. Gating is thought to be associated with the membrane-spanning S4 segment, which contains highly conserved positive charges. Many of the α1-subunit genes give rise to alternatively spliced products. At least for high-voltage activated channels, it is likely that native channels comprise co-assemblies of α1, β and α2-δ subunits. The γ subunits have not been proven to associate with channels other than α1 s. The α2-δ1 and α2-δ2 subunits bind gabapentin and pregabalin. In many cell types, P and Q current components cannot be adequately separated and many researchers in the field have adopted the terminology ‘P/Q-type’ current when referring to either component. Abbreviations: FPL64176, 2,5-dimethyl-4-[2(phenylmethyl)benzoyl]-H-pyrrole-3-carboxylate; SB-209712, (1,6,bis{1-[4-(3-phenylpropyl)piper-idinyl]}hexane); (—)-(S)-BAYK8664, (—)-(S)-methyl-1,4-dihydro-2,6-dimethyl-3-nitro-4-(2-trifluromethylphenyl)-pyridine-5-carboxylate; SNX482, 41 amino acid peptide-(GVDKAGCRYMFGGCSVNDDCCPRLGCHSLFSYCAWDLTFSD); SZ(+)-(S)-202-791, isopropyl 4-(2,1,3-benzoxadiazol-4-yl)-1,4-dihydro-2,6-dimethyl-5-nitro-3-pyridinecarboxylate Catterall WA (2000). Structure and regulation of voltage-gated Ca2+ channels. Annu Rev Cell Dev Biol 16: 521–555. Catterall WA, Perez-Reyes E, Snutch TP, Striessing J (2005). International Union of Pharmacology. XLVIII. Nomenclature and structure-function relationships of voltage-gated calcium channels. Pharmacol Rev 57: 411–425. Davies A, Hendrich J, Van Minh AT, Wratten J, Douglas L, Dolphin AC (2007). Functional biology of the alpha(2)delta subunits of voltage-gated calcium channels. Trends Pharmacol Sci 28: 220–228. Dolphin AC (2003). G protein modulation of voltage-gated calcium channels. Pharmacol Rev 55: 607–627. Elmslie KS (2004). Calcium channel blockers in the treatment of disease. J Neurosci Res 75: 733–741. Ertel EA, Campbell KP, Harpold MM, Hofmann F, Mori Y, Perez-Reyes E et al. (2000). Nomenclature of voltage-gated calcium channels. Neuron 25: 533–535. Hofmann F, Lacinova L, Klugbauer N (1999). Voltage-dependent calcium channels; from structure to function. Rev Physiol Biochem Pharmacol 139: 35–87. Kochegarov AA (2003). Pharmacological modulators of voltage-gated calcium channels and their therapeutic application. Cell Calcium 33: 145–162. Lewis RJ, Garcia ML (2003). Therapeutic potential of venom peptides. Nat Rev Drug Discov 2: 790–802. Lory P, Chemin J (2007). Towards the discovery of novel T-type calcium channel blockers. Expert Opin Ther Targets 11: 717–722. Nelson MT, Todorovic SM, Perez-Reyes E (2006). The role of T-type calcium channels in epilepsy and pain. Curr Pharm Des 12: 2189–2197. Perez-Reyes E (2003). Molecular physiology of low-voltage-activated T-type calcium channels. Physiol Rev 83: 117–161. Taylor CP, Anelotti T, Fauman E (2007). Pharmacology and mechanism of action of pregabalin; the calcium channel alpha2-delta subunit as a target for antiepileptic drug discovery. Epilepsy Res 73: 137–150. Terlau H, Olivera BM (2004). Conus venoms: a rich source of novel ion channel-targeted peptides. Physiol Rev 84: 41–68. Triggle DJ (2006). L-type calcium channels. Curr Pharm Des 12: 443–457. Triggle DJ (2007). Calcium channel antagonists: clinical uses - past, present and future. Biochem Pharmacol 74: 1–9. Trimmer JS, Rhodes KJ (2004). Localisation of voltage-gated ion channels in mammalian brain. Annu Rev Physiol 66: 477–519. Yu FH, Catterall WA (2004). The VGL-chanome: a protein superfamily specialized for electrical signaling and ionic homeostasis. Sci STKE 2004 (253): re15. We recommend that any citations to information in the Guide are presented in the following format: Alexander SPH, Mathie A, Peters JA (2008). Guide to Receptors and Channels (GRAC), 3rd edn. Br J Pharmacol 153 (Suppl. 2): S1-S209. Overview: CatSper channels (CatSper1-4; nomenclature as agreed by NC-IUPHAR, Clapham and Garbers, 2005) are putative 6TM, voltage-gated, calcium permeant channels that are presumed to assemble as a tetramer of α-like subunits and mediate the current ICatSper. In mammals, CatSper subunits are structurally most closely related to individual domains of voltage-activated calcium channels (Cav) (Ren et al., 2001). CatSper1 (Ren et al., 2001), CatSper2 (Quill et al., 2001) and CatSpers 3 and 4 (Lobley et al., 2003; Lin et al., 2005; Qi et al., 2007), in common with a recently identified putative 2TM auxiliary CatSperβ protein (Liu et al., 2007), are restricted to the testis and localised to the principle piece of sperm tail. CatSper channel subunits expressed singly, or in combination, fail to functionally express in heterologous expression systems (Quill et al., 2001; Ren et al., 2001). The properties of CatSper1 tabulated above are derived from whole cell voltage-clamp recordings comparing currents endogenous to spermatozoa isolated from the corpus epididymis of wild-type and Catsper1(-/-) mice (Kirichok et al., 2006). ICatSper is also undetectable in the spermatozoa of Catsper2(-/-), Catsper3(-/-), or Catsper4(-/-) mice and CatSper 1 associates with CatSper 2, 3, or 4 in heterologous expression systems (Qi et al., 2007). Moreover, targeted disruption of Catsper1, 2, 3, or 4 genes results in an identical phenotype in which spermatozoa fail to exhibit the hyperactive movement (whip-like flagellar beats) necessary for penetration of the egg cumulus and zona pellucida and subsequent fertilization. Such disruptions are associated with a deficit in alkalinization and depolarization-evoked Ca2+ entry into spermatozoa (Carlson et al., 2003, 2005; Qi et al., 2007). Thus, it is likely that the CatSper pore is formed by a heterotetramer of CatSpers1-4 (Qi et al., 2007). The driving force for Ca2+ entry is principally determined by a mildly outwardly rectifying K+ channel (KSper) that, like CatSpers, is activated by intracellular alkalinization (Navarro et al., 2007). KSper is not yet identified, but its properties are most consistent with mSlo3, a protein detected only in testis (Navarro et al., 2007). Clapham DE, Garbers DL (2005). International Union of Pharmacology. L. Nomenclature and structure-function relationships of CatSper and two-pore channels. Pharmacol Rev 57: 451–454. Quill TA, Wang D, Garbers DL (2006). Insights into sperm cell motility through sNHE and the CatSpers. Mol Cell Endocrinol 250: 84–92. Zhang D, Gopalakrishnan M (2005). Sperm ion channels: molecular targets for the next generation of contraceptive medicines? J Androl 26: 643–653. Carlson AE et al. (2005). J Biol Chem 280: 32238–32244. Carlson AE et al. (2003). Proc Natl Acad Sci USA 100: 14864–14868. Kirichok Y et al. (2006). Nature 439: 737–740. Lin J-L et al. (2005). Biol Reprod 73: 1235–1242. Liu J et al. (2007). J Biol Chem 282: 18945–18952. Lobley A et al. (2003). Reprod Biol Endocrinol 1: 53. Navarro B et al. (2007). Proc Natl Acad Sci USA 104: 7688–7692. Qi H et al. (2007). Proc Natl Acad Sci USA 104: 1219–1223. Quill TA et al. (2001). Proc Natl Acad Sci USA 98: 12527–12531. Ren D et al. (2001). Nature 413: 603–609. We recommend that any citations to information in the Guide are presented in the following format: Alexander SPH, Mathie A, Peters JA (2008). Guide to Receptors and Channels (GRAC), 3rd edn. Br J Pharmacol 153 (Suppl. 2): S1-S209. Overview: Chloride channels are a functionally and structurally diverse group of anion selective channels involved in processes including the regulation of the excitability of neurones, skeletal, cardiac and smooth muscle, cell volume regulation, transepithelial salt transport, the acidification of internal and extracellular compartments, the cell cycle and apoptosis (reviewed by Nilius and Droogmans, 2003). Excluding the transmitter-gated GABA and glycine receptors (see separate tables), well characterised chloride channels can be classified as the voltage-sensitive ClC subfamily, calcium-activated channels, high (maxi) conductance channels, the cystic fibrosis transmembrane conductance regulator (CFTR) and volume regulated channels. No official recommendation exists regarding the classification of chloride channels. Functional chloride channels that have been cloned from, or characterised within, mammalian tissues are listed. ClC-family: The mammalian ClC family (reviewed by Jentsch et al., 2002; Nilius and Droogmans, 2003; Chen, 2005; Jentsch et al., 2005a, b; Dutzler, 2006) contains 9 members that fall into three groups; ClC-1, ClC-2, hClC-Ka (rClC-K1) and hClC-Kb (rClC-K2); ClC-3 to ClC-5, and ClC-6 and −7. ClC-1 and ClC-2 are plasma membrane chloride channels as are ClC-Ka and ClC-Kb (largely expressed in the kidney) when associated with barttin (ENSG00000162399), a 320 amino acid 2TM protein (Estévez et al., 2001). The localisation of CIC-3 (ENSG00000109572), ClC-4 (ENSG00000073464) and ClC-5 (ENSG00000171365) is likely to be predominantly intracellular and recent reports indicate that ClC-4 and ClC-5 (and by inference ClC-3) function as Cl−/H+ antiporters, rather than classical Cl− channels (Picollo and Pusch, 2005; Scheel et al., 2005; reviewed by Miller, 2006 & Pusch et al., 2006). An intracellular location has been demonstrated for ClC-6 (ENSG00000011021) and ClC-7 (ENSG00000103249) also (reviewed by Jentsch et al., 2005b). Alternative splicing increases the structural diversity within the ClC family (e.g. for ClC-2, ClC-3 ClC-5 and ClC-6). The crystal structure of two bacterial ClC channels has been described (Dutzler et al., 2002). Each ClC subunit, with a complex topology of 17 intramembrane α-helices, contributes a single pore to a dimeric ‘double-barrelled’ ClC channel that contains two independently-gated pores, confirming the predictions of previous functional and structural investigations (reviewed by Estévez and Jentsch, 2002; Babini and Pusch, 2004; Chen, 2005; Dutzler, 2006). As found for ClC-4 and ClC-5, the prokaryotic ClC homologue functions as an H+ /Cl− antiporter, rather than as an ion channel (Accardi and Miller, 2004). ClC channels other than ClC-3 display the permeability sequence Cl− > Br– > I− (at physiological pH); for ClC-3, I− > Cl−. ClC-1 has significant opening probability at resting membrane potential, accounting for 75% of the membrane conductance at rest in skeletal muscle, and is important for repolarization and for stabilization of the membrane potential. S-(—)CPP, A-9-C and niflumic acid act intracellularly and exhibit a strongly voltage-dependent block with strong inhibition at negative voltages and relief of block at depolarized potentials (Liantonio et al., 2007 and reviewed by Pusch et al., 2002). Mutations in the ClC-1 gene result in myotonia congenita. Although ClC-2 can be activated by cell swelling, it does not correspond to the VRAC channel (see below). Alternative potential physiological functions for ClC-2 are reviewed by Jentsch et al. (2005b). Disruption of the ClC-2 gene in mice is associated with testicular and retinal degeneration. Functional expression of human ClC-Ka and ClC-Kb requires the presence of barttin (Estévez et al., 2001; Scholl et al., 2006). The rodent homologue (ClC-K1) of ClC-Ka demonstrates limited expression as a homomer, but its function is enhanced by barttin which increases both channel opening probablility in the physiological range of potentials and single channel conductance (Estévez et al., 2001; Scholl et al., 2006). Knock out of the ClC-K1 channel induces nephrogenic diabetes insipidus. Classic (type III) Bartter's syndrome and Gitelman's variant of Bartter's syndrome are associated with mutations of the ClC-Kb chloride channel (reviewed by Jentsch et al., 2005b; Uchida and Sasaki, 2005). ClC-Ka is approximately 5 to 6-fold more sensitive to block by 3-phenyl-CPP and DIDS than ClC-Kb (Liantonio et al., 2002). The biophysical and pharmacological properties of ClC-3, and the relationship of the protein to the endogenous volume-regulated anion channel(s) VRAC (see Guan et al., 2006 and below) are controversial and further complicated by the inference that ClC-3 is a Cl−/H+ exchanger, rather than an ion channel (Picollo and Pusch, 2005). Activation of heterologously expressed ClC-3 by cell swelling in response to hypotonic solutions is disputed, as are other aspects of regulation, including inhibition by PKC. Lack of chloride ion channel function of ClC-3 heterologously expressed in HEK 293 cells, and inserted in to the plasma membrane, has additionally been claimed. However, phosphorylation by exogenously introduced CaM kinase II may be required for high activity of ClC-3 in this paradigm. In ClC-3 knock-out mice (Clcn3−/-), volume regulated anion currents (ICl,swell) persist (Stobrawa et al., 2001; Arreola et al., 2002), and demonstrate kinetic, ionic selectivity and pharmacological properties similar to ICl,swell recorded from cells of wild-type (Clcn3+/+) animals, indicating that ClC-3 is not indispensable for such regulation (Yamamoto-Mizuma et al., 2004). However, both ClC-3 antisense and novel anti-ClC-3 antibodies are reported to reduce VRAC function in several cell systems (e.g. Hermoso et al., 2002; Wang et al., 2003), and the sensitivity of ICl,swell to regulators such as PKC, [ATP]i and [Mg2+]i differs between cells of Clcn3(+/+) and Clcn3(-/-) mice (Yamamoto-Mizuma et al., 2004). A splice variant of ClC-3 (i.e. ClC-3B) upregulated by NHERF, is expressed in the plasma membrane of epithelial cells and mediates outwardly rectifying currents activated by depolarisation. In association with CFTR, ClC-3B is activated by PKA. ClC-3B is a candidate for the outwardly rectifying chloride channel ORCC (Ogura et al., 2002). Results obtained from ClC-3 knock-out mice suggest an endosomal/synaptic vesicle location for the channel and a role, via the dissipation of electrical potential, in the acidification of vesicles. Mice lacking ClC-3 display total degeneration of the hippocampus and retinal degeneration (Stobrawa et al., 2001; Jentsch et al., 2005b). Loss-of-function mutations of ClC-5 are associated with proteinuria, hypercalciuria and kidney stone formation (Dent's disease). A ClC 5 knock-out provides a mouse model of this disease. Disruption of the ClC-7 gene in mice leads to osteopetrosis, blindness and lysosomal dysfunction (Jentsch et al., 2005b). CFTR: CFTR, a 12TM, ABC type protein, is a cAMP-regulated epithelial cell membrane Cl− channel involved in normal fluid transport across various epithelia. The most common mutation in CFTR (i.e. the deletion mutant, Δ508) results in impaired trafficking of CFTR and reduces its incorporation into the plasma membrane causing cystic fibrosis. In addition to acting as an anion channel per se, CFTR may act as a regulator of several other conductances including inhibition of the epithelial Na channel (ENaC), calcium activated chloride channels (CaCC) and volume regulated anion channel (VRAC), activation of the outwardly rectifying chloride channel (ORCC), and enhancement of the sulphonylurea sensitivity of the renal outer medullary potassium channel (ROMK2), (reviewed by Nilius and Droogmans, 2003). CFTR also regulates TRPV4, which provides the Ca2+ signal for regulatory volume decrease in airway epithelia (Arniges et al., 2004). The activities of CFTR and the chloride-bicarbonate exchangers SLC26A3 (DRA) and SLC26A6 (PAT1) are mutually enhanced by a physical association between the regulatory (R) domain of CFTR and the STAS domain of the SCL26 transporters, an effect facilitated by PKA-mediated phosphorylation of the R domain of CFTR (Ko et al., 2004). CFTR contains two cytoplasmic nucleotide binding domains (NBDs) that bind ATP. A single open-closing cycle is hypothesised to involve, in sequence: binding of ATP at the N-terminal NBD1, ATP binding to the C-terminal NBD2 leading to the formation of an intramolecular NBD1-NBD2 dimer associated with the open state, and subsequent ATP hydrolysis at NBD2 facilitating dissociation of the dimer and channel closing, and the initiation of a new gating cycle (Vergani et al., 2005; Aleksandrov et al., 2007). Phosphorylation by PKA at sites within a cytoplasmic regulatory (R) domain are required for the binding ATP to gate CFTR (Gadsby et al., 2006). PKC (and PKGII within intestinal epithelial cells via guanylin-stimulated cGMP formation) positively regulate CFTR activity. Calcium activated chloride channel: Chloride channels activated by intracellular calcium (CaCC) are widely expressed in excitable and non-excitable cells where they perform diverse functions (Hartzell et al., 2005). The molecular nature of CaCC is unclear. Members of the initial putative calcium-activated chloride channel proteins (the CLCA family) have