The Concise Guide to Pharmacology 2013/14: Ion Channels

The Concise Guide to PHARMACOLOGY 2013/14 provides concise overviews of the key properties of over 2000 human drug targets with their pharmacology, plus links to an open access knowledgebase of drug targets and their ligands (www.guidetopharmacology.org), which provides more detailed views of target and ligand properties. The full contents can be found at http://onlinelibrary.wiley.com/doi/10.1111/bph.12444/full. Ion channels are one of the seven major pharmacological targets into which the Guide is divided, with the others being G protein-coupled receptors, ligand-gated ion channels, catalytic receptors, nuclear hormone receptors, transporters and enzymes. These are presented with nomenclature guidance and summary information on the best available pharmacological tools, alongside key references and suggestions for further reading. A new landscape format has easy to use tables comparing related targets. It is a condensed version of material contemporary to late 2013, which is presented in greater detail and constantly updated on the website www.guidetopharmacology.org, superseding data presented in previous Guides to Receptors and Channels. It is produced in conjunction with NC-IUPHAR and provides the official IUPHAR classification and nomenclature for human drug targets, where appropriate. It consolidates information previously curated and displayed separately in IUPHAR-DB and the Guide to Receptors and Channels, providing a permanent, citable, point-in-time record that will survive database updates.


Aquaporins
Overview: Aquaporins and aquaglyceroporins (provisional nomenclature) 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) [54], 12 additional members of the family have been identified, although little is known about the functional properties of two of these (AQP11; Q8NBQ7 and AQP12A; Q8IXF9). The other 11 aquaporins can be divided into two families (aquaporins and aquaglyceroporins) depending on whether they are permeable to glycerol [53]. 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 [52]. Functional AQPs exist as tetramers but, unusually, each subunit contains a separate pore, so each channel has four pores.  [71]. CatSper1 [71], CatSper2 [70] and CatSpers 3 and 4 [61,66,69], in common with a recently identified putative 2TM auxiliary CatSperβ protein [65] and two putative 1TM associated CatSperγ and CatSperδ proteins [58,73], are restricted to the testis and localised to the principle piece of sperm tail.  [70][71]. 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 [62] and also mature human sperm [63,72]. ICatSper is also undetectable in the spermatozoa of Catsper2 (-/-) , Catsper3 (-/-) , or Catsper4 (-/-)  in association with the auxiliary subunits (β, γ, δ) that are also essential for function [58]. CatSper channels are required for the increase in intracellular Ca 2+ concentration in sperm evoked by egg zona pellucida glycoproteins [74]. The driving force for Ca 2+ entry is principally determined by a mildly outwardly rectifying K + channel (KSper) that, like CatSpers, is activated by intracellular alkalinization [68]. Mouse KSper is encoded by mSlo3, a protein detected only in testis [67- 68,75]. In human sperm, such alkalinization may result from the activation of Hv1, a proton channel [64]. Mutations in CatSpers are associated with syndromic and non-syndromic male infertility [60]. In human ejaculated spermatozoa, progesterone (8,17]. In addition, certain prostaglandins (e.g. PGF1α, PGE1) also potentiate CatSper mediated currents [63,72].

Chloride channels
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 in [92]).
Excluding the transmitter-gated GABAA and glycine receptors (see separate tables), well characterised chloride channels can be classified as certain members of the voltage-sensitive ClC subfamily, calcium-activated channels, high (maxi) conductance channels, the cystic fibrosis transmembrane conductance regulator (CFTR) and volume regulated channels [155]. 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 with the exception of several classes of intracellular channels (e.g. CLIC) that are reviewed by in [96].

ClC family
Overview: The mammalian ClC family (reviewed in [77,87,92,94,108]) contains 9 members that fall, on the basis of sequence homology, 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. ClC-Ka and ClC-Kb are also plasma membrane channels (largely expressed in the kidney and inner ear) when associated with barttin (BSND, Q8WZ55), a 320 amino acid 2TM protein [97]. The localisation of the remaining members of the ClC family is likely to be predominantly intracellular in vivo, although they may traffic to the plasma membrane in overexpression systems.
Numerous recent reports indicate that ClC-4, ClC-5, ClC-6 and ClC-7 (and by inference ClC-3) function as Cl -/H + antiporters (secondary active transport), rather than classical Clchannels [103,114,125,134,146]; reviewed in [77,139]). It has recently been reported that the activity of ClC-5 as a Cl -/H + exchanger is important for renal endocytosis [127]. Alternative splicing increases the structural diversity within the ClC family. The crystal structure of two bacterial ClC proteins has been described [95] and a eukaryotic ClC transporter (CmCLC) has recently been described at 3.5 Å resolution [99]. Each ClC subunit, with a complex topology of 18 intramembrane segments, 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 in [87,94,108,139]). As found for ClC-4, ClC-5, ClC-6 and ClC-7, the prokaryotic ClC homologue (ClC-ec1) and CmCLC function as H + /Cl antiporters, rather than as ion channels [76,99]. The generation of monomers from dimeric ClC-ec1 has firmly established that each ClC subunit is a functional unit for transport and that cross-subunit interaction is not required for Cl -/H + exchange in ClC transporters [141].  Comments: ClC channels display the permeability sequence Cl -> Br -> I -(at physiological pH). 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 stabilization of the membrane potential. S-(-)CPP, 9-A-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 ( [115] and reviewed in [138] . Alternative potential physiological functions for ClC-2 are reviewed in [137]. Functional expression of human ClC-Ka and ClC-Kb requires the presence of barttin [97,147] reviewed in [98]. The properties of ClC-Ka/barttin and ClC-Kb/barttin tabulated are those observed in mammalian expression systems: in oocytes the channels display time-and voltage-dependent gating. 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 [97,101,147] reviewed in [98]). ClC-Ka is approximately 5 to 6-fold more sensitive to block by 3-phenyl-CPP and DIDS than ClC-Kb, while newly synthesized benzofuran derivatives showed the same blocking affinity (<10 μM) on both CLC-K isoforms [116]. The biophysical and pharmacological properties of ClC-3, and the relationship of the protein to the endogenous volume-regulated anion channel(s) VRAC [78,105] are controversial and further complicated by the possibility that ClC-3 may function as both a Cl -/H + exchanger and an ion channel [78, 134,156]. The functional properties tabulated are those most consistent with the close structural relationship between ClC-3, ClC-4 and ClC-5. Activation of heterologously expressed ClC-3 by cell swelling in response to hypotonic solutions is disputed, as are many other aspects of its regulation. Dependent upon the predominant extracellular anion (e.g. SCNversus Cl -), CIC-4 can operate in two transport modes: a slippage mode in which behaves as an ion channel and an exchanger mode in which unitary transport rate is 10-fold lower [79]. Similar findings have been made for ClC-5 [158]. ClC-7 associates with a β subunit, Ostm1, which increases the stability of the former [112] and is essential for its function [114].

CFTR
Overview: CFTR, a 12TM, ABC transporter-type protein, is a cAMP-regulated epithelial cell membrane Clchannel involved in normal fluid transport across various epithelia. Of the 1700 mutations identified in CFTR, the most common is the deletion mutant ΔF508 (a class 2 mutation) which results in impaired trafficking of CFTR and reduces its incorporation into the plasma membrane causing cystic fibrosis (reviewed in [88]). Channels carrying the ΔF508 mutation that do traffic to the plasma membrane demonstrate gating defects. Thus, pharmacological resto-ration the function of the ΔF508 mutant would require a compound that embodies 'corrector' (i.e. facilitates folding and trafficking to the cell surface) and 'potentiator' (i.e. promotes opening of channels at the cell surface) activities [88]. 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 in [126]). CFTR also regulates TRPV4, which provides the Ca 2+ signal for regulatory volume decrease in airway epithelia [81]. 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 [109]. Comments: In addition to the agents listed in the table, the novel small molecule, ataluren, induces translational read through of nonsense mutations in CFTR (reviewed in [150]). Corrector compounds that aid the folding of DF508CFTR to increase the amount of protein expressed and potentially delivered to the cell surface include VX-532 (which is also a potentiator), VRT-325, KM11060, Corr-3a and Corr-4a see [155] for details and structures of Corr-3a and Corr-4a

Calcium activated chloride channel
Overview: Chloride channels activated by intracellular calcium (CaCC) are widely expressed in excitable and non-excitable cells where they perform diverse functions [106]. The molecular nature of CaCC has been uncertain with both CLCA, TWEETY and BEST genes having been considered as likely candidates [92,107,117]. It is now accepted that CLCA expression products are unlikely to form channels per se and probably function as cell adhesion proteins, or are secreted [133]. Similarly, TWEETY gene products do not recapictulate the properties of endogenous CaCC. The bestrophins encoded by genes BEST1-4 have a topology more consistent with ion channels [107] and form chloride channels that are activated by physiological concentrations of Ca 2+ , but whether such activation is direct is not known [107]. However, currents generated by bestrophin over-expression do not resemble native CaCC currents. The evidence for and against bestrophin proteins forming CaCC is critically reviewed by Duran et al. in their 2010 paper [92]. Recently, a new gene family, TMEM16 (anoctamin) consisting of 10 members (TMEM16A-K; anoctamin 1-10) has been identified and there is firm evidence that some of these members form chloride channels [91,110]. TMEM16A (anoctamin 1; Ano 1) produces Ca 2+ -activated Clcurrents with kinetics similar to native CaCC currents recorded from different cell types [86,142,148,157]. Knockdown of TMEM16A greatly reduces currents mediated by calcium-activated chloride channels in submandibular gland cells [157] and smooth muscle cells from pulmonary artery [118]. In TMEM16A (-/-) mice secretion of Ca 2+ -dependent Clsecretion by several epithelia is reduced [132,142]. Alternative splicing regulates the voltage-and Ca 2+ -dependence of TMEM16A and such processing may be tissue-specific manner and thus contribute to functional diversity [100]. There are also reports that TMEM16B (anoctamin 2; Ano 2) supports CaCC activity (e.g. [135]) and in TMEM16B (-/-) mice Ca-activated Clcurrents in the main olfactory epithelium (MOE) and in the vomeronasal organ are virtually absent [85].  Comments: Blockade of ICl(Ca) by niflumic acid, DIDS and 9-A-C is voltage-dependent whereas block by NPPB is voltageindependent [106]. Extracellular niflumic acid; DCDPC and 9-A-C (but not DIDS) exert a complex effect upon ICl(Ca) in vascular smooth muscle, enhancing and inhibiting inwardly and outwardly directed currents in a manner dependent upon [Ca 2+ ]i (see [113] for summary). Considerable crossover in pharmacology with large conductance Ca 2+ -activated K + channels also exists (see [104] for overview). Two novel compounds, CaCCinh-A01

Subunits
and CaCCinh-B01 have recently been identified as blockers of calcium-activated chloride channels in T84 human intestinal epithelial cells [89] for structures). Significantly, other novel compounds totally block currents mediated by TMEM116A, but have only a modest effect upon total current mediated by CaCC native to T84 cells or human bronchial epithelial cells, suggesting that TMEM16A is not the predominant CaCC in such cells [124]. CaMKII modulates CaCC in a tissue dependent manner (reviewed by [106,113]). CaMKII inhibitors block activation of ICl(Ca) in T84 cells but have no effect in parotid acinar cells. In tracheal and arterial smooth muscle cells, but not portal vein myocytes, inhibition of CaMKII reduces inactivation of ICl(Ca). Intracellular Ins(3,4,5,6)P4 may act as an endogenous negative regulator of CaCC channels activated by Ca 2+ , or CaMKII.
Smooth muscle CaCC are also regulated positively by Ca 2+dependent phosphatase, calcineurin (see [113] for summary).

Maxi chloride channel
Overview: Maxi Clchannels are high conductance, anion selective, channels initially characterised in skeletal muscle and subsequently found in many cell types including neurones, glia, cardiac muscle, lymphocytes, secreting and absorbing epithelia, macula densa cells of the kidney and human placenta syncytiotrophoblasts [144]. The physiological significance of the maxi Clchannel is uncertain, but roles in cell volume regulation and apoptosis have been claimed. Evidence suggests a role for maxi Clchannels as a conductive pathway in the swelling-induced release of ATP from mouse mammary C127i cells that may be important for autocrine and paracrine signalling by purines [93,143]. A similar channel mediates ATP release from macula densa cells within the thick ascending of the loop of Henle in response to changes in luminal NaCl concentration [83]. A family of human high conductance Clchannels (TTYH1-3) that resemble Maxi Clchannels has been cloned [152], but alternatively, Maxi Clchannels have also been suggested to correspond to the voltage-dependent anion channel, VDAC, expressed at the plasma membrane [82,128].

Channels
ATP is a voltage dependent permeant blocker of single channel activity (PATP/PCl = 0.08-0.1); channel activity increased by patch-excision; channel opening probability (at steady-state) maximal within approximately ± 20 mV of 0 mV, opening probability decreased at more negative and (commonly) positive potentials yielding a bell-shaped curve; channel conductance and opening probability regulated by annexin 6 Maxi Clis also activated by G protein-coupled receptors and cell swelling. tamoxifen and toremifene are examples of triphenylethylene anti-oestrogens Comments: Differing ionic conditions may contribute to variable estimates of γ reported in the literature. Inhibition by arachidonic acid (and cis-unsaturated fatty acids) is voltageindependent, occurs at an intracellular site, and involves both channel shut down (Kd = 4-5 μM) and a reduction of γ (Kd = 13-14 μM). Blockade of channel activity by SITS, DIDS, Gd 3+ and ara-chidonic acid is paralleled by decreased swelling-induced release of ATP [93,143]. Channel activation by anti-oestrogens in whole cell recordings requires the presence of intracellular nucleotides and is prevented by pre-treatment with 17β-estradiol, dibutyryl cAMP, or intracellular dialysis with GDPβS [90]. Activation by tamoxifen is suppressed by low concentrations of okadaic acid, suggesting that a dephosphorylation event by protein phosphatase PP2A occurs in the activation pathway [90]. In contrast, 17β-estradiol and tamoxifen appear to directly inhibit the maxi Clchannel of human placenta reconstituted into giant liposomes and recorded in excised patches [140].

Volume regulated chloride channels
Overview: Volume activated chloride channels (also termed VSOAC, volume-sensitive organic osmolyte/anion channel; VRC, volume regulated channel and VSOR, volume expansion-sensing outwardly rectifying anion channel) participate in regulatory volume decrease (RVD) in response to cell swelling. VRAC may also be important for several other processes including the regulation of membrane excitability, transcellular Cltransport, angiogenesis, cell proliferation, necrosis, apoptosis, glutamate release from astrocytes, insulin (INS, P01308) release from pancreatic β cells and resistance to the anti-cancer drug, cisplatin (reviewed by [84,123,126,129]). VRAC may not be a single entity, but may instead represent a number of different channels that are expressed to a variable extent in different tissues and are differentially activated by cell swelling. In addition to ClC-3 expression products (see above) several former VRAC candidates including MDR1 P-glycoprotein, Icln, Band 3 anion exchanger and phospholemman are also no longer considered likely to fulfil this function (see reviews [126,145]).

Channels
outward rectification due to voltage dependence of γ; inactivates at positive potentials in many, but not all, cell types; time dependent inactivation at positive potentials; intracellular ionic strength modulates sensitivity to cell swelling and rate of channel activation; rate of swelling-induced activation is modulated by intracellular ATP concentration; ATP dependence is independent of hydrolysis and modulated by rate of cell swelling; inhibited by increased intracellular free Mg 2 + concentration; swelling induced activation of several intracellular signalling cascades may be permissive of, but not essential to, the activation of VRAC including: the Rho-Rho kinase-MLCK; Ras-Raf-MEK-ERK; PIK3-NOX-H2O2 and Src-PLCγ-Ca 2 + pathways; regulation by PKCα required for optimal activity; cholesterol depletion enhances activity; activated by direct stretch of β1-integrin VRAC is also activated by cell swelling and low intracellular ionic strength. VRAC is also blocked by chromones, extracellular nucleotides and nucleoside analogues Comments: In addition to conducting monovalent anions, in many cell types the activation of VRAC by a hypotonic stimulus can allow the efflux of organic osmolytes such as amino acids and polyols that may contribute to RVD.
Other chloride channels: In addition to some intracellular chloride channels that are not considered here, plasma membrane channels other than those listed have been functionally described. Many cells and tissues contain outwardly rectifying chloride channels (ORCC) that may correspond to VRAC active under isotonic conditions. A cAMP-activated Clchannel that does not correspond to CFTR has been described in intestinal Paneth cells [154]. A Cl channel activated by cGMP with a dependence on raised intracellular Ca 2+ has been recorded in various vascular smooth muscle cells types, which has a pharma-cology and biophysical characteristics very different from the 'conventional' CaCC [119,136]. It has been proposed that bestrophin-3 (BEST3, Q8N1M1) is an essential component of the cGMP-activated channel [120]. A proton-activated, outwardly rectifying anion channel has also been described [111].

Connexins and Pannexins
Overview: Gap junctions are essential for many physiological processes including cardiac and smooth muscle contraction, regulation of neuronal excitability and epithelial electrolyte transport [162][163][164]. Gap junction channels allow the passive diffusion of molecules of up to 1,000 Daltons which can include nutrients, metabolites and second messengers (such as IP3) as well as cations and anions. 21 connexin genes (Cx23, Cx25, Cx26, Cx30, Cx30.2, Cx30.3, Cx31, Cx31.1, Cx31.9, Cx32, Cx36, Cx37, Cx40, Cx40.1, Cx43, Cx45, Cx46, Cx47, Cx50, Cx59, Cx62) and 3 pannexin genes (Px1, Px2, Px3; which are structur-ally related to the invertebrate innexin genes) code for gap junction proteins (provisional nomenclature) in humans. Each connexin gap junction comprises 2 hemichannels or 'connexons' which are themselves formed from 6 connexin molecules. The various connexins have been observed to combine into both homomeric and heteromeric combinations, each of which may exhibit different functional properties. It is also suggested that individual hemichannels formed by a number of different connexins might be functional in at least some cells [165]. Connexins have a common topology, with four α-helical transmembrane domains, two extracellular loops, a cytoplasmic loop, and N-and C-termini located on the cytoplasmic membrane face. In mice, the most abundant connexins in electrical synapses in the brain seem to be Cx36, Cx45 and Cx57 [168]. Mutations in connexin genes are associated with the occurrence of a number of pathologies, such as peripheral neuropathies, cardiovascular diseases and hereditary deafness. The pannexin genes Px1 and Px2 are widely expressed in the mammalian brain [169]. Like the connexins, at least some of the pannexins can form hemichannels [162,166]. Comments: Connexins are most commonly named according to their molecular weights, so, for example, Cx23 is the connexin protein of 23 kDa. This can cause confusion when comparing between species -for example, the mouse connexin Cx57 is orthologous to the human connexin Cx62. No natural toxin or specific inhibitor of junctional channels has been identified yet however two compounds often used experimentally to block connexins are carbenoxolone and flufenamic acid [167]. At least some pannexin hemichannels are more sensitive to carbenoxolone than connexins but much less sensitive to flufenamic acid [161]. It has been suggested that 2-aminoethoxydiphenyl borate (2-APB) may be a more effective blocker of some connexin channel subtypes (Cx26, Cx30, Cx36, Cx40, Cx45, Cx50) compared to others (Cx32, Cx43, Cx46, [160]).

Cyclic nucleotide-regulated channels
Overview: Cyclic nucleotide-gated (CNG) channels are responsible for signalling in the primary sensory cells of the vertebrate visual and olfactory systems. A standardised nomenclature for CNG channels has been proposed by the NC-IUPHAR subcommittee on voltage-gated ion channels [176].
CNG channels are voltage-independent cation channels formed as tetramers. Each subunit has 6TM, with the pore-forming domain between TM5 and TM6. CNG channels were first found in rod photoreceptors [175,177], where light signals through rhodopsin and transducin to stimulate phosphodiesterase and reduce intracellular cGMP level. This results in a closure of CNG channels and a reduced 'dark current'. Similar channels were found in the cilia of olfactory neurons [178] and the pineal gland [174]. The cyclic nucleotides bind to a domain in the C terminus of the subunit protein: other channels directly binding cyclic nucleotides include HCN, eag and certain plant potassium channels.

Hyperpolarisation-activated, cyclic nucleotide-gated (HCN)
The hyperpolarisation-activated, cyclic nucleotide-gated (HCN) channels are cation channels that are activated by hyperpolarisation at voltages negative to ∼-50 mV. The cyclic nucleotides cAMP and cGMP directly activate the channels and shift the activation curves of HCN channels to more positive voltages, thereby enhancing channel activity. HCN channels underlie pacemaker currents found in many excitable cells including cardiac cells and neurons [173,179]. In native cells, these currents have a variety of names, such as Ih, Iq and If. The four known HCN channels have six transmembrane domains and form tetramers. It is believed that the channels can form heteromers with each other, as has been shown for HCN1 and HCN4 [170]. A standardised nomenclature for HCN channels has been proposed by the NC-IUPHAR subcommittee on voltage-gated ion channels [176].  [172] have proven useful in identifying and studying functional HCN channels in native cells. zatebradine and cilobradine are also useful blocking agents.

Epithelial sodium channels (ENaC)
Overview: The epithelial sodium channels (ENaC) mediates sodium reabsorption in the aldosterone-sensitive distal part of the nephron and the collecting duct of the kidney. ENaC is found on other tight epithelial tissues such as the airways, distal colon and exocrine glands. ENaC activity is tightly regulated in the kidney by aldosterone, angiotensin II (AGT, P01019), vasopressin, insulin (INS, P01308) and glucocorticoids; this fine regulation of ENaC is essential to maintain sodium balance between daily intake and urinary excretion of sodium, circulating volume and blood pressure. ENaC expression is also vital for clearance of foetal lung fluid, and to maintain air-surface-liquid [195,199]. Sodium reabsorption is suppressed by the 'potassium-sparing' diuretics amiloride and triamterene. ENaC is a heteromultimeric channel made of homologous α β and γ subunits. The primary structure of αENaC subunit was identified by expression cloning [188]; β and γ ENaC were identified by functional complementation of the α subunit [189]. Each ENaC subunit contains 2 TM α helices connected by a large extracellular loop and short cytoplasmic amino-and carboxy-termini. The stoichiometry of the epithelial sodium channel in the kidney and related epithelia is, by homology with the structurally related channel ASIC1a, thought to be a heterotrimer of 1α:1β:1γ subunits [193]. P552-02 (7.6x10 -9 M), benzamil (∼1x10 -8 M), amiloride (1x10 -7 -2x10 -7 M), triamterene (∼5x10 -6 M) [189,196] γ ≈ 4-5 pS, PNa/PK > 20; tonically open at rest; expression and ion flux regulated by circulating aldosterone-mediated changes in gene transcription. The action of aldosterone, which occurs in 'early' (1.5-3 h) and 'late' (6-24 hr) phases is competitively antagonised by spironolactone, its active metabolites and eplerenone. Glucocorticoids are important functional regulators in lung/airways and this control is potentiated by thyroid hormone; but the mechanism underlying such potentiation is unclear [185,206,209]. The density of channels in the apical membrane, and hence GNa, can be controlled via both serum and glucocorticoid-regulated kinases (SGK1, 2 and 3) [190][191] and via cAMP/PKA [203]; and these protein kinases appear to act by inactivating Nedd-4/2, a ubiquitin ligase that normally targets the ENaC channel complex for internalization and degradation [186,190]. ENaC is constitutively activated by soluble and membrane-bound serine proteases, such as furin, prostasin (CAP1), plasmin and elastase [197][198]204,[207][208]. The activation of ENaC by proteases is blocked by a protein, SPLUNC1, secreted by the airways and which binds specifically to ENaC to prevent its cleavage [192]. Pharmacological inhibitors of proteases (e.g. camostat acting upon prostasin) reduce the activity of ENaC [202]. Phosphatidylinositides such as PtIns(4,5)P2 and PtIns(3,4,5)P3) stabilise channel gating probably by binding to the β and γ ENaC subunits, respectively [201,205], whilst C terminal phosphorylation of β and γ-ENaC by ERK1/2 has been reported to inhibit the withdrawal of the channel complex from the apical membrane [212]. This effect may contribute to the cAMP-mediated increase in sodium conductance.

Channels
Comments: Data in the table refer to the αβγ heteromer. There are several human diseases resulting from mutations in ENaC subunits. Liddle's syndrome (including features of salt-sensitive hypertension and hypokalemia), is associated with gain of function mutations in the β and γ subunits leading to defective ENaC ubiquitylation and increased stability of active ENaC at the cell surface [208,[210][211]. Enzymes that deubiquitylate ENaC increase its function in vivo. Pseudohypoaldosteronism type 1 (PHA-1) can occur through either mutations in the gene encoding the mineralocorticoid receptor, or loss of function mutations in genes encoding ENaC subunits [187]. Regulation of ENaC by phosphoinositides may underlie insulin (INS, P01308)-evoked renal Na + retention that can complicate the clinical management of type 2 diabetes using insulin-sensitizing thiazolidinedione drugs [194].

IP 3 receptor
Overview

Comments:
The absence of a modulator of a particular isoform of receptor indicates that the action of that modulator has not been determined, not that it is without effect.

Potassium channels
Overview: Potassium channels are fundamental regulators of excitability. They control the frequency and the shape of action potential waveform, the secretion of hormones and neurotransmitters and cell membrane potential. Their activity may be regulated by voltage, calcium and neurotransmitters (and the signalling pathways they stimulate). They consist of a primary pore-forming a subunit often associated with auxiliary regulatory subunits. Since there are over 70 different genes encoding K channels α subunits in the human genome, it is beyond the scope of this guide to treat each subunit individually. Instead, channels have been grouped into families and subfamilies based on their structural and functional properties. The three main families are the 2TM (two transmembrane domain), 4TM and 6TM families. A standardised nomenclature for potassium channels has been proposed by the NC-IUPHAR subcommittees on potassium channels [213][214][215][216].

Inwardly rectifying potassium channels
Overview: The 2TM domain family of K channels are also known as the inward-rectifier K channel family. This family includes the strong inward-rectifier K channels (KIR2.x), the G-protein-activated inward-rectifier K channels (KIR3.x) and the ATP-sensitive K channels (KIR6.x, which combine with sulphonylurea receptors (SUR)). The pore-forming a subunits form tetramers, and heteromeric channels may be formed within subfamilies (e.g. KIR3.2 with KIR3.3).

Two-P potassium channels
Overview: The 4TM family of K channels are thought to underlie many leak currents in native cells. They are open at all voltages and regulated by a wide array of neurotransmitters and biochemical mediators. The primary pore-forming αsubunit contains two pore domains (indeed, they are often referred to as two-pore domain K channels or K2P) and so it is envisaged that they form functional dimers rather than the usual K channel tetramers. There is some evidence that they can form heterodimers within subfamilies (e.g. K2P3.1 with K2P9.1). There is no current, clear, consensus on nomenclature of 4TM K channels, nor on the division into subfamilies [213]. The suggested division into subfamilies, below, is based on similarities in both structural and functional properties within subfamilies.

Ryanodine receptor
Overview: The ryanodine receptors (RyRs, provisional nomenclature) are found on intracellular Ca 2+ storage/release organelles. The family of RyR genes encodes three highly related Ca 2+ release channels: RyR1, RyR2 and RyR3, which assemble as large tetrameric structures. These RyR channels are ubiquitously expressed in many types of cells and participate in a variety of important Ca 2+ signaling phenomena (neurotransmission, secretion, etc.).
In addition to the three mammalian isoforms described below, various nonmammalian isoforms of the ryanodine receptor have been identified [218]. The function of the ryanodine receptor channels may also be influenced by closely associated proteins such as the tacrolimus (FK506)-binding protein, calmodulin [219], triadin, calsequestrin, junctin and sorcin, and by protein kinases and phosphatases.

Sodium leak channel, non-selective
Overview: The sodium leak channel, non selective (NC-IUPHAR tentatively recommends the nomenclature NaVi2.1) is structurally a member of the family of voltage-gated sodium channel family (Nav1.1 -Nav1.9) [221,228]. In contrast to the latter, NaVi2.1, is voltage-insensitive (denoted in the subscript 'vi' in the tentative nomenclature) and possesses distinctive ion selectivity and pharmacological properties. NaVi2.1, which is insensitive to tetrodotoxin (10 μM), has been proposed to mediate the tetrodotoxin-resistant and voltage-insensitive Na + leak current (IL-Na) observed in many types of neurone [222].
However, whether NaVi2.1 is constitutively active has been challenged [226]. NaVi2.1 is widely distributed within the central nervous system and is also expressed in the heart and pancreas specifically, in rodents, within the islets of Langerhans [221][222]. Comments: In native and recombinant expression systems NaVi2.1 can be activated by stimulation of NK1 (in hippocampal neurones), neurotensin (in ventral tegmental area neurones) and M3 muscarinic acetylcholine receptors (in MIN6 pancreatic β-cells) and in a manner that is independent of signalling through G-proteins [223,226]. Pharmacological and molecular biological evidence indicates such modulation to occur though a pathway that involves the activation of Src family tyrosine kinases. It is suggested that NaVi2.1 exists as a macromolecular complex with M3 receptors [226] and peptide receptors [223], in the latter instance in association with the protein UNC-80, which recruits Src to the channel complex [223,227]. By contrast, stimulation of Navi2.1 by decreased extracellular Ca 2+ concentration is G-protein dependent and involves a Ca 2+ -sensing G protein-coupled receptor and UNC80 which links Navi2.1 to the protein UNC79 in the same complex [224]. NaVi2.1 null mutant mice have severe disturbances in respiratory rhythm and die within 24 hours of birth [222]. Navi2.1 heterozygous knockout mice display increased serum sodium concentrations in comparison to wildtype littermates and a role for the channel in osmoregulation has been postulated [225].

Transient receptor potential channels
Overview: The TRP superfamily of channels (nomenclature agreed by NC-IUPHAR; [244,351]), whose founder member is the Drosophila Trp channel, exists in mammals as six families; TRPC, TRPM, TRPV, TRPA, TRPP and TRPML based on amino acid homologies. TRP subunits contain six putative transmembrane domains and assemble as homo-or hetero-tetramers to form cation selective channels with diverse modes of activation and varied permeation properties (reviewed by [314]). Established, or potential, physiological functions of the individual members of the TRP families are discussed in detail in the recommended reviews and a compilation edited by Islam [271]. The established, or potential, involvement of TRP channels in disease is reviewed in [279,307] and [308], together with a special edition of Biochemica et Biophysica Acta on the subject [307]. The pharmacology of most TRP channels is poorly developed [351]. Broad spectrum agents are listed in the tables along with more selective, or recently recognised, ligands that are flagged by the inclusion of a primary reference. Most TRP channels are regulated by phosphoinostides such as PtIns(4,5)P2 and IP3 although the effects reported are often complex, occasionally contradictory, and likely be dependent upon experimental conditions (reviewed by [309,327,343]). Such regulation is generally not included in the tables.
TRPA (ankyrin) family TRPA1 is the sole mammalian member of this group (reviewed by [259]). In some [234,274,330,334], but not other [272,303], studies TRPA1 is activated by noxious cold.
One study suggests that activation of TRPA1 is secondary to a cold-induced elevation of [Ca 2+ ]i [357], but this has been refuted [274]. Additionally, TRPA1 has been proposed to be a component of a mechanosensitive transduction channel of vertebrate hair cells [246,303], but TRPA1 (-/-) mice demonstrate no impairment in hearing, or vestibular function [238,284]. There is consensus that TRPA1 acts as a nociceptor for environmental irritants [235].

TRPC (canonical) family
Members of the TRPC subfamily (reviewed by [229][230][241][242]258,277,316,324]) fall into the subgroups outlined below. TRPC2 (not tabulated) is a pseudogene in man. It is generally accepted that all TRPC channels are activated downstream of Gq/11-coupled receptors, or receptor tyrosine kinases (reviewed by [321,337,351]). A comprehensive listing of G-protein coupled receptors that activate TRPC channels is given in [229]. Hetero-oligomeric complexes of TRPC channels and their association with proteins to form signalling complexes are detailed in [230] and [278]. TRPC channels have frequently been proposed to act as store-operated channels (SOCs) (or compenents of mulimeric complexes that form SOCs), activated by depletion of intracellular calcium stores (reviewed by [230,243,317,322,329,354]), but this is controversial. All members of the TRPC family are blocked by 2-APB and SKF96356 [265][266]. Activation of TRPC channels by lipids is discussed by [241].
TRPC1/C4/C5 subgroup TRPC4/C5 may be distinguished from other TRP channels by their potentiation by micromolar concentrations of La 3+ .
TRPC3/C6/C7 subgroup All members are activated by diacylglycerol independent of protein kinase C stimulation [266].

TRPM (melastatin) family
Members of the TRPM subfamily (reviewed by [257,265,317,356]) fall into the five subgroups outlined below.
TRPM1/M3 subgroup TRPM1 exists as five splice variants and is involved in normal melanocyte pigmentation [312] and is also a visual transduction channel in retinal ON bipolar cells [283]. TRPM3 (reviewed by [313]) exists as multiple splice variants four of which (mTRPM3α1, mTRPM3α2, hTRPM3a and hTRPM31325) have been characterised and found to differ significantly in their biophysical properties. TRPM3 has recently been found to contribute to the detection of noxious heat [346].
TRPM2 TRPM2 functions as a sensor of redox status in cells and is also activated by heat (reviewed by [353]). Numerous splice variants of TRPM2 exist which differ in their activation mechanisms [254].
TRPM4/5 subgroup TRPM4 and TRPM5 are thermosensitive and have the distinction within all TRP channels of being impermeable to Ca 2+ [351]. A splice variant of TRPM4 (i.e.TRPM4b) and TRPM5 are molecular candidates for endogenous calciumactivated cation (CAN) channels [262]. TRPM4 has been shown to be an important regulator of Ca 2+ entry in to mast cells [339] and dendritic cell migration [236]. TRPM5 in taste receptor cells of the tongue appears essential for the transduction of sweet, amino acid and bitter stimuli [289].
TRPM8 Is a channel activated by cooling and pharmacological agents evoking a 'cool' sensation and participates in the thermosensation of cold temperatures [240,245,252] reviewed by [282,291,302,344].

TRPML (mucolipin) family
The TRPML family [323,325,355] consists of three mammalian members (TRPML1-3). TRPML channels are probably restricted to intracellular vesicles and mutations in the gene (MCOLN1) encoding TRPML1 (mucolipin-1) are the cause of the neurodegenerative disorder mucolipidosis type IV (MLIV) in man. TRPML1 is a cation selective ion channel that is important for sorting/transport of endosomes in the late endocytotic pathway and specifically fusion between late endosome-lysosome hybrid vesicles. TRPML2 (MCLN2) remains to be functionally characterised in detail. TRPML3 is important for hair cell maturation, stereocilia maturation and intracellular vesicle transport. A naturally occurring gain of function mutation in TRPML3 (i.e. A419P) results in the varitint waddler (Va) mouse phenotype (reviewed by [310,325]).

TRPP (polycystin) family
The TRPP family (reviewed by [249,251,260,268,349]) subsumes the polycystins that are divided into two structurally distinct groups, polycystic kidney disease 1-like (PKD1-like) and polycystic kidney disease 2-like (PKD2-like). Members of the PKD1-like group, in mammals, include PKD1 (reclassified as TRPP1), PDKREJ, PKD1L1, PKD1L2 and PKD1L3. The PKD2-like members comprise PKD2, PKD2L1 and PKD2L2, which have renamed TRPP2, TRPP3 and TRPP5, respectively [300]. PKDREJ (Q9NTG1), PKD1L1 (Q8TDX9), mouse PKD1L2 (Q7TN88), PKD1L3 (Q7Z443) and TRPP5 (PKD2L2, Q9NZM6) are not listed in the table due to lack of functional data. Similarly, TRPP1 (P98161) is also omitted because although one study [233] has reported the induction of a cation conductance in CHO cells transfected with TRPP1, there is no unequivocal evidence that TRPP1 is a channel per se and in other studies (e.g. [250,264]) TRPP1 is incapable of producing currents.  [320,333,335]). Numerous splice variants of TRPV1 have been described, some of which modulate the activity of TRPV1, or act in a dominant negative manner when co-expressed with TRPV1 [331]. The pharmacology of TRPV1 channels is discussed in detail in [263] and [345]. TRPV2 is probably not a thermosensor in man [315], but has recently been implicated in innate immunity [290]. TRPV3 and TRPV4 are both thermosensitive, with the latter also having a mechanosensing function [255].

Subunits
Channel Blockers (IC50) -2-APB, Gd 3+ , La 3+ , mefenamic acid [281], pioglitazone (independent of PPAR-γ) [295], rosiglitazone, troglitazone  antagonists produce depolarizing shifts in V½ [301]. The V½ for the native channel is far more positive than that of heterologously expressed TRPM8 [301]. It should be noted that (-)menthol and structurally related compounds can elicit release of Ca 2+ from the endoplasmic reticulum independent of activation of TRPM8 [293]. Intracellular pH modulates activation of TRPM8 by cold and icilin, but not (-)-menthol [231]. Comments: Agents activating TRPA1 in a covalent manner are thiol reactive electrophiles that bind to cysteine and lysine residues within the cytoplasmic domain of the channel [267,292]. TRPA1 is activated by a wide range of endogenous and exogenous compounds and only a few representative examples are mentioned in the table: an exhaustive listing can be found in [235]. In addition, TRPA1 is potently activated by intracellular zinc (EC50 = 8 nM) [232,269].    [247,251] and [332]. Broadly similar single channel conductance, mono-and di-valent cation selectivity and sensitivity to blockers are observed for TRPP2 co-expressed with TRPP1 [250]. Ca 2+ , Ba 2+ and Sr 2+ permeate TRPP3, but reduce inward currents carried by Na + . Mg 2+ is largely impermeant and exerts a voltage dependent inhibition that increases with hyperpolarization.

Voltage-gated calcium channels
Overview: Calcium (Ca 2+ ) channels are voltage-gated ion channels present in the membrane of most excitable cells. The nomenclature for Ca 2+ channels was proposed by [359] and approved by the NC-IUPHAR subcommittee on Ca 2+ channels [358]. Ca 2+ 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 dihydropyridineinsensitive (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 membranespanning 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 α1s. The α2-δ1 and α2-δ2 subunits bind gabapentin and pregabalin.

Voltage-gated proton channel
Overview: The voltage-gated proton channel (provisionally denoted Hv1) is a putative 4TM proton-selective channel gated by membrane depolarization and which is sensitive to the transmembrane pH gradient [361][362][363]372,374]. The structure of Hv1 is homologous to the voltage sensing domain (VSD) of the superfamily of voltage-gated ion channels (i.e. segments S1 to S4) and contains no discernable pore region [372,374]. Proton flux through Hv1 is instead most likely mediated by a water wire completed in a crevice of the protein when the voltage-sensing S4 helix moves in response to a change in transmembrane potential [371,377]. Hv1 expresses largely as a dimer mediated by intracellular C-terminal coiled-coil interactions [367]  Activated by membrane depolarization mediating macroscopic currents with time-, voltage-and pH-dependence; outwardly rectifying; voltage dependent kinetics with relatively slow current activation sensitive to extracellular pH and temperature, relatively fast deactivation; voltage threshold for current activation determined by pH gradient (ΔpH = pHo -pHi) across the membrane

Comments:
The voltage threshold (Vthr) for activation of Hv1 is not fixed but is set by the pH gradient across the membrane such that Vthr is positive to the Nernst potential for H + , which ensures that only outwardly directed flux of H + occurs under physiological conditions [361][362][363]. Phosphorylation of Hv1 within the N-terminal domain by PKC enhances the gating of the channel [368]. Tabulated IC50 values for Zn 2 + and Cd 2+ are for heterologously expressed human and mouse Hv1 [372,374]. Zn 2+ is not a conventional pore blocker, but is coordinated by two, or more, external protonation sites involving histamine residues [372]. Zn 2+ binding may occur at the dimer interface between pairs of histamine residues from both monomers where it may interfere with channel opening [369]. Mouse knockout studies demonstrate that Hv1 participates in charge compensation in granulocytes during the respiratory burst of NADPH oxidase-dependent reactive oxygen species production that assists in the clearance of bacterial pathogens [373]. Additional physiological functions of Hv1 are reviewed by [361].

Voltage-gated sodium channels
Overview: Sodium channels are voltage-gated sodium-selective ion channels present in the membrane of most excitable cells. Sodium channels comprise of one pore-forming α subunit, which may be associated with either one or two β subunits [380]. α-Subunits consist of four homologous domains (I-IV), each containing six transmembrane segments (S1-S6) and a pore-forming loop. The positively charged fourth transmembrane segment (S4) acts as a voltage sensor and is involved in channel gating. The crystal structure of the bacterial NavAb channel has revealed a number of novel structural features compared to earlier potassium channel structures including a short selectivity filter with ion selectivity determined by interactions with glutamate side chains [381]. Interestingly, the pore region is penetrated by fatty acyl chains that extend into the central cavity which may allow the entry of small, hydrophobic pore-blocking drugs [381]. Auxiliary β1, β2, β3 and β4 subunits consist of a large extracellular N-terminal domain, a single transmembrane segment and a shorter cytoplasmic domain.
The nomenclature for sodium channels was proposed by Goldin et al., (2000) [379] and approved by the NC-IUPHAR subcommittee on sodium channels (Catterall et al., 2005, [378]). Comments: Sodium channels are also blocked by local anaesthetic agents, antiarrythmic drugs and antiepileptic drugs. There are two clear functional fingerprints for distinguishing different subtypes. These are sensitivity to tetrodotoxin (NaV1.5, NaV1.8 and NaV1.9 are much less sensitive to block) and rate of inactivation (NaV1.8 and particularly NaV1.9 inactivate more slowly).