Abstract
In the Nav channel family the lipophilic drugs/toxins binding sites and the presence of fenestrations in the channel pore wall are well defined and categorized. No such classification exists in the much larger Kv channel family, although certain lipophilic compounds seem to deviate from binding to well-known hydrophilic binding sites. By mapping different compound binding sites onto 3D structures of Kv channels, there appear to be three distinct lipid-exposed binding sites preserved in Kv channels: the front and back side of the pore domain, and S2-S3/S3-S4 clefts. One or a combination of these sites is most likely the orthologous equivalent of neurotoxin site 5 in Nav channels. This review describes the different lipophilic binding sites and location of pore wall fenestrations within the Kv channel family and compares it to the knowledge of Nav channels.
Introduction
Voltage-gated ion channels are transmembrane proteins that are selectively permeable to physiological important ions such as Na+, K+, Ca2+, and Cl-. Under influence of the membrane potential (Vm) these channels change their conductance. In the conductive open (or activated) state, ions flow down their electrochemical gradient through the channel pore. The flux of these ions elicits an electrical current that directly influences the Vm. For voltage-gated sodium and potassium channels (Nav and Kv), the main focus of this review, the Vm will shift towards the ion's equilibrium potential, which under normal conditions is depolarizing and repolarizing, respectively (). Although Nav and Kv channels differ in selectivity from one another, their structure is quite similar. However, a main difference is that Nav channels are characterized by one large α-subunit containing four recognizable domains (DI–IV), whereas Kv channels are formed by tetramerization of four individual α-subunits. In both cases, these four entities comprise six transmembrane segments (S1–S6), which are divided into a voltage sensing domain (VSD, S1–S4) and a pore-forming domain (PD, S5–S6) that are connected by the S4–S5 linker. The four PDs assemble into the ion permeation pathway (or pore) that is surrounded by four VSDs. In the non-conductive closed (or deactivated) state, ion permeation is prevented by the intracellular activation gate, located at the point where the four S6 helices cross. The aperture-like opening and closure of this gate is controlled by the VSD (; ; ; ). The main component of the VSD is the S4 segment that physically moves in response to a change in Vm, due to the presence of positively charged residues (arginine and lysine) that detect changes in the membrane electric field (). The S4–S5 linker is a component of the electro-mechanical coupling that translates S4 movements into opening or closing of the activation gate (; ). After opening, fast inactivation occurs in Nav and in some Kv channels, which is caused by the physical occlusion of the pore by an inactivation particle. For Nav channels this inactivation particle is the linker between DIII and DIV, while in Kv channels it is located at the N-terminus of each subunit, hence termed N-type inactivation (; West et al., 1992; ). Alternatively, Kv channel inactivation can occur via the slower C- or U-type inactivation mechanism that makes the channels non or less conductive (; ; ).
Binding of drugs/toxins to Nav and Kv channels may alter the activation, deactivation, and/or inactivation process(es), which may cause or alleviate aberrant electrical excitability. Therefore, knowledge about the different binding sites is key for drug development and pharmacovigilance. The binding sites for these drugs/toxins are well defined and categorized within the Nav channel family, as opposed to the much larger Kv channel family. Most binding sites are enveloped by water, locating either inside or outside the channel's pore. However, some compounds bind to a site(s) that does not fit any of the hydrophilic binding sites. For instance, brevetoxins and ciguatoxins bind to a conserved hydrophobic site within the Nav channel family, termed neurotoxin site 5 (; ). For the Kv channel family no such site has been described, but certain compounds have been shown to deviate from binding to hydrophilic binding sites like; retigabine, gambierol, psora-4, polyunsaturated fatty acids (PUFAs), ICA-compounds (Vennekamp et al., 2004; ; ; ). It is notable that these are rather lipophilic compounds and there has been, and still is, a growing interest in such compounds for their use in treating neurological disorders (e.g., as anti-convulsant).
So, is there a unifying picture of the lipid exposed/accessible drug/toxin binding sites within the large Kv channel family and even between Kv and Nav channels? Several lipophilic binding sites have been described in different Kv channels, while in fact some may converge to just one binding region preserved between Kv channel (sub)families. In this review, the well-documented Nav lipophilic binding sites, neurotoxin site 2, site 5, and the access to the local anaesthetic (LA) binding site within the pore through fenestrations is compared to what has been reported for Kv channels.
Voltage-Gated Sodium Channels
The Nav channel family contains nine isoforms (Nav1.1 to Nav1.9) that display a high sequence homology, especially within the transmembrane segments (; ). This facilitated the categorization of drug/toxin binding sites within the Nav channel family. Over the past decades a detailed picture emerged on where compounds bind within these channels and resulted in a well-documented classification of seven different sites (site 1 to 7) and a LA binding site (Stevens et al., 2011; ). As the focus of this review is on the binding sites that involve lipid soluble and/or transmembrane binding compounds only binding site 5 and site 2 will be briefly discussed. The LA binding site is also mentioned as some compounds can reach their binding site via hydrophobic fenestrations in the pore wall of the channel protein. To maintain an orderly overview, all Nav residues are numbered according to the Nav1.4 channel when possible. In case the sequence could not be aligned, as for bacterial Nav structures, it will be noted and the original numbering is maintained.
The Closed State Accessible LA Binding Site: Pore-Accessibility Through Channel Fenestrations
LA compounds and anti-arrhythmic drugs inhibit Nav channels by occlusion of the pore. Most LA compounds have a similar structure consisting of a tertiary hydrophilic amine domain (head) linked with an aromatic hydrophobic ring domain, with a total length of 10–15Å (). Three types of block can be observed. First type is the use dependent open state block, or high affinity block, which occurs after channel opening and LA compounds enter the pore via the intracellular side (; ; ). The second type is flicker block, or fast block, which is only observed when the channel's inactivation process is modulated. For example, by other compounds such as batrachotoxin (BTX) (CAS No.:23509-16-2) that binds at site 2 (; Uehara and Moczydlowski, 1986; Zamponi et al., 1993). The third and least common type is the resting or hydrophilic block that establishes when the channel is closed. LA compounds are thought to enter the ion conductive pore and find their binding site trough fenestrations in the lipid exposed part of the PD (). LA compounds are protonated but for crossing the cell membrane they need to be deprotonated. This is possible when their pKA value is close to the physiological pH values in the extracellular environment. After traversing the cell membrane the compounds are protonated again to exert their pharmacological effect (). As the binding site is located within the pore, block occurs with the highest affinity when the channel is opened or used, while applying frequent channel activating stimuli. Two residues within the S6 of domain four (DIV-S6) have been identified to be important for LA binding/modulation, namely, F1586 and Y1593, (Nav1.4 numbering). F1586 probably binds to the alkylamino head, while Y1593 interacts with the aromatic ring structure (Ragsdale et al., 1994; Yarov-Yarovoy et al., 2002) (Figures 1A, B). As these residues reside within the pore and certain compounds display closed state block, this requires a hydrophobic pathway to the binding site when the channel gate is closed ().
Figure 1
These pathways, or fenestrations, in the lipid exposed part of the PD were first observed in the crystal structure of bacterial Nav channels such as NavAb (Payandeh et al., 2011; Payandeh et al., 2012), NavMs () and NavRh (Zhang et al., 2012). It should be noted that bacterial Nav channels are constructed out of four separate α-subunits instead of one large subunit. For the bacterial Nav channels, four fenestrations are observed between the different domains where the radius of the fenestration varies from 0.8Å minimally to 2.59-2.83Å (). Nonetheless, these fenestrations are wide enough for small LA compounds and anti-arrhythmic drugs to pass. The narrowest point in the fenestration, termed “bottleneck”, is created by the amino acid residues M174, T175, F203, T206, and M209 (NavAb numbering), with F203 being the most important residue (Figure 1C). These amino acids will sterically hinder the passage of compounds through the fenestration. Mutation of F203 to an alanine increased the size of the fenestration allowing easier access of flecainide (CAS No.:54143-55-4, polar surface area=59.6Å) to its binding site within the pore, with as result an increased tonic, closed state, block (). The mutation did not affect lidocaine block as this compound is smaller (polar surface area=32.3Å) and can easily traverse the wild type fenestration (). The F203W mutation on the other hand reduces fenestration size and consequently the access of both lidocaine and flecainide is reduced, decreasing tonic block (). Computational modelling using the Nav1.4 3D structure resulted in the observation of four fenestrations just as in bacterial Nav channels, with the exception that the size of two out of the four fenestrations seemed inadequate for compound access, namely the fenestration constructed by DII–DIII and DIV–DI (). Fenestrations in between DI-DII and DIII-DIV seemed sufficiently wide for compounds to cross (Figure 1C). The bottleneck of these fenestrations is formed by residues N790, L438, and M786 for the fenestration between DI-DII and by I1582, I1583, and F1586 (if rotated) for the fenestration between DIII-DIV. These fenestrations lining residues, are well conserved in the different Nav channel isoforms. Mutating DIV-S6 residue I1582 appeared to create an extra pore access pathway (Ragsdale et al., 1994) and allowed the external blocker QX314 (CAS No.:24003-58-5, polar surface area: 29.1Å), which is a charged LA compound at physiological pH and therefore not able to traverse the membrane, to access the LA binding site when added in the extracellular environment (Sunami et al., 2001).
Apart from LAs that have a distinct binding site, sevoflurane (CAS No.: 28523-86-6), an inhalational anaesthetic, has a more complex binding profile. Sevoflurane binding results in a decrease of the peak sodium current, a hyperpolarised shift in the voltage dependence of inactivation and a slowing of the recovery from inactivation (; Ouyang et al., 2009). Within the bacterial sodium channel NaChBac, binding regions have been located at the pore region, selectivity filter, and the S4–S5 linker/S6 interface (Ouyang et al., 2007; Raju et al., 2013). MD simulations suggested that the binding of sevoflurane to the selectivity filter and the S4–S5 linker occurs mainly when the channel is in the activated/open state, while in the closed state the channel gate and the VSD are targeted (). A state independent binding site is possibly the central cavity that is accessed by the fenestrations. Residues T220 and F227 (NavChBac numbering) are proposed to be responsible for sevoflurane binding in the central cavity, which are the homologue residues for LA binding in Nav1.4 (F1764, Y1771).
Binding Site 2 Compounds, Though Being Lipophilic, Bind to the Inner Pore
Compounds binding at site 2 of the Nav channel include batrachotoxin (; ), grayanotoxin (Yuki et al., 2001; ), CAS No.: 54781-61-2), and alkaloids from plant such as veratridine (Ulbricht, 1969; Sutro, 1986), CAS No.:71-62-5), aconitine and mesaconitine (; ) (CAS No.: 302-27-2). All these compounds are lipid soluble and need to traverse the cell membrane before influencing the channel in a use-dependent manner, comparable to the action of LA compounds (; ; ; ; Sutro, 1986; ; ; Yuki et al., 2001; Wang and Wang, 2003). Their effects can be a combination of: (1) a hyperpolarised shift of the voltage dependence of activation, (2) inhibition of the fast inactivation process, leading to persistent sodium currents, (3) decrease in ion conductance, and/or (4) decrease of the Na+ selectivity. The location of binding site 2 is thought to be at the S6 of all four domains and to overlap with the LA binding site or at least allosterically hinder LA binding.
To describe neurotoxin site 2 in more detail, we focus on the most potent site 2 toxin reported to date, batrachotoxin (BTX) a steroidal alkaloid indirectly produced by the South-American poison dart frogs of the genus Phyllobates (). Binding of BTX is highly irreversible and only possible when the channel is in its activated open state (; ). BTX shifts the voltage dependence of channel activation by approximately −40 mV toward more hyperpolarized potentials (), reducing the channel's ion conductance and selectivity. BTX also reduces the affinity for LA, which is thought to be caused by non-competitive antagonism (i.e., allosteric effect) because of overlapping binding sites as BTX and LA share the binding residue F1586 (). Residue mutations affecting BTX in Nav1.4 were first described in DI-S6 and DIV-S6: I439K, N440K, L443K, F1586K, and N1591K, respectively (Wang and Wang, 1998; Wang and Wang, 1999). Afterwards the DIII-S6 residues Ser1283 and leu1287 were identified (Wang et al., 2000) and finally DII-S6 residues N790 and L794 (Wang et al., 2001). All these residues line the inside of the pore (Figure 2). Non-pore lining residues involved in BTX sensitivity locate at the putative hinge region of the channel gate, Gly1282 and Phe1284, respectively (). While Phe1284 is important for the stabilisation of the ammonium group of BTX, Gly1282 is not a direct binding receptor for BTX and mutations of this residue affect binding allosterically by changing the channels gating properties. Modeling studies suggest that BTX binds to the pore but does not completely prevent ion conduction because of its “horseshoe-like” structure (). Only upon mutation of DII-S6 N790 BTX becomes a full blocker (Wang et al., 2007).
Figure 2
Binding Site 5 Locates Between DIS6 and DIVS5
Some of the toxins that bind to site 5 of Nav channels are brevetoxins (; Poli et al., 1986) and ciguatoxins (; ), both originate from marine dinoflagellates (Gambierdiscus toxicus) and are structurally comparable. Brevetoxins are produced by unarmoured marine dinoflagellates (e.g., Karenia brevis, Gymnodinium breve, or Ptychodiscus brevis). Ingestion of these toxins can lead to poisoning and death of marine animals and cause the disease neurotoxic shellfish poisoning in humans (; ). Eleven different brevetoxins have been discovered today with brevetoxin A (PbTX1, CAS No.:98112-41-5) and brevetoxin B (PbTX2 and 3, CAS No.:79580-28-2 and 85079-48-7) being the most investigated ones. These toxins are about 30Å in length, 6Å in width, and 6Å high. They are composed of four recognizable parts: a lactone ring (the head) linked with a linker to a multiple carbon ring (the tail) ending in a rest-group (). While the head is responsible for the effect, without the linker and the tail the toxin cannot modulate the channel as it allows the head region to reach its binding site (Rein et al., 1994). PbTX binds to the Nav channel at the DI-S6, DIV-S5, and DIV-S6 segments (Trainer et al., 1991; Trainer et al., 1994; ). The region spans within DI-S6 from Ala418/Thr422 to Lys465/Lys477 and within DIV-S5 from Glu1510 to lys1557 with possible extension to Lys1565 (Figure 3). The tail of the molecule will be situated at the S5–S6 extracellular loop while the head is able to reach, due to the long linker and tail region, the inactivation gate at the intracellular side of the channel (Trainer et al., 1991; Trainer et al., 1994). Interaction of the head with the inactivation gate reduces channel inactivation, leading to a persistent sodium current (Sheridan and Adler, 1989; Schreibmayer and Jeglitsch, 1992).
Figure 3
Like brevetoxins, ciguatoxins (CTX) bind at site 5 and their effects are consequently similar. Ciguatoxins are classified based on their geographic origin, with P-CTX () and C-CTX () standing for Pacific and Caribbean, respectively. In all Nav channel isoforms P-CTX1 (CAS No.:11050 21 8), the most potent CTX discovered to date, induces a hyperpolarizing shift in the voltage dependence of channel activation (; ; Strachan et al., 1999) concomitantly with a shift in the voltage dependence of inactivation in some (). As CTX and brevetoxin share the same binding site, they are logically going into competition with each other (). Gambierol (CAS No.: 146763-62-4, origin Gambierdiscus Toxicus) has a similar structure as brevetoxins and CTXs, being a lipophilic multi-ring polyether toxin, but has no effect on the sodium currents (). However, when administered simultaneously it decreases the effect of CTX and brevetoxin, suggesting that gambierol acts as a competitive antagonist that binds to site 5 or at least exerts a negative allosteric effect.
Voltage-Gated Potassium Channels
The Kv channel family is impressively large compared to this of Nav channels, due to an extensive library of genes encoding different α-subunits that in some subfamilies can “mix-and-match” to form functional Kv channels. Additionally, alternative splicing, RNA editing, and post-translational modification further expand on the Kv channel family (). This explains why the binding sites of the much larger Kv channel family are less well categorized than those of the Nav channel family. The next part will highlight different binding sites and pore wall fenestrations within several, but not all, members of the Kv channel family. After a short overview of the well-documented extracellular and intracellular exposed “hydrophilic” binding sites, we will discuss in detail the lesser-known lipid embedded “hydrophobic” binding site(s) in different Kv channels.
Extracellular and Intracellular Exposed “Hydrophilic” Binding Sites
The well-documented Kv channel binding sites can be topologically located on the intracellular or extracellular side of the channels. Extracellularly the most well-known conserved binding sites are those of the pore blockers and VSD targeting gating modifiers, while intracellularly the inner pore block is the most notable one (Wulff et al., 2009). Certain toxins from a variety of venomous animals target the extracellular binding sites. For example, scorpion toxins like charybdotoxin (CTX, CAS No.: 95751-30-7) and agitoxin (AgTx, CAS No.: 168147-41-9) target the water enveloped extracellular mouth of Kv channels, thereby physically occluding the permeation pore (; ). Other scorpion toxins, like ergotoxin (ErgTx1, CAS No.: 8006-25-5) and BeKm-1 (CAS No.: 524962-01-4), cause an incomplete block of Kv current. This is because they only partially occlude the permeation pathway, which is known as “turret-block” (Xu et al., 2003; Zhang et al., 2003; ). Gating modifier toxins, like tarantula toxins (Hanatoxin, SGTx1, and VSTx1), bind to the paddle motif (S3b helices, S3–S4 linkers, and S4 helices) at the extracellular protein-lipid interface of the VSD. This binding site cannot be characterized as strictly hydrophilic as the hydrophobic residues of these amphipathic peptide toxins allow it to partially partition into the membrane, and interact with the VSD (; ; ; Swartz, 2007; ; ). Intracellularly, compounds can bind in the inner cavity, either physically blocking ion permeation (e.g., quaternary ammonium ions) or allosterically modulating the gating machinery of Kv channels (e.g., 4-aminopyridine, CAS No.: 504-24-5) (; ; ). Also, the quaternary ammonium ion TEA can occlude the external Kv pore, similar to the scorpion toxins mentioned above (). Apart from the gating modifier binding site, which is partially enveloped by water, the binding sites mentioned here are exposed to an aqueous environment. Hence, referring them here as “hydrophilic” binding sites.
Lipid Embedded “Hydrophobic” Binding Site(s) and Pore Wall Fenestrations in the Shaker-Type Kv Family, Kv7 Family, and Kv10.1/Kv11.1 Channel
The next part reviews the potential lipophilic binding sites and pore wall fenestrations in different Kv channel types. To define the binding site(s) of certain compounds, the PD is divided into a “front side” and “back side”. Residues of the front side point towards the VSD of the same α-subunit, while residues of the “back side” point in the opposite direction, thus towards the VSD of a neighboring α-subunit (Figures 4D, 5D and 6C).
Figure 4
Figure 5
The Shaker-Type Kv Channel Family
Whereas gambierol does not modulate Nav channels, it is capable of inhibiting Kv1 and Kv3 channels (; ; ). Gambierol's inhibitory mechanism has been extensively studied, whereby a threonine on S6, T427 in Kv3.1, is an important determinant (). Substitution of the polar threonine by a hydrophobic valine abolishes the high gambierol affinity (). Additional determinants are a leucine and phenylalanine on S5: L348 and F351 in Kv3.1, respectively. Kv1 channels possess a threonine residue equivalent to T427 (T401 in Kv1.2), explaining their similar gambierol sensitivity (; ; ). These residues are mostly positioned on the front side of the PD (Figure 4).
On the other hand, psora-4 (CAS No.: 724709-68-6), a potent inhibitor of Kv1.3, has been shown to predominantly bind to the back side of the PD (Figure 4) (Vennekamp et al., 2004; ). A single psora-4 molecule acts as a central pore blocker of Kv1 channels (Kv1.1-Kv1.5, and Kv1.7), thereby preventing ions from permeating. However, four additional drug molecules can bind the lipid-exposed pocket on the back side of the PD, thereby causing the selectivity filter to narrow. Thus, the binding of five psora-4 molecules leads to a stable non-conducting state. The residues identified in Kv1.5 as playing a key role in psora-4 action map on the back side of S5–S6 (Figure 4). Additionally, some residues in S4 and the S4–S5 linker also seem involved ().
The inhalation anaesthetic, sevoflurane, seems to bind within the central cavity and to a similar hydrophobic pocket as psora-4 (Stock et al., 2018). Apart from these binding regions, the polar lipophilic molecule has been shown to interact with the S4–S5 linker, pore helix, segment S6, and even the VSD, likewise to the many binding regions of sevoflurane on the Nav channel (; ). These sites are primarily dehydrated and lipid accessible, which is highly favourable for the polar lipophilic sevoflurane molecule (Stock et al., 2018). The specific residues involved in sevoflurane binding are not known, but one residue within the S4–S5 linker (G329 according to Kv1.2 numbering) has been identified to play an important role ().
PUFAs are charged lipophilic compounds that position at the lipid membrane interface, although in small quantities (Yazdi et al., 2016). Nonetheless, PUFAs may play an important role in the treatment of arrhythmias and epilepsy, due to their modulating effect on, but not limited to, voltage-gated ion channels (; ; ; ). The Shaker Kv channel has been extensively studied as one of the targets of PUFAs. They interact with the channel at several sites, but the major one seems located within the lipid-facing cleft between S3 and S4 (S3–S4 cleft) of the VSD (). This hydrophobic cleft is perfectly shaped to accommodate the lipophilic carbon tail of the PUFAs, causing the negatively charged carboxyl head group to be positioned close to the S4 segment. In this way, PUFAs electrostatically affect the VSD by trapping S4 toward the extracellular position, stabilizing the open state of Shaker Kv channels (; Yazdi et al., 2016). Accordingly, a series of point mutations on the lipid facing side of S3-S4 (I325C, T329C, A359C, and I360C, Shaker numbering) had a significant impact on the PUFA-induced hyperpolarizing shift in the channel's voltage dependence of activation (Figure 4) (). Although the binding site for gating modifier toxins is in close proximity to that of PUFAs, the action sites do most likely not overlap as the residues important for PUFA action are more deeply embedded into the lipid bilayer (). Interestingly, dehydroabietic acid (DHAA) and some of its derivatives, the most potent being Wu32and Wu122, have a similar effect on Shaker Kv channels. The carboxyl group of DHAA is positioned at roughly the same site, namely the S3–S4 cleft (Ottosson et al., 2017). Wu32 possibly interacts with residues between the S2–S3 and/or S3–S4 cleft as five cysteine mutations in S3, which have been shown to alter its affinity and/or efficacy, point towards S2 (I320C and F324C, shaker numbering) and S4 (I318C, P322C, and T326C), respectively (Ottosson et al., 2017).
The Kv7 Family
The well characterized retigabine (RTG, CAS No.: 150812-12-7) binding site is localized between the front side of one PD and the back side of an adjacent PD of Kv7.2–Kv7.5 (KCNQ2-5) channels (Figure 5) (Schenzer et al., 2005; ). These channels are predominantly expressed in neurons where they underlie the native M-current that plays a major role in regulating neuronal excitability (Wang et al., 1998; ; Schroeder et al., 2000). RTG amplifies the Kv7.2–Kv7.5 currents by stabilizing the open-channel conformation, by which it acts as a brake on neuronal excitability in vivo (). Hence, the potential use of RTG and RTG-derived compounds as anticonvulsants (Rundfeldt, 1997; Wang et al., 2018). RTG binding has been attributed to several conserved residues on S5–S6, lining a hydrophobic pocket near the channel gate of Kv7.2–Kv7.5. According to Kv7.2 numbering these residues are: W236, L243, L275, L299, and G301 (Figure 5). Kv7.1 channels lack these amino acids (apart from L243), explaining their RTG-insensitivity (Schenzer et al., 2005; ). Because of the role of Kv7 in diseases of neuronal hyperexcitability, the search for positive allosteric modulators such as RTG, for which the clinical use is currently discontinued (Wang et al., 2018), is pursued. For example, BMS-204352, ML-213, and the acrylamide compound (S)-1 act on the canonical RTG binding site (; ; Wang et al., 2018).
The hydrophobic pocket of the RTG binding site also seems to be able to accommodate endogenous hydrophilic neurotransmitters like γ-aminobutyric acid (GABA), which directly activates Kv7.3 and Kv7.5 via W236. In contrast to RTG, GABA does not readily cross the plasma membrane to reach its site of action. Based on the Kv1.2–Kv2.1 paddle chimera structure the tryptophan also seems to be accessible from the extracellular side (). It is possible that this accessibility is dependent of the state of the channel, such that in certain conformations the mostly hydrophobic binding pocket can be reached by hydrophilic compounds like GABA. Additionally, not all Kv7 channel openers interact with residue W236. Zinc pyrithione for instance interacts with two residues at the back side of S5 and the pore helix (L249 and L275) (Figure 5) (Schenzer et al., 2005; Xiong et al., 2007; ).
ICA-compounds (ICAgen, Durham, NC, US), which are benzanilide Kv7 channel openers, were developed as RTG alternatives (). Of these, the most well-documented is ICA-27243 (ICA43), which has been shown to be more selective than RTG (Wickenden et al., 2008). The individual residues that determine ICA43 binding have not been identified, but the C-terminal end of S2 and the N-terminal part of S3 (S2–S3 cleft) are proposed to be involved (Padilla et al., 2009). Later, Wang AW, et al. continued the investigation of the mechanism of action of ICA-compounds on Kv7 channels, but focussed on ICA-069673 (ICA73) (Wang et al., 2017). They identified two key residues in S3 of Kv7.2: A181 and F168, respectively. Mutation of these residues did not affect RTG-mediated gating, but did alter the action of ICA73 (Wang et al., 2017). Furthermore, it has been shown that ICA43 and ICA73 are resistant to mutation of the RTG binding site, supporting that not all Kv7 channel openers bind to the PD, but also can interact with a VSD site (Padilla et al., 2009; Wang et al., 2017). However, it remains debated whether residues A181 and F168 are involved in ICA binding directly or allosterically (Wang et al., 2018). In case these residues are binding ICA73, the position of residue A181 toward the lipid-exposed surface of the VSD suggests the presence of a drug binding site at the lipid-exposed cleft of S2–S3 and/or S3–S4 in Kv7 channels, similar to the interaction of Wu32 with Shaker Kv channels (Figure 5).
PUFAs have also been described to electrostatically affect Kv7.1 channels, resulting in a negative shift of the conductance-voltage curve. This modulation of Kv7.1 channels by PUFAs is similar to what has been described for shaker Kv channels (; ). Hence, the binding site and mechanism of action of PUFAs are most likely similar for both channels ().
The hERG (Kv11.1) and EAG (Kv10.1) Channel
The human ether-a-go-go related gene (hERG) type 1 encodes for the Kv11.1 channel, which functions as the rapid component of the delayed rectifier K+ current contributing to the repolarization of cardiac action potentials (Vandenberg et al., 2012). Alteration of the native functioning of Kv11.1 channels, either genetically or pharmacologically, can disrupt this repolarization, leading to various cardiac rhythm disorders (Thomas et al., 2006). Furthermore, the role of Kv11.1 is not limited to the heart, as it also seems to play a role in the central nervous system, digestive, secretory, and reproductive system, and even cancer (). Hence, it has become common practice to screen compounds on hERG channel activity during the early stages of drug development, as unintentional side-effects may lead to disease and sudden-death (; Thomas et al., 2006; ). Most hERG inhibitors interact with residues inside the channel's permeation pathway, either located on the pore helix (T623,S624, and V625, according to hERG numbering) or on segment S6 (G648, Y652, and F656) (; ; ; Sanchez-Chapula et al., 2002; Sanchez-Chapula et al., 2003; ; ). On the other hand, some small molecule hERG activators have been discovered who deviate from this binding site, like ICA-105574 (ICA74) and PD-118057 (PD57). The most critical binding determinants for ICA74 are F557 and L622 (), which topologically would situate the binding site at the front side of the PD (Figure 6). Apart from these critical binding determinants several other residues have been proposed to line the hydrophobic ICA binding pocket of Kv11.1 (F619, T623, M645, L646, M651, Y652, and F656), situating the ICA74 binding site between the front side of one α-subunit and the back side of an adjacent α-subunit (). PD57 seems to bind the same hydrophobic pocket, with key residues being L646 on segment S6 and F619 on the pore helix of an adjacent subunit (Figure 6) (Perry et al., 2009).
Figure 6
Interestingly, ICA74 binds to a similar hydrophobic pocket in the related ether-a-go-go (EAG) type 1 channel (Kv10.1), although eliciting an opposite effect as in Kv11.1. ICA74 inhibits Kv10.1 currents by enhancing channel inactivation (). The key residues for ICA74 binding in Kv11.1 (F557 and L622) are indeed conserved in Kv10.1 (F359 and L434). Furthermore, only three residues (F619, L646, and M651) of the proposed ICA binding pocket in Kv11.1 appeared not to be conserved in the Kv10.1 channel. This strongly suggests that ICA74 binds to the same hydrophobic site in Kv10.1 and Kv11.1 channels ().
Lateral Pore Wall Fenestrations
The reports of compounds that can access the inner cavity from lipid exposed side-pocket through lateral pore wall fenestrations in Kv channels is still very limited. In Kv7.1 a pore wall fenestration is formed upon interaction with the β-subunit KCNE1 such that adamantane compounds, AC-1 (CAS No.: 878489-28-2) and its analogs (ACs), can reach their binding site (; Wrobel et al., 2016). Interestingly, AC-1 does not affect currents generated by homomeric Kv7.1, channels, nor Kv7.1 co-expressed with other KCNE isoforms (KCNE2-5). Thus, the “β-subunit-induced fenestrations” seem to be required for AC binding. Within these fenestrations many residues have been identified as important for AC-1 activity (V334, F335, I337, F340, and A344, according Kv7.1 numbering), but its position relative to the central cavity and lipophilic side-pocket could not be elucidated (Wrobel et al., 2016).
In Kv11.1, a lateral pore wall fenestration is possibly formed upon mutation of residue F557 to leucine (F557L), explaining the decrease in current inhibition of six known hERG blockers (dofetilide, haloperidol, terfenadine, astemizole, cisapride, and amiodarone) (Saxena et al., 2016). For Kv1.5 it has been proposed that psora-4 molecules can move in the I502A mutant between the central cavity and the lipophilic side-pockets through fenestration between segments S5–S6 ().Hence, the presence of pore wall fenestrations has thus far only been observed upon β-subunit interaction with Kv7.1 and to be induced by mutations in Kv1.5 and Kv11.1 (; Saxena et al., 2016; Wrobel et al., 2016). Although the presence of fenestrations has not been reported yet for wild-type channels, the likelihood that some Kv channel types express lateral pore wall fenestrations increases. If present, these fenestrations may be similar to those characterized in Nav channels, which allow LA's to pass between the central cavity and lipophilic side-pockets (Payandeh et al., 2011; ; Payandeh et al., 2012; Zhang et al., 2012; ; Wrobel et al., 2016).
Unifying the Lipid Exposed/Accessible Drug/Toxin Binding Sites of Kv and Nav Channels
Whereas a classification exists for the different drug/toxin binding sites in Nav channels, such a categorization is currently lacking for the Kv channel family. In this review we take a first step and describe the hydrophobic binding sites reported in different Kv channel families. A compound like gambierol (Figures 4 and 7) has been shown to mostly bind to the front side of the PD, while RTG (Figures 5 and 7) PD57, and ICA74 (Figure 6) also interact with the back side of the PD of an adjacent α-subunit. When mapping all the sites it appears that gambierol, ICA74, RTG, and PD57 bind to an analogues binding site present in different Kv channel types (Figure 7) (Schenzer et al., 2005; ; ; Perry et al., 2009; ; ). This indicates that an analogues lipophilic binding site is conserved between the different Kv channel types.
Figure 7
Zinc pyrithione, on the other hand, seems to solely bind to the back side of the PD, implying that the front and back side of the PD could serve as distinct binding sites (Figures 5 and 7) (Xiong et al., 2007). Psora-4 and sevoflurane are less specific regarding their binding site, as they both bind to the front and back side of the PD, among others (; ; Stock et al., 2018). Although certain compounds solely bind to the front or back side of the PD it seems that all residues point toward a similar lipophilic region, leading to the speculation that these seemingly distinct binding sites may converge to just one conserved lipophilic binding region in Kv channels. This binding site is then most likely similar to neurotoxin site 5 in Nav channels (Figure 3) (; ), located between DIS6-DIVS5 (Figures 3, 4, 5) (). The idea that these binding sites are orthologous equivalents is because gambierol presumably binds to site 5 in Nav channels (). In the case of Kv channels four such binding sites are present due to its tetrameric nature, as opposed to Nav channels who only have one neurotoxin site 5 (Schenzer et al., 2005; ; ; ; Stock et al., 2018).
Another lipid facing binding site is located on the VSD, in particular the cleft between segments S2–S3 and S3–S4. PUFAs, DHAA and its derivatives, and ICA-compounds allegedly bind to these clefts in Shaker and Kv7 channels, respectively (Figures 4D, 5D, and 7) (Padilla et al., 2009; ; Ottosson et al., 2017; Wang et al., 2017). This leads to the assumption that also this lipophilic binding site is conserved between different Kv channel types. Additionally, certain compounds of Nav (LAs and sevoflurane) and Kv (AC-1, psora-4, and several hERG blockers) channels have been proposed to use hydrophobic lateral pore wall fenestrations to reach their binding sites. The location of these fenestrations in Nav channels are situated between DI–DII and DIII–DIV (Figure 1), while in Kv channels they are most likely present between segments S5–S6, allowing lipid soluble compounds to reach their binding site even when the channel is in its closed state (Payandeh et al., 2011; ; Payandeh et al., 2012; Zhang et al., 2012; ; ; ; Saxena et al., 2016; Wrobel et al., 2016).
Of note, the residues reported to affect drug/toxin affinity were in this review mapped on available 3D structures that are snapshots of the channel in a certain state, which should not be the high affinity state for the respective drug/toxin. As mentioned, several drugs/toxins are state dependent and bind with highest affinity to a certain conformation of the channel (e.g., the closed or open channel configuration). Consequently, the residues reported to be important for drug/toxin effect might orient differently when the conformation of the channel changes. Thus, when the channel is in its high affinity drug/toxin state, the orientation of the residues might be slightly different resulting in possibly broader binding regions than highlighted in the figures. Furthermore, several of the residues reported to be important for drug/toxin effect are likely not the binding partners of the drugs/toxins but alter affinity in an allosteric way. Nonetheless, there seem to be three distinct lipid-exposed binding sites preserved in Kv channels: the front and back side of the PD, and S2–S3/S3–S4 clefts. Future experiments will determine if the front and back PD binding sites are two distinct entities or if they converge to just one larger lipophilic binding site region.
Funding
This review was supported by a grant of the FWO (Fonds voor Wetenschappelijk onderzoek) G0C6220N (DJS) and by a GOA grant of the University of Antwerp.
Statements
Author contributions
All authors contributed in writing this review.
Conflict of interest
The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.
References
1
AhernC. A.PayandehJ.BosmansF.ChandaB. (2016). The hitchhiker's guide to the voltage-gated sodium channel galaxy. J. Gen. Physiol.147, 1–24. doi: 10.1085/jgp.201511492
2
AlabiA. A.BahamondeM. I.JungH. J.KimJ. I.SwartzK. J. (2007). Portability of paddle motif function and pharmacology in voltage sensors. Nature450, 370–375. doi: 10.1038/nature06266
3
AmeriA.ShiQ.AschoffJ.PetersT. (1996). Electrophysiological effects of aconitine in rat hippocampal slices. Neuropharmacology35, 13–22. doi: 10.1016/0028-3908(95)00153-0
4
ArmstrongC. M.HollingworthS. (2018). A perspective on Na and K channel inactivation. J. Gen. Physiol.150, 7–18. doi: 10.1085/jgp.201711835
5
ArmstrongC. M.LobodaA. (2001). A model for 4-aminopyridine action on K channels: similarities to tetraethylammonium ion action. Biophys. J.81, 895–904. doi: 10.1016/S0006-3495(01)75749-9
6
ArmstrongC. M. (1971). Interaction of tetraethylammonium ion derivatives with the potassium channels of giant axons. J. Gen. Physiol.58, 413–437. doi: 10.1085/jgp.58.4.413
7
BabcockJ. J.LiM. (2013). hERG channel function: beyond long QT. Acta Pharmacol. Sin.34, 329–335. doi: 10.1038/aps.2013.6
8
BadenD. G. (1983). Marine food-borne dinoflagellate toxins. Int. Rev. Cytol.82, 99–150. doi: 10.1016/S0074-7696(08)60824-4
9
BanerjeeA.LeeA.CampbellE.MackinnonR. (2013). Structure of a pore-blocking toxin in complex with a eukaryotic voltage-dependent K(+) channel. Elife2, e00594. doi: 10.7554/eLife.00594
10
BarberA. F.LiangQ.AmaralC.TreptowW.CovarrubiasM. (2011). Molecular mapping of general anesthetic sites in a voltage-gated ion channel. Biophys. J.101, 1613–1622. doi: 10.1016/j.bpj.2011.08.026
11
BarberA. F.CarnevaleV.KleinM. L.EckenhoffR. G.CovarrubiasM. (2014). Modulation of a voltage-gated Na+ channel by sevoflurane involves multiple sites and distinct mechanisms. Proc. Natl. Acad. Sci. United States America111, 6726–6731. doi: 10.1073/pnas.1405768111
12
BarnesS.HilleB. (1988). Veratridine modifies open sodium channels. J. Gen. Physiol.91, 421–443. doi: 10.1085/jgp.91.3.421
13
BavroV. N.De ZorziR.SchmidtM. R.MunizJ. R.ZubcevicL.SansomM. S.et al. (2012). Structure of a KirBac potassium channel with an open bundle crossing indicates a mechanism of channel gating. Nat. Struct. Mol. Biol.19, 158–163. doi: 10.1038/nsmb.2208
14
BenoitE.LegrandA. M.DuboisJ. M. (1986). Effects of ciguatoxin on current and voltage clamped frog myelinated nerve fibre. Toxicon24, 357–364. doi: 10.1016/0041-0101(86)90195-9
15
BentzenB. H.SchmittN.CalloeK.Dalby BrownW.GrunnetM.OlesenS. P. (2006). The acrylamide (S)-1 differentially affects Kv7 (KCNQ) potassium channels. Neuropharmacology51, 1068–1077. doi: 10.1016/j.neuropharm.2006.07.001
16
BenzI.KohlhardtM. (1991). Responsiveness of cardiac Na+ channels to antiarrhythmic drugs: the role of inactivation. J. Membr. Biol.122, 267–278. doi: 10.1007/BF01871427
17
BezanillaF. (2008). How membrane proteins sense voltage. Nat. Rev. Mol. Cell Biol.9, 323–332. doi: 10.1038/nrm2376
18
BidardJ. N.VijverbergH. P.FrelinC.ChungueE.LegrandA. M.BagnisR.et al. (1984). Ciguatoxin is a novel type of Na+ channel toxin. J. Biol. Chem.259, 8353–8357.
19
BlunckR.BatulanZ. (2012). Mechanism of electromechanical coupling in voltage-gated potassium channels. Front. Pharmacol.3, 166. doi: 10.3389/fphar.2012.00166
20
BolandL. M.DrzewieckiM. M. (2008). Polyunsaturated fatty acid modulation of voltage-gated ion channels. Cell Biochem. Biophys.52, 59–84. doi: 10.1007/s12013-008-9027-2
21
BorjessonS. I.ElinderF. (2011). An electrostatic potassium channel opener targeting the final voltage sensor transition. J. Gen. Physiol.137, 563–577. doi: 10.1085/jgp.201110599
22
BorjessonS. I.HammarstromS.ElinderF. (2008). Lipoelectric modification of ion channel voltage gating by polyunsaturated fatty acids. Biophys. J.95, 2242–2253. doi: 10.1529/biophysj.108.130757
23
CahalanM. D. (1978). Local anesthetic block of sodium channels in normal and pronase-treated squid giant axons. Biophys. J.23, 285–311. doi: 10.1016/S0006-3495(78)85449-6
24
CatterallW. A.GainerM. (1985). Interaction of brevetoxin A with a new receptor site on the sodium channel. Toxicon23, 497–504. doi: 10.1016/0041-0101(85)90034-0
25
CatterallW. A.RiskM. (1981). Toxin T4(6) from Ptychodiscus brevis (formerly Gymnodinium breve) enhances activation of voltage-sensitive sodium channels by veratridine. Mol. Pharmacol.19, 345–348.
26
CatterallW. A. (1980). Neurotoxins that act on voltage-sensitive sodium channels in excitable membranes. Annu. Rev. Pharmacol. Toxicol.20, 15–43. doi: 10.1146/annurev.pa.20.040180.000311
27
CesteleS.CatterallW. A. (2000). Molecular mechanisms of neurotoxin action on voltage-gated sodium channels. Biochimie82, 883–892. doi: 10.1016/S0300-9084(00)01174-3
28
CourtneyK. R. (1988). Why do some drugs preferentially block open sodium channels? J. Mol. Cell Cardiol.20, 461–464. doi: 10.1016/S0022-2828(88)80073-7
29
CuelloL. G.JoginiV.CortesD. M.PerozoE. (2010). Structural mechanism of C-type inactivation in K(+) channels. Nature466, 203–208. doi: 10.1038/nature09153
30
CuypersE.Abdel-MottalebY.KopljarI.RainierJ. D.RaesA. L.SnydersD. J.et al. (2008). Gambierol, a toxin produced by the dinoflagellate Gambierdiscus toxicus, is a potent blocker of voltage-gated potassium channels. Toxicon : Off. J. Int. Soc. Toxinol.51, 974–983. doi: 10.1016/j.toxicon.2008.01.004
31
DalyJ. W.WitkopB.BommerP.BiemannK. (1965). Batrachotoxin. The active principle of the Colombian arrow poison frog, Phyllobates bicolor. J. Am. Chem. Soc.87, 124–126. doi: 10.1021/ja01079a026
32
De Lera RuizM.KrausR. L. (2015). Voltage-Gated Sodium Channels: Structure, Function, Pharmacology, and Clinical Indications. J. Med. Chem.58, 7093–7118. doi: 10.1021/jm501981g
33
Del CaminoD.KanevskyM.YellenG. (2005). Status of the intracellular gate in the activated-not-open state of shaker K+ channels. J. Gen. Physiol.126, 419–428. doi: 10.1085/jgp.200509385
34
DoyleD. A.Morais CabralJ.PfuetznerR. A.KuoA.GulbisJ. M.CohenS. L.et al. (1998). The structure of the potassium channel: molecular basis of K+ conduction and selectivity. Science280, 69–77. doi: 10.1126/science.280.5360.69
35
DuY.GardenD. P.WangL.ZhorovB. S.DongK. (2011). Identification of new batrachotoxin-sensing residues in segment IIIS6 of the sodium channel. J. Biol. Chem.286, 13151–13160. doi: 10.1074/jbc.M110.208496
36
DuboisJ. M.SchneiderM. F.KhodorovB. I. (1983). Voltage dependence of intramembrane charge movement and conductance activation of batrachotoxin-modified sodium channels in frog node of Ranvier. J. Gen. Physiol.81, 829–844. doi: 10.1085/jgp.81.6.829
37
ErikssonM. A.RouxB. (2002). Modeling the structure of agitoxin in complex with the Shaker K+ channel: a computational approach based on experimental distance restraints extracted from thermodynamic mutant cycles. Biophys. J.83, 2595–2609. doi: 10.1016/S0006-3495(02)75270-3
38
FernandezD.GhantaA.KauffmanG. W.SanguinettiM. C. (2004). Physicochemical features of the HERG channel drug binding site. J. Biol. Chem.279, 10120–10127. doi: 10.1074/jbc.M310683200
39
FlewellingL. J.NaarJ. P.AbbottJ. P.BadenD. G.BarrosN. B.BossartG. D.et al. (2005). Brevetoxicosis: red tides and marine mammal mortalities. Nature435, 755–756. doi: 10.1038/nature435755a
40
FrieseJ.GleitzJ.GutserU. T.HeubachJ. F.MatthiesenT.WilffertB.et al. (1997). Aconitum sp. alkaloids: the modulation of voltage-dependent Na+ channels, toxicity and antinociceptive properties. Eur. J. Pharmacol.337, 165–174. doi: 10.1016/S0014-2999(97)01268-5
41
Gamal El-DinT. M.LenaeusM. J.ZhengN.CatterallW. A. (2018). Fenestrations control resting-state block of a voltage-gated sodium channel. Proc. Natl. Acad. Sci. U. S. A115, 13111–13116. doi: 10.1073/pnas.1814928115
42
GargV.Stary-WeinzingerA.SachseF.SanguinettiM. C. (2011). Molecular determinants for activation of human ether-a-go-go-related gene 1 potassium channels by 3-nitro-n-(4-phenoxyphenyl) benzamide. Mol. Pharmacol.80, 630–637. doi: 10.1124/mol.111.073809
43
GargV.SachseF. B.SanguinettiM. C. (2012). Tuning of EAG K(+) channel inactivation: molecular determinants of amplification by mutations and a small molecule. J. Gen. Physiol.140, 307–324. doi: 10.1085/jgp.201210826
44
GargV.Stary-WeinzingerA.SanguinettiM. C. (2013). ICA-105574 interacts with a common binding site to elicit opposite effects on inactivation gating of EAG and ERG potassium channels. Mol. Pharmacol.83, 805–813. doi: 10.1124/mol.112.084384
45
GingrichK. J.BeardsleyD.YueD. T. (1993). Ultra-deep blockade of Na+ channels by a quaternary ammonium ion: catalysis by a transition-intermediate state? J. Physiol.471, 319–341. doi: 10.1113/jphysiol.1993.sp019903
46
GrantA. O.DietzM. A.GilliamF. R.StarmerC. F. (1989). Blockade of cardiac sodium channels by lidocaine. Single-channel analysis. Circ. Res.65, 1247–1262. doi: 10.1161/01.res.65.5.1247
47
HerzogW. H.FeibelR. M.BryantS. H. (1964). The Effect of Aconitine on the Giant Axon of the Squid. J. Gen. Physiol.47, 719–733. doi: 10.1085/jgp.47.4.719
48
HillA. P.SundeM.CampbellT. J.VandenbergJ. I. (2007). Mechanism of block of the hERG K+ channel by the scorpion toxin CnErg1. Biophys. J.92, 3915–3929. doi: 10.1529/biophysj.106.101956
49
HilleB. (1977). Local anesthetics: hydrophilic and hydrophobic pathways for the drug-receptor reaction. J. Gen. Physiol.69, 497–515. doi: 10.1085/jgp.69.4.497
50
HilleB. (2001). Ion Channels of Excitable Membranes.
51
HorishitaT.EgerE. I.2ndHarrisR. A. (2008). The effects of volatile aromatic anesthetics on voltage-gated Na+ channels expressed in Xenopus oocytes. Anesth Analg.107, 1579–1586. doi: 10.1213/ane.0b013e318184b966
52
HoshiT.ZagottaW. N.AldrichR. W. (1990). Biophysical and molecular mechanisms of Shaker potassium channel inactivation. Science250, 533–538. doi: 10.1126/science.2122519
53
HoshiT.ZagottaW. N.AldrichR. W. (1991). Two types of inactivation in Shaker K+ channels: effects of alterations in the carboxy-terminal region. Neuron7, 547–556. doi: 10.1016/0896-6273(91)90367-9
54
HuangL. Y.MoranN.EhrensteinG. (1982). Batrachotoxin modifies the gating kinetics of sodium channels in internally perfused neuroblastoma cells. Proc. Natl. Acad. Sci. U. S. A..79, 2082–2085. doi: 10.1073/pnas.79.6.2082
55
InserraM. C.IsraelM. R.CaldwellA.CastroJ.DeuisJ. R.HarringtonA. M.et al. (2017). Multiple sodium channel isoforms mediate the pathological effects of Pacific ciguatoxin-1. Sci. Rep.7, 42810. doi: 10.1038/srep42810
56
JanL. Y.JanY. N. (2012). Voltage-gated potassium channels and the diversity of electrical signalling. J. Physiol.590, 2591–2599. doi: 10.1113/jphysiol.2011.224212
57
JansenS. A.KleerekooperI.HofmanZ. L.KappenI. F.Stary-WeinzingerA.Van Der HeydenM. A. (2012). Grayanotoxin poisoning: ‘mad honey disease' and beyond. Cardiovasc. Toxicol.12, 208–215. doi: 10.1007/s12012-012-9162-2
58
JaraskovaL.LindersJ. T. M.Van Der VekenL. J. E.WillemsensG. H. M.BischoffF. P. (2005). N-2 Adamantanyl-2-Phenoxy-Acetamide Derivatives as 11-Beta Hydroxysteroid Dehydrogenase Inhibitors.
59
JungH. J.LeeJ. Y.KimS. H.EuY. J.ShinS. Y.MilescuM.et al. (2005). Solution structure and lipid membrane partitioning of VSTx1, an inhibitor of the KvAP potassium channel. Biochemistry44, 6015–6023. doi: 10.1021/bi0477034
60
JungH. H.JungH. J.MilescuM.LeeC. W.LeeS.LeeJ. Y.et al. (2010). Structure and orientation of a voltage-sensor toxin in lipid membranes. Biophys. J.99, 638–646. doi: 10.1016/j.bpj.2010.04.061
61
KaczmarskiJ. A.CorryB. (2014). Investigating the size and dynamics of voltage-gated sodium channel fenestrations. Channels (Austin)8, 264–277. doi: 10.4161/chan.28136
62
KamiyaK.MitchesonJ. S.YasuiK.KodamaI.SanguinettiM. C. (2001). Open channel block of HERG K(+) channels by vesnarinone. Mol. Pharmacol.60, 244–253. doi: 10.1124/mol.60.2.244
63
KamiyaK.NiwaR.MorishimaM.HonjoH.SanguinettiM. C. (2008). Molecular determinants of hERG channel block by terfenadine and cisapride. J. Pharmacol. Sci.108, 301–307. doi: 10.1254/jphs.08102FP
64
KimR. Y.YauM. C.GalpinJ. D.SeebohmG.AhernC. A.PlessS. A.et al. (2015). Atomic basis for therapeutic activation of neuronal potassium channels. Nat. Commun.6, 8116. doi: 10.1038/ncomms9116
65
KlemicK. G.ShiehC. C.KirschG. E.JonesS. W. (1998). Inactivation of Kv2.1 potassium channels. Biophys. J.74, 1779–1789. doi: 10.1016/S0006-3495(98)77888-9
66
KonokiK.BadenD. G.ScheuerT.CatterallW. A. (2019). Molecular Determinants of Brevetoxin Binding to Voltage-Gated Sodium Channels. Toxins11, 513. doi: 10.3390/toxins11090513
67
KopljarI.LabroA. J.CuypersE.JohnsonH. W.RainierJ. D.TytgatJ.et al. (2009). A polyether biotoxin binding site on the lipid-exposed face of the pore domain of Kv channels revealed by the marine toxin gambierol. Proc. Natl. Acad. Sci. U. S. A.106, 9896–9901. doi: 10.1073/pnas.0812471106
68
KubischC.SchroederB. C.FriedrichT.LutjohannB.El-AmraouiA.MarlinS.et al. (1999). KCNQ4, a novel potassium channel expressed in sensory outer hair cells, is mutated in dominant deafness. Cell96, 437–446. doi: 10.1016/S0092-8674(00)80556-5
69
LabroA. J.SnydersD. J. (2012). Being flexible: the voltage-controllable activation gate of kv channels. Front. Pharmacol.3, 168. doi: 10.3389/fphar.2012.00168
70
LangeW.GeissendorferJ.SchenzerA.GrotzingerJ.SeebohmG.FriedrichT.et al. (2009). Refinement of the binding site and mode of action of the anticonvulsant Retigabine on KCNQ K+ channels. Mol. Pharmacol.75, 272–280. doi: 10.1124/mol.108.052282
71
LeafA. (2007). Prevention of sudden cardiac death by n-3 polyunsaturated fatty acids. J. Cardiovasc. Med. (Hagerstown)8 Suppl 1, S27–S29. doi: 10.2459/01.JCM.0000289270.98105.b3
72
LeeC. W.KimS.RohS. H.EndohH.KoderaY.MaedaT.et al. (2004). Solution structure and functional characterization of SGTx1, a modifier of Kv2.1 channel gating. Biochemistry43, 890–897. doi: 10.1021/bi0353373
73
Lees-MillerJ. P.DuanY.TengG. Q.DuffH. J. (2000). Molecular determinant of high-affinity dofetilide binding to HERG1 expressed in Xenopus oocytes: involvement of S6 sites. Mol. Pharmacol.57, 367–374.
74
LefevreF.AronsonN. (2000). Ketogenic diet for the treatment of refractory epilepsy in children: A systematic review of efficacy. Pediatrics105, E46. doi: 10.1542/peds.105.4.e46
75
LenaeusM. J.Gamal El-DinT. M.IngC.RamanadaneK.PomesR.ZhengN.et al. (2017). Structures of closed and open states of a voltage-gated sodium channel. Proc. Natl. Acad. Sci. U. S. A114, E3051–E3060. doi: 10.1073/pnas.1700761114
76
LepageK.RainierJ.JohnsonH.BadenD.MurrayT. (2007). Gambierol Acts as a Functional Antagonist of Neurotoxin Site 5 on Voltage-Gated Sodium Channels in Cerebellar Granule Neurons. J. Pharmacol. Exp. Ther.323, 174–179. doi: 10.1124/jpet.107.124271
77
LewisR. J.VernouxJ.-P.BreretonI. M. (1998). Structure of Caribbean Ciguatoxin Isolated from Caranx latus. J. Am. Chem. Soc.120, 5914–5920. doi: 10.1021/ja980389e
78
LiangQ.AndersonW. D.JonesS. T.SouzaC. S.HosoumeJ. M.TreptowW.et al. (2015). Positive Allosteric Modulation of Kv Channels by Sevoflurane: Insights into the Structural Basis of Inhaled Anesthetic Action. PloS One10, e0143363. doi: 10.1371/journal.pone.0143363
79
LiinS. I.Silvera EjnebyM.Barro-SoriaR.SkarsfeldtM. A.LarssonJ. E.Starck HarlinF.et al. (2015). Polyunsaturated fatty acid analogs act antiarrhythmically on the cardiac IKs channel. Proc. Natl. Acad. Sci. U. S. A.112, 5714–5719. doi: 10.1073/pnas.1503488112
80
LinY.-Y.RiskM.RayS. M.Van EngenD.ClardyJ.GolikJ.et al. (1981). Isolation and structure of brevetoxin B from the “red tide” dinoflagellate Ptychodiscus brevis (Gymnodinium breve). J. Am. Chem. Soc.103, 6773–6775. doi: 10.1021/ja00412a053
81
LinfordN. J.CantrellA. R.QuY.ScheuerT.CatterallW. A. (1998). Interaction of batrachotoxin with the local anesthetic receptor site in transmembrane segment IVS6 of the voltage-gated sodium channel. Proc. Natl. Acad. Sci. U. S. A.95, 13947–13952. doi: 10.1073/pnas.95.23.13947
82
LombetA.BidardJ. N.LazdunskiM. (1987). Ciguatoxin and brevetoxins share a common receptor site on the neuronal voltage-dependent Na+ channel. FEBS Lett.219, 355–359. doi: 10.1016/0014-5793(87)80252-1
83
LongS. B.TaoX.CampbellE. B.MackinnonR. (2007). Atomic structure of a voltage-dependent K+ channel in a lipid membrane-like environment. Nature450, 376–382. doi: 10.1038/nature06265
84
LuzhkovV. B.AqvistJ. (2001). Mechanisms of tetraethylammonium ion block in the KcsA potassium channel. FEBS Lett.495, 191–196. doi: 10.1016/S0014-5793(01)02381-X
85
ManvilleR. W.PapanikolaouM.AbbottG. W. (2018). Direct neurotransmitter activation of voltage-gated potassium channels. Nat. Commun.9, 1847. doi: 10.1038/s41467-018-04266-w
86
MarbanE.YamagishiT.TomaselliG. F. (1998). Structure and function of voltage-gated sodium channels. J. Physiol.508 ( Pt 3), 647–657. doi: 10.1111/j.1469-7793.1998.647bp.x
87
Martinez-MoralesE.KopljarI.RainierJ. D.TytgatJ.SnydersD. J.LabroA. J. (2016). Gambierol and n-alkanols inhibit Shaker Kv channel via distinct binding sites outside the K(+) pore. Toxicon120, 57–60. doi: 10.1016/j.toxicon.2016.07.017
88
MarzianS.StansfeldP. J.RapediusM.RinneS.Nematian-ArdestaniE.AbbruzzeseJ. L.et al. (2013). Side pockets provide the basis for a new mechanism of Kv channel-specific inhibition. Nat. Chem. Biol.9, 507–513. doi: 10.1038/nchembio.1271
89
MccuskerE. C.BagnerisC.NaylorC. E.ColeA. R.D'avanzoN.NicholsC. G.et al. (2012). Structure of a bacterial voltage-gated sodium channel pore reveals mechanisms of opening and closing. Nat. Commun.3, 1102. doi: 10.1038/ncomms2077
90
Mcnaughton-SmithG. A.GrossM. F.WickendenA. D. (2002). Benzanilides as potassium channel openers, United States patent application 6372767.
91
MihailescuM.KrepkiyD.MilescuM.GawrischK.SwartzK. J.WhiteS. (2014). Structural interactions of a voltage sensor toxin with lipid membranes. Proc. Natl. Acad. Sci. U. S. A111, E5463–E5470. doi: 10.1073/pnas.1415324111
92
MitchesonJ. S.ChenJ.LinM.CulbersonC.SanguinettiM. C. (2000). A structural basis for drug-induced long QT syndrome. Proc. Natl. Acad. Sci. U. S. A.97, 12329–12333. doi: 10.1073/pnas.210244497
93
MurataM.LegrandA. M.IshibashiY.FukuiM.YasumotoT. (1990). Structures and configurations of ciguatoxin from the moray eel Gymnothorax javanicus and its likely precursor from the dinoflagellate Gambierdiscus toxicus. J. Am. Chem. Soc.112, 4380–4386. doi: 10.1021/ja00167a040
94
NicholsonG.LewisR. (2006). Ciguatoxins: Cyclic Polyether Modulators of Voltage-gated Ion Channel Function. Marine Drugs4, 82–118. doi: 10.3390/md403082
95
OttossonN. E.Silvera EjnebyM.WuX.YazdiS.KonradssonP.LindahlE.et al. (2017). A drug pocket at the lipid bilayer-potassium channel interface. Sci. Adv.3, e1701099. doi: 10.1126/sciadv.1701099
96
OuyangW.JihT. Y.ZhangT. T.CorreaA. M.HemmingsH. C. Jr. (2007). Isoflurane inhibits NaChBac, a prokaryotic voltage-gated sodium channel. J. Pharmacol. Exp. Ther.322, 1076–1083. doi: 10.1124/jpet.107.122929
97
OuyangW.HeroldK. F.HemmingsH. C.Jr. (2009). Comparative effects of halogenated inhaled anesthetics on voltage-gated Na+ channel function. Anesthesiology110, 582–590. doi: 10.1097/ALN.0b013e318197941e
98
PadillaK.WickendenA. D.GerlachA. C.MccormackK. (2009). The KCNQ2/3 selective channel opener ICA-27243 binds to a novel voltage-sensor domain site. Neurosci. Lett.465, 138–142. doi: 10.1016/j.neulet.2009.08.071
99
PayandehJ.ScheuerT.ZhengN.CatterallW. A. (2011). The crystal structure of a voltage-gated sodium channel. Nature475, 353–358. doi: 10.1038/nature10238
100
PayandehJ.Gamal El-DinT. M.ScheuerT.ZhengN.CatterallW. A. (2012). Crystal structure of a voltage-gated sodium channel in two potentially inactivated states. Nature486, 135–139. doi: 10.1038/nature11077
101
PerryM.SachseF. B.AbbruzzeseJ.SanguinettiM. C. (2009). PD-118057 contacts the pore helix of hERG1 channels to attenuate inactivation and enhance K+ conductance. Proc. Natl. Acad. Sci. U. S. A106, 20075–20080. doi: 10.1073/pnas.0906597106
102
PettersenE. F.GoddardT. D.HuangC. C.CouchG. S.GreenblattD. M.MengE. C.et al. (2004). UCSF Chimera–a visualization system for exploratory research and analysis. J. Comput. Chem.25, 1605–1612. doi: 10.1002/jcc.20084
103
PoliM. A.MendeT. J.BadenD. G. (1986). Brevetoxins, unique activators of voltage-sensitive sodium channels, bind to specific sites in rat brain synaptosomes. Mol. Pharmacol.30, 129–135.
104
RagsdaleD. S.McpheeJ. C.ScheuerT.CatterallW. A. (1994). Molecular determinants of state-dependent block of Na+ channels by local anesthetics. Science265, 1724–1728. doi: 10.1126/science.8085162
105
RajuS. G.BarberA. F.LebardD. N.KleinM. L.CarnevaleV. (2013). Exploring volatile general anesthetic binding to a closed membrane-bound bacterial voltage-gated sodium channel via computation. PloS Comput. Biol.9, e1003090. doi: 10.1371/journal.pcbi.1003090
106
ReinK. S.BadenD. G.GawleyR. E. (1994). Conformational Analysis of the Sodium Channel Modulator, Brevetoxin A, Comparison with Brevetoxin B Conformations, and a Hypothesis about the Common Pharmacophore of the “Site 5”. Toxins. J. Organ. Chem.59, 2101–2106. doi: 10.1021/jo00087a027
107
RundfeldtC. (1997). The new anticonvulsant retigabine (D-23129) acts as an opener of K+ channels in neuronal cells. Eur. J. Pharmacol.336, 243–249. doi: 10.1016/S0014-2999(97)01249-1
108
Sanchez-ChapulaJ. A.Navarro-PolancoR. A.CulbersonC.ChenJ.SanguinettiM. C. (2002). Molecular determinants of voltage-dependent human ether-a-go-go related gene (HERG) K+ channel block. J. Biol. Chem.277, 23587–23595. doi: 10.1074/jbc.M200448200
109
Sanchez-ChapulaJ. A.FerrerT.Navarro-PolancoR. A.SanguinettiM. C. (2003). Voltage-dependent profile of human ether-a-go-go-related gene channel block is influenced by a single residue in the S6 transmembrane domain. Mol. Pharmacol.63, 1051–1058. doi: 10.1124/mol.63.5.1051
110
SaxenaP.Zangerl-PlesslE. M.LinderT.WindischA.HohausA.TiminE.et al. (2016). New potential binding determinant for hERG channel inhibitors. Sci. Rep.6, 24182. doi: 10.1038/srep24182
111
SchenzerA.FriedrichT.PuschM.SaftigP.JentschT. J.GrotzingerJ.et al. (2005). Molecular determinants of KCNQ (Kv7) K+ channel sensitivity to the anticonvulsant retigabine. J. Neurosci.25, 5051–5060. doi: 10.1523/JNEUROSCI.0128-05.2005
112
SchreibmayerW.JeglitschG. (1992). The sodium channel activator Brevetoxin-3 uncovers a multiplicity of different open states of the cardiac sodium channel. Biochim. Biophys. Acta1104, 233–242. doi: 10.1016/0005-2736(92)90035-K
113
SchroederB. C.HechenbergerM.WeinreichF.KubischC.JentschT. J. (2000). KCNQ5, a novel potassium channel broadly expressed in brain, mediates M-type currents. J. Biol. Chem.275, 24089–24095. doi: 10.1074/jbc.M003245200
114
SheridanR. E.AdlerM. (1989). The actions of a red tide toxin from Ptychodiscus brevis on single sodium channels in mammalian neuroblastoma cells. FEBS Lett.247, 448–452. doi: 10.1016/0014-5793(89)81389-4
115
StevensM.PeigneurS.TytgatJ. (2011). Neurotoxins and their binding areas on voltage-gated sodium channels. Front. Pharmacol.2, 71. doi: 10.3389/fphar.2011.00071
116
StockL.HosoumeJ.CirqueiraL.TreptowW. (2018). Binding of the general anesthetic sevoflurane to ion channels. PloS Comput. Biol.14, e1006605. doi: 10.1371/journal.pcbi.1006605
117
StrachanL. C.LewisR. J.NicholsonG. M. (1999). Differential actions of pacific ciguatoxin-1 on sodium channel subtypes in mammalian sensory neurons. J. Pharmacol. Exp. Ther.288, 379–388.
118
SunJ.MackinnonR. (2017). Cryo-EM Structure of a KCNQ1/CaM Complex Reveals Insights into Congenital Long QT Syndrome. Cell169, 1042–1050 e1049. doi: 10.1016/j.cell.2017.05.019
119
SunamiA.GlaaserI. W.FozzardH. A. (2001). Structural and gating changes of the sodium channel induced by mutation of a residue in the upper third of IVS6, creating an external access path for local anesthetics. Mol. Pharmacol.59, 684–691. doi: 10.1124/mol.59.4.684
120
SutroJ. B. (1986). Kinetics of veratridine action on Na channels of skeletal muscle. J. Gen. Physiol.87, 1–24. doi: 10.1085/jgp.87.1.1
121
SwartzK. J. (2007). Tarantula toxins interacting with voltage sensors in potassium channels. Toxicon : Off. J. Int. Soc. Toxinol.49, 213–230. doi: 10.1016/j.toxicon.2006.09.024
122
ThomasD.KarleC. A.KiehnJ. (2006). The cardiac hERG/IKr potassium channel as pharmacological target: structure, function, regulation, and clinical applications. Curr. Pharm. Des.12, 2271–2283. doi: 10.2174/138161206777585102
123
TrainerV. L.ThomsenW. J.CatterallW. A.BadenD. G. (1991). Photoaffinity labeling of the brevetoxin receptor on sodium channels in rat brain synaptosomes. Mol. Pharmacol.40, 988–994.
124
TrainerV. L.BadenD. G.CatterallW. A. (1994). Identification of peptide components of the brevetoxin receptor site of rat brain sodium channels. J. Biol. Chem.269, 19904–19909.
125
UeharaA.MoczydlowskiE. (1986). Blocking mechanisms of batrachotoxin-activated Na channels in artificial bilayers. Membr. Biochem.6, 111–147. doi: 10.3109/09687688609065446
126
UlbrichtW. (1969). The effect of veratridine on excitable membranes of nerve and muscle. Ergeb. Physiol.61, 18–71. doi: 10.1007/BFb0111446
127
VandenbergJ. I.PerryM. D.PerrinM. J.MannS. A.KeY.HillA. P. (2012). hERG K(+) channels: structure, function, and clinical significance. Physiol. Rev.92, 1393–1478. doi: 10.1152/physrev.00036.2011
128
VennekampJ.WulffH.BeetonC.CalabresiP. A.GrissmerS.HanselW.et al. (2004). Kv1.3-blocking 5-phenylalkoxypsoralens: a new class of immunomodulators. Mol. Pharmacol.65, 1364–1374. doi: 10.1124/mol.65.6.1364
129
WangW.MackinnonR. (2017). Cryo-EM Structure of the Open Human Ether-a-go-go-Related K(+) Channel hERG. Cell169, 422–430 e410. doi: 10.1016/j.cell.2017.03.048
130
WangS. Y.WangG. K. (1998). Point mutations in segment I-S6 render voltage-gated Na+ channels resistant to batrachotoxin. Proc. Natl. Acad. Sci. U. S. A.95, 2653–2658. doi: 10.1073/pnas.95.5.2653
131
WangS. Y.WangG. K. (1999). Batrachotoxin-resistant Na+ channels derived from point mutations in transmembrane segment D4-S6. Biophys. J.76, 3141–3149. doi: 10.1016/S0006-3495(99)77465-5
132
WangG. K.WangS. Y. (2003). Veratridine block of rat skeletal muscle Nav1.4 sodium channels in the inner vestibule. J. Physiol.548, 667–675. doi: 10.1113/jphysiol.2002.035469
133
WangH. S.PanZ.ShiW.BrownB. S.WymoreR. S.CohenI. S.et al. (1998). KCNQ2 and KCNQ3 potassium channel subunits: molecular correlates of the M-channel. Science282, 1890–1893. doi: 10.1126/science.282.5395.1890
134
WangS. Y.NauC.WangG. K. (2000). Residues in Na(+) channel D3-S6 segment modulate both batrachotoxin and local anesthetic affinities. Biophys. J.79, 1379–1387. doi: 10.1016/S0006-3495(00)76390-9
135
WangS. Y.BarileM.WangG. K. (2001). Disparate role of Na(+) channel D2-S6 residues in batrachotoxin and local anesthetic action. Mol. Pharmacol.59, 1100–1107. doi: 10.1124/mol.59.5.1100
136
WangS. Y.TikhonovD. B.MitchellJ.ZhorovB. S.WangG. K. (2007). Irreversible block of cardiac mutant Na+ channels by batrachotoxin. Channels (Austin)1, 179–188. doi: 10.4161/chan.4437
137
WangA. W.YangR.KurataH. T. (2017). Sequence determinants of subtype-specific actions of KCNQ channel openers. J. Physiol.595, 663–676. doi: 10.1113/JP272762
138
WangC. K.LamotheS. M.WangA. W.YangR. Y.KurataH. T. (2018). Pore- and voltage sensor-targeted KCNQ openers have distinct state-dependent actions. J. Gen. Physiol.150, 1722–1734. doi: 10.1085/jgp.201812070
139
WaterhouseA. MProcterJ. B.MartinD. M. A.ClampM.BartonG. J. (2009). Jalview Version 2 - a multiple sequence alignment editor and analysis workbench. Bioinformatics25 (9), 1189–1191. doi: 10.1093/bioinformatics/btp033
140
WaterhouseA.BertoniM.BienertS.StuderG.TaurielloG.GumiennyR.et al. (2018). SWISS-MODEL: homology modelling of protein structures and complexes. Nucleic Acids Res.46, W296–W303. doi: 10.1093/nar/gky427
141
WestJ. W.PattonD. E.ScheuerT.WangY.GoldinA. L.CatterallW. A. (1992). A cluster of hydrophobic amino acid residues required for fast Na(+)-channel inactivation. Proc. Natl. Acad. Sci. U. S. A89, 10910–10914. doi: 10.1073/pnas.89.22.10910
142
WickendenA. D.KrajewskiJ. L.LondonB.WagonerP. K.WilsonW. A.ClarkS.et al. (2008). N-(6-chloro-pyridin-3-yl)-3,4-difluoro-benzamide (ICA-27243): a novel, selective KCNQ2/Q3 potassium channel activator. Mol. Pharmacol.73, 977–986. doi: 10.1124/mol.107.043216
143
WrobelE.RothenbergI.KrispC.HundtF.FraenzelB.EckeyK.et al. (2016). KCNE1 induces fenestration in the Kv7.1/KCNE1 channel complex that allows for highly specific pharmacological targeting. Nat. Commun.7, 1–13. doi: 10.1038/ncomms12795
144
WulffH.CastleN. A.PardoL. A. (2009). Voltage-gated potassium channels as therapeutic targets. Nat. Rev. Drug Discovery8, 982–1001. doi: 10.1038/nrd2983
145
XiongQ.SunH.LiM. (2007). Zinc pyrithione-mediated activation of voltage-gated KCNQ potassium channels rescues epileptogenic mutants. Nat. Chem. Biol.3, 287–296. doi: 10.1038/nchembio874
146
XuC. Q.ZhuS. Y.ChiC. W.TytgatJ. (2003). Turret and pore block of K+ channels: what is the difference? Trends Pharmacol. Sci.24, 446–448author reply 448–449. doi: 10.1016/S0165-6147(03)00223-2
147
Yarov-YarovoyV.McpheeJ. C.IdsvoogD.PateC.ScheuerT.CatterallW. A. (2002). Role of amino acid residues in transmembrane segments IS6 and IIS6 of the Na+ channel alpha subunit in voltage-dependent gating and drug block. J. Biol. Chem.277, 35393–35401. doi: 10.1074/jbc.M206126200
148
YazdiS.SteinE.ElinderF.AnderssonM.LindahlE. (2016). The Molecular Basis of Polyunsaturated Fatty Acid Interactions with the Shaker Voltage-Gated Potassium Channel. PloS Comput. Biol.12, e1004704. doi: 10.1371/journal.pcbi.1004704
149
YukiT.YamaokaK.YakehiroM.SeyamaI. (2001). State-dependent action of grayanotoxin I on Na(+) channels in frog ventricular myocytes. J. Physiol.534, 777–790. doi: 10.1111/j.1469-7793.2001.00777.x
150
ZamponiG. W.DoyleD. D.FrenchR. J. (1993). Fast lidocaine block of cardiac and skeletal muscle sodium channels: one site with two routes of access. Biophys. J.65, 80–90. doi: 10.1016/S0006-3495(93)81042-7
151
ZhangM.KorolkovaY. V.LiuJ.JiangM.GrishinE. V.TsengG. N. (2003). BeKm-1 is a HERG-specific toxin that shares the structure with ChTx but the mechanism of action with ErgTx1. Biophys. J.84, 3022–3036. doi: 10.1016/S0006-3495(03)70028-9
152
ZhangX.RenW.DecaenP.YanC.TaoX.TangL.et al. (2012). Crystal structure of an orthologue of the NaChBac voltage-gated sodium channel. Nature486, 130–134. doi: 10.1038/nature11054
Summary
Keywords
hydrophobic binding sites, voltage-gated potassium channels, voltage-gated sodium channels, channel fenestrations, lipophilic compounds
Citation
Van Theemsche KM, Van de Sande DV, Snyders DJ and Labro AJ (2020) Hydrophobic Drug/Toxin Binding Sites in Voltage-Dependent K+ and Na+ Channels. Front. Pharmacol. 11:735. doi: 10.3389/fphar.2020.00735
Received
31 January 2020
Accepted
04 May 2020
Published
15 May 2020
Volume
11 - 2020
Edited by
Mounir Tarek, Centre National de la Recherche Scientifique (CNRS), France
Reviewed by
Michael E. O'Leary, Cooper Medical School of Rowan University, United States; Roope Mannikko, University College London, United Kingdom
Updates
Copyright
© 2020 Van Theemsche, Van de Sande, Snyders and Labro.
This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
*Correspondence: Alain J. Labro, alain.labro@uantwerpen.be
†These authors share first authorship
This article was submitted to Pharmacology of Ion Channels and Channelopathies, a section of the journal Frontiers in Pharmacology
Disclaimer
All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article or claim that may be made by its manufacturer is not guaranteed or endorsed by the publisher.