Pharmac. Ther. Vol. 48, pp. 237-258, 1990 Printed in Great Britain. All rights reserved

0163-7258/90 $0.00 + 0.50 © 1990 Pergamon Press pie

Associate Editor: M. J. LEWIS

POTASSIUM CHANNEL OPENERS AND VASCULAR SMOOTH MUSCLE RELAXATION G. EDWARDS and A. H. WESTON Smooth Muscle Research Group, Department of Physiological Sciences, Stopford Building, University of Manchester, Manchester, MI3 9PT, U.K. Abstract--Potassium channel openers comprise a diverse group of chemical agents which open plasmalemmal K-channels. They show selectivity for smooth muscle, although K-channels in cardiac and skeletal muscle, neurones and the pancreatic fl-cell are also affected at relatively high concentrations. In addition, at least one endogenous K-channel opener of vascular origin---endothelium-derived hyperpolarizing factor--exists and in man plays a role in modulating blood vessel tone. The type of K-channel involved in the actions of both exogenous and endogenous K-channel openers is still uncertain, although a prime candidate in smooth muscle seems similar to the [ATPi]-modulated K-channel in the pancreatic fl-cell. This review focuses attention on the action of these agents in vascular smooth muscle and on the possible clinical exploitation of their powerful vasorelaxant properties. CONTENTS 1. Introduction 2. K-Channel Classification 2.1. K-channel subtypes 2.1.1. Voltage-gated channels 2.1.2. Ion-gated channels 2.1.3. Ligand-gated channels 2.1.4. Second-messenger/intracellular metabolite-gated channels 3. Synthetic K-Channel Openers 3.1. Chemical structures 3.1.I. Benzopyrans 3.1.2. Cyanoguanidines and related compounds 3.1.3. Nicotinamides 3.1.4. Thioformamide derivatives 3.1.5. Diazoxide and minoxidil sulfate 3.1.6. Niguldipine 3.2. Interactions with specific K-channels 3.3. In vitro pharmacodynamics 3.3.1. Effects on mechanical activity 3.3.2. Effects on 42K or S6Rb efflux 3.3.3. Effects on membrane potential 3.3.4. Effects on intracellular Ca 2÷ stores 3.3.5. Effects on transmitter release 3.4. In vivo pharmacodynamics 3.5. Clinical uses and potential 3.5.1. Essential hypertension 3.5.2. Angina pectoris 3.5.3. Cerebral vasospasm 3.5.4. Chronic occlusive arterial disease (COAD) 3.5.5. Myocardial ischemia 4. Endogenous K-Channel Openers 4.1. Endothelium-derived hyperpolarizing factor (EDHF) 4.2. Arachidonic acid and phospholipids 4.3. Calcitonin gene-related peptide 5. Conclusion Acknowledgements References

1. I N T R O D U C T I O N

237 238 238 238 239 239 240 240 240 240 241 241 241 242 242 242 245 245 245 246 246 246 246 247 248 249 249 249 249 250 250 250 251 251 251 251

the regulation o f b l o o d supply to specific regions. Regional changes in tone affect not only local b l o o d flow b u t also alter general systemic pressure. M a n y factors c o m b i n e to influence a given level o f vascular

Blood vessels exhibit varying degrees of tone which b o t h resists distension due to b l o o d flow a n d allows 237

238

G. EDWARDSand A. H. WESTON

tone and these range from the intrinic electrical activity of the vascular smooth muscle cells to extrinsic influences such as those exerted by locally-released neurotransmitters and mediators (see review by Longmore and Weston, 1989). Recently, attention has become focused on the role of potassium (K) channels in the modulation of vascular tone. The general role which changes in plasmalemmal K-conductance play in the modulation of excitability is well-understood. In the heart (Noble, 1984), central nervous system (North, 1989) and in smooth muscles (Bfilbring and Tomita, 1987), the opening of K-channels raises the membrane potential and shifts the cell into a condition in which it is less easily excitable. In neurones, the role of different K-channels within the same cell in controlling patterns of excitability has long been recognized (Hille, 1984). More recently, advances have occurred in understanding the detailed role of K-channels. This has resulted from a combination of factors, one of the most important of which has been the development of whole cell clamp and patch clamp techniques (Hamill et al., 1981) which have enabled K-currents to be measured in isolated single cells and in membrane patches. In parallel with this methodological breakthrough have been developments in the pharmacology of K-channels. The isolation and purification of toxins which block K-channels has been extremely important and several such agents have emerged, each relatively specific for a particular K-channel type (reviewed by Moczydlowski et al., 1988; Castle et al., 1989; Strong, 1990). Apamin (Habermann, 1984; Moczydlowski et al., 1988) charybdotoxin (GimenezGallego et aL, 1986; Moczydlowski et al., 1988) and dendrotoxin (Moczydlowski et al., 1988) have been most useful in confirming and assessing channel presence and function. In addition to these naturally occurring substances, the sulfonylureas, used clinically in diabetes mellitus as oral hypoglycemic agents are proving invaluable in the study of ATPdependent K-channels (Bailey et al., 1989; de Weille et al., 1989). Furthermore the development of Kchannel blockers, selective for cardiac muscle and potentially useful as Class III anti-arrhythmic agents has now reached the clinical trial stage (Colatsky and Follmer, 1989). Perhaps the most recent pharmacological development is the emergence of a new group of agents known as the K-channel openers. These substances, typified by cromakalim, pinacidil and nicorandil open K-channels in a variety of smooth muscles (Hamilton and Weston, 1989). Their availability, together with the methodological and other chemical developments already described has given a new insight into the role of K-channels in excitable cells and into their possible uses for the treatment of hypertension (Weston, 1990). The purpose of this review is to outline the developments in K-channel pharmacology which have already occurred and especially their relevance to vascular smooth muscle relaxation. This provides a basis for tile understanding of K-channels in the modulation of tone in vascular and other smooth muscles and for the clinical exploitation of this phenomenon.

2. K-CHANNEL CLASSIFICATION 2.1. K-CHANNELSUBTYPES The criteria for classification of K-channels into subtypes are derived from a combination of (a) channel gating properties, (b) channel conductance and (c) channel pharmacology. The properties of K-channels in smooth muscle are less wellunderstood than in other excitable systems. Contributing factors here include the doubtful suitability of cultured smooth muscle cells due to their rapid de-differentiation in culture and the harsh enzymatic conditions often necessary to separate cells acutely from their mother tissue. Based on their gating, conductance and pharmacological characteristics K-channels can be divided into four basic groups. Wherever possible, reference will be made to smooth muscle, although few data are available relative to other tissue systems. The nomenclature employed has been taken from Watson and Abbott (1990). For further details of K-channel types see Rudy (1988). 2.1.1. Voltage-Gated Channels

2.1.1.1. K w --T he delayed outward rectifier channel. This channel contributes to the macroscopic outward K-current observed in many cells and was probably first recognized in the squid giant axon (Hodgkin and Huxley, 1952) although it was not designated as such until relatively recently (Conti and Neher, 1980). In neurones (Conti and Neher, 1980) and in skeletal muscle (Standen et al., 1985), the channels have a conductance of 15-20 pS and are activated by membrane depolarization after a finite time delay. The current flowing through Kv rises sigmoidally on activation. Under a constant depolarizing stimulus, a slow, exponential inactivation occurs, requiring up to several seconds for completion. Blockade of Kv can be achieved using Cs, Ba and intracellular TEA. 4-aminopyridine (4AP) is also sometimes effective. In smooth muscle, a 4AP-sensitive outward Kcurrent probably carried by Kv has been identified in pulmonary artery (Okabe et al., 1987), rabbit jejunum (Benham and Bolton, 1983), bladder (Kl6ckner and Isenberg, 1985) and in intestinal smooth muscle (Ohya et al., 1986). In these tissues, the channel may open during the repolarization phase of slow electrical waves and it may also function in the termination of agonist-induced depolarization. 2.1.1.2. K A - - T h e transient outward channel. The current carried by KA is often known as 'the A current' (Ia) and was first described in molluscan neurones (Connor and Stevens, 1971). KA channels have a typical conductance of 15-20 pS and can be blocked by 4AP. They activate and inactivate very rapidly on exposure to a depolarizing stimulus. In neurones, the interplay between Kv and KA channels functions to control neuronal firing rate, latency and action potential repolarization (Hille, 1984). In smooth muscle, a current with properties similar to that carried by KA in other tissues has been described in pulmonary artery (Ohya et al., 1986), although its precise function in this tissue is unknown.

Potassium channel openers and smooth muscle relaxation 2.1.1.3. K;R--The inward rectifier channel. Historically, the earliest K-currents to be described exhibited outward rectification, a property which became regarded as 'normal'. The discovery of K-currents which increased in magnitude at membrane potentials more negative than those typical of resting cells gave rise to the term 'anomalous rectifier'. Such K m channels have been described in sea urchin eggs (Hagiwara and Jaffe, 1979) skeletal and cardiac muscle (Adrian, 1969; Noble 1984) and in neurones (North, 1989). The channels which exhibit a typical conductance of approximately 20 pS can be blocked by Cs, Ba or intracellular TEA. In intestinal smooth muscle, an inwardly rectifying K-current has been described (Benham et al., 1987) and this may in part contribute to the K conductance seen under resting conditions. An inward rectifier current has also been described in submucosal arterioles (Edwards and Hirst, 1988). In this tissue pronounced inward rectification was observed at resting membrane potentials, strongly suggestive of a role in maintaining resting K conductance. A similar K-current also exists in rat cerebral arteries (Edwards et al., 1988). 2.1.2. Ion -Gated Channels Although K-channels which can be gated by changes in Na concentration have been described (Bertrand et al., 1989), this section will only deal with those K-channels gated by Ca. At present three such broad groups of calcium-dependent K-channels have been described and these can be separated by their single channel conductance and their ease of blockade by a variety of pharmacological agents. 2.1.2.1. BKc,--The big conductance calcium-activated channel. BKc, channels have been described in a very wide variety of cells ranging from neurones (Meech, 1978) and skeletal muscle (Miller et al., 1985) to the pancreas (Findlay et al., 1985) and salivary glands (Maruyama et al., 1983). The channels typically exhibit large conductances (150-200 pS) and they can be blocked using extracellular TEA (Kd < 1 mM), quinine, Ba or charybdotoxin. The channels are also voltage-gated and are activated on membrane depolarization. In mammalian smooth muscle, such channels have been identified in intestinal and arterial smooth muscles (Benham et al., 1984; Bolton et al., 1985; Inoue et al., t985) in airways (McCann and Welsh, 1986), ureter (Shuba, 1981) and taenia caeci (Inomata and Kao, 1979). The channels almost certainly function to terminate excitatory processes which are triggered or maintained by an increase in [Cai2. ] and/or involve membrane depolarization. In this respect, they may be responsible for the early fast outward current which follows the upstroke of smooth muscle action potentials and for the spontaneous transient outward currents which can be elicited in tissues like rabbit portal vein (Benham and Bolton, 1986). 2. ! .2.2. IKc~--The intermediate conductance calciumactivated channel. IKca channels have been identified in red blood cells (Hamill, 1981) and in neurones (Herman, 1986). They are essentially not gated by JPT 48;2--1

239

voltage but exhibit an increased opening probability following an increase in [Ca2+ ]. Some inward rectification is also observed and typical single channel conductance is 18-50 pS. The channel is blocked by quinine, quinidine, TEA or charybdotoxin. Broadly similar channels have also been described in rabbit portal vein (Inoue et al., 1986), but in this tissue, single channel conductance is approximately 80 pS and the channels are open at resting potentials. This suggests a possible role in the maintenance of resting membrane potential. 2.1.2.3. S Kc,-- The small conductance calciumactivated channel. The SKca channel exhibits a typical conductance of 10--14 pS and was first described in cultured rat skeletal muscle cells (Blatz and Magleby, 1986) and in guinea-pig hepatocytes (Cook and Haylett, 1985). These channels are activated by an increase in [Ca 2+ ] and can be blocked by apamin. They exhibit little voltage sensitivity and in general these channels are more sensitive than BKca channels to [Ca2+]. An analogous current/channel has been described in portal vein (Inoue et al., 1985) and in bladder smooth muscle (Isenberg and Kl6ckner, 1986). 2.1.3. Ligand-Gated Channels Ligand-gated channels are modulated by the interaction of iigands with plasmalemmal receptors. For reasons which are not understood, cells which have been separated using enzymatic techniques and/or grown in culture may fail to respond to typical agonists. For this reason, relatively few ligand-gated channels have so far been described. 2.1.3.1. Acetylcholine and 5-hydroxytryptamine receptor-gated channels. At least two K-channels gated by th.e interaction of acetylcholine (ACh) with muscartalc receptors have been described. In the heart KACh is the channel activated by ACh and is responsible for the characteristic inhibitory effects of ACh seen in atrial muscle (Noble, 1984). In vertebrate neurones, a different channel designated KM is inactivated by ACh and is responsible for the characteristic excitatory effects of the transmitter in this tissue (Brown and Adams, 1980; Brown, 1988). Interestingly, KM is stimulated by fl-adrenoceptor agonists (Sims et al., 1988). In Aplysia 5-hydroxytryptamine closes (inactivates) a K-channel designated KS.HT, an effect which underlies the excitatory effects of this neurotransmitter (Siegelbaum et al., 1982). 2.1.3.2. Otl-Adrenoceptor and NANC-receptor gated channels. In guinea pig taenia caeci, the relaxant actions of noradrenaline via ~-adrenoceptors and those of the non-adrenergic, non-cholinergic (NANC) inhibitory transmitter are inhibited by apamin (Banks et al., 1979; Shuba and Vladimirova, 1980). Since this bee venom toxin is a selective blocker of SKca channels (Burgess et al., 1980; Blatz and Magleby, 1986), it is reasonable to conclude that such channels exist in the taenia and that they can be activated via cq-adrenoceptors and by the receptor for the NANC transmitter.

240

G. EDWARDSand A. H. WESTON

2.1.4. Second-Messenger/Intracellular Gated Channels

Metabofite-

This final category of K-channels is not really a category in its own right. It simply serves to describe further some of the characteristics of K-channels which might also be ligand- and/or ion- and/or voltage-gated. 2.1.4.1. Inositol trisphosphate- and diacylglycerolgated channels. Inositol trisphosphate (IP3) inhibits a Ca-dependent K-current in rabbit nodose ganglia (Weinreich, 1986) and together with inositol 1,3,4,5 tetrakisphosphate (IP4) activates Ca-dependent Kchannels in lacrymal acinar cells (Morris et al., 1987). Diacylglycerol (DAG) inhibits both Ca and voltagedependent K-currents in a variety of cells via the activation of protein kinase C (Kaczmarek and Strumwasser, 1984; Baraban et al., 1985; Farley and Auerbach, 1986). Evidence has been obtained that both IP 3 and resultant intracellular Ca mobilization induce a transient hyperpolarization via a Cadependent K-current followed by a sustained depolarization via blockade of a DAG-sensitive K-current (Higashida and Brown 1986).

2.1.4.2. Cyclic A M P - g a t e d channels. In several tissues an increase in cAMP concentrations modulates Kcurrents by activation of cAMP-dependent protein kinases. In Aplysia the 5-HT-induced inhibition of the Ks..T channel already mentioned (Section 2.1.3.1) is mediated by this intracellular mechanism (Schuster et al., 1985). Similar cAMP-gated channels have been described in a variety of tissues (Kaczmarek and Strumwasser, 1984; Ewald et al., 1985; Avenet et al., 1987). 2.1.4.3. A TP-dependent channels. K-channels which close as [ATPi] increases have been described in cardiac muscle (Noma, 1983) and in skeletal muscle (Spruce et al., 1985, 1987). Under experimental conditions in these tissues, an increase in the ATP: ADP ratio closes the channels; under normal physiological conditions the channels are in a closed state. Typical channel conductance lies in the range 40-80 pS. Similar KATP channels have also been described in pancreatic/J-cells (Cook and Hales, 1984). Although the channel open state probability is also low in this tissue (approximately 99% of KATPchannels in pancreatic/J-cells are thought to be closed under normal resting conditions) the high density of the channels in /?-cells probably means that sufficient numbers are

open at normal plasma glucose levels for this channel to be a major determinant of the resting membrane potential (Cook, D. L. et al., 1988) An increase in [ATPi] (when plasma glucose levels are high) closes further KATP channels with resultant membrane depolarization, Ca entry and insulin secretion. The opening of KATVin pancreatic/j-cells can be blocked by low concentrations of oral hypoglycemic drugs like glibenclamide (Henquin and Meissner, 1982; Ashford et al., 1988), agents which are also effective blockers of KATp in other cells, albeit at higher concentrations. Preliminary experiments suggest that similar channels may also be present in the CNS (Sturgess et al., 1985; Ashford et al., 1988). The recent demonstration of KATP channels in vascular smooth muscle (Standen et al., 1989) and their possible activation by cromakalim is discussed in detail in Section 3.2.

3. SYNTHETIC K-CHANNEL OPENERS 3.1. CHEMICALSTRUCTURES Synthetic K-channel opening drugs encompass seven distinct chemical classes of which the benzopyrans are the most widely studied group (Table 1). Variants on the cyanoguanidines, typified by pinacidil, are also under world-wide study. Developments within these two classes contrast with those based on nicorandil which possesses only a small number of active structural variants. The most recently described novel structural group, characterized by the thioformamide RP-49356, has led to the synthesis of a large number of derivatives, the activity of which is currently under detailed scrutiny. 3.1.1. Benzopyrans

The current world-wide standard for a potassium channel opener is cromakalim (formerly BRL 34915), a racemic mixture of two enantiomeric forms designated BRL 38226 and BRL 38227 (Fig. 1). The chemical development of cromakalim was described by Evans et al. (1983) and by Ashwood et al. (1986). Largely for regulatory considerations (Barber, 1989; Seymour, 1990), development of cromakalim recently ceased in favor of the active enantiomer BRL 38227, now given the generic name lemakalim (Fig. 1). Structure activity studies have shown the critical importance of the carbonyl substituent in the pyrrolidine ring, together with the nature of the substituent

TABLE1. Chemical Classes of Potassium Channel Opener Under Development Chemical class Typical member Origin benzopyran (a) with chiral centres at C3 and C4 (b) without chiral centres at C3 and C4 cyanoguanidine nicotinamide thioformamide pyrimidine oxide benzothiadiazine dihydropyridine

lemakalim(cromakalim)

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by ATP is reduced in the presence of ADP. This has led to the conclusion that it is the ratio of [ATPi]:[ADPi] rather than [ATPi] itself which effectively determines the open state probability of the ATP-sensitive K-channel (Kakei et al., 1986; Dunne and Petersen, 1986; Misler et al., 1986; Findlay, 1988; Corkey et al., 1988). A further complication is that ATP-sensitive K-channels inactivate (and are incapable of opening) in the total absence of ATP, probably as a result of channel dephosphorylation (see Ashcroft, 1988; Rorsman and Trube, 1990). This is suggested by the requirement for the presence of Mg 2+ and a low concentration of ATP at the intracellular side of the membrane to maintain the channel in an active state which will allow opening (OhnoShosaku et al., 1987; Findlay, 1987a,b). Thus in isolated membrane patches, maximal opening of the ATP-sensitive K-channel can be expected to occur either immediately after removal of ATP or when there is a low concentration of ATP at the cytoplasmic side of a patch rather than when ATP is completely absent. In patches isolated from cardiac myocytes, pinacidil or RP49356 at concentrations of 300/~ M can open channels closed by a low [ATPi] (Escande

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Nlguldipine FIG. 7. Structural formula of niguidipine, a dihydropyridine calcium antagonist with K-channel opening properties.

244

G. EDWARDSand A. H. WESTON

et al., 1989; Thuringer and Escande, 1989; Martin and Chinn, 1990), and also in the absence of ATP if the channel has not become inactivated (Thuringer and Escande, 1989). In vitro, nicorandil, pinacidil and cromakalim (1-100pM) shorten cardiac action potentials (Cain and Metzler, 1985; Osterrieder, 1988). In similar concentrations these agents abolish inward rectification by K ÷ channels in the heart, an effect apparently sensitive to intracellular ATP (Kakei et al., 1986; Iijima and Taira, 1987; Osterrieder, 1988). It therefore seems likely that the opening of ATP-sensitive channels in the heart might be responsible for the effects of the potassium channel openers. Recently, Thuringer and Escande (1989) have shown that RP49356 acts by decreasing the sensitivity of KA~P to [ATPi]. In RINm5F cells, RP 49356, pinacidil and cromakalim (each 200 pM) can open channels which are under inhibition by a low concentration of ATP (0.1 mM) but when the channels are inhibited by more physiological levels of [ATP~] (4mM) and [ADPi] (I mM), even extremely high concentrations of these agents are barely effective (Dunne, 1990). In contrast, diazoxide (100 #M) which is a potent, selective opener of the KATP in pancreatic fl-cells (Trube et aL, 1986; Dunne et al., 1987; Sturgess et al., 1988), almost totally reverses the inhibition of opening of the ATP-sensitive K-channel by 4 mM ATP in the presence of 1 mM ADP (Dunne, 1990). In mouse isolated pancreatic fl-cells, diazoxide (100 p M) almost abolishes insulin secretion whereas cromakalim or pinacidil (each 500pM) produce only about 40 or 60% inhibition, respectively; minoxidil sulfate paradoxically stimulates insulin secretion (Garrino et al., 1989). Such data contrast with the effects of these potassium channel openers on the rat isolated portal vein in which diazoxide is the least potent of these K-channel openers at inhibiting spontaneous mechanical activity; maximal effects are produced by l pM cromakalim, 3pM pinacidil, 10pM minoxidil and 100 pM diazoxide (Newgreen et al., 1990). Thus, although diazoxide apparently shows no tissue selectivity, other potassium channel openers seem to be far more potent in vascular smooth muscle than in any other tissue. The different rank orders of potency of these agents in the pancreas and in vascular smooth muscle suggest that either the channel through which they exert their effects or their channel recognition site is different in these two tissue types. In spite of these differences, an action by cromakalim at a smooth muscle K-channel similar to KATp in pancreatic fl-cells is consistent with the finding of Standen et al. (1989) that a low concentration of cromakalim (1 pM) is capable of increasing the open-state probability of a K-selective channel which is closed by l mM ATP and inhibited by glibenclamide. The high conductance of the K-channel in these experiments (135 pS at 0 mV with 60 mM extracellular K + and 120raM intracellular K÷; Standen et al., 1989) contrasts with the much lower conductances of ATP-sensitive K-channel in either the heart or the pancreatic fl-cell (50-80pS with symmetrical 140mM K ÷ gradient; see review by Ashcroft, 1988). Although the vascular ATP-sensitive K-channels were inhibited by glibenclamide (Standen et al., 1989), the high concentration used (20 pM) was

far in excess of the concentration which selectively inhibits ATP-sensitive K-channels in pancreatic fl-cells (0.1-1 riM). The selectivity of such a high concentration of glibenclamide for a single channel type in vascular smooth muscle cells is questionable, casting doubt over whether the effect of such concentrations of glibenclamide on vascular smooth muscle really indicates the involvement of an ATP-sensitive K-channel similar to that described in the heart or pancreatic fl-cell. Recently Gelband et al. (1990) have shown that glibenclamide can block BKca. There have been suggestions that cromakalim opens Ca2+-dependent K-channels in vascular smooth muscle. Kreye et al. (1987) found that cromakalim stimulated efflux from rabbit aorta preloaded with the K-substitute 86Rb. Since this effect was inhibited by Ca2+-channel blockers (D600, nifedipine or trifluoperazine) the authors concluded that the channel opened by cromakalim was calcium dependent. Nakao et al. (1988) drew a similar conclusion from their observations that cromakalim was unable to hyperpolarize guinea-pig mesenteric artery or vein in a calcium-free medium. Other reports have indicated that cromakalim increases the open state probability of a large conductance Ca2÷-dependent K-channel in cultured aortic smooth muscle cells or following incorporation of these channels into planar lipid bilayers (Kusano et al., 1987; Gelband et al., 1989, 1990). Preliminary reports also indicated that BKc~ may be the site of action of pinacidil (Hermsmeyer, 1988). Ca2+-dependent K-channels are subdivided into three groups according to their conductance (see Section 2.1.2). BKc~ and IKc~ are selectively blocked by charybdotoxin derived from the venom of the scorpion Leiurus quinquestriatus (Miller et al., 1985). SKc~ is selectively blocked by the bee venom, apamin (see review by Habermann, 1984). However, attempts to block the cromakalim-activated K-channel with either charybdotoxin or apamin have failed (Strong et al., 1989; Weir and Weston, 1986). Beech and Bolton (1989) investigated the effects of a range of K-channel blockers on the current flowing through a variety of voltage or calcium-dependent K-channels in single cells freshly isolated from rabbit portal vein. A similar rank order of potency was found for the ability of the blockers to inhibit either the delayed outward rectifier current carried by Kv or the current induced by cromakalim. Since, under their conditions, the channels opened by cromakalim were not voltage-sensitive it was suggested by Beech and Bolton (1989) that cromakalim could increase the open-state proability of Kv by a mechanism which inhibited its voltage sensor. Low concentrations of cromakalim ( < l p M ) stimulate 42K efflux from preloaded tissues but have very little effect on 86Rb efflux; higher concentrations (> 1/aM) increase both 42K and 86Rb efflux (Bray and Weston, 1989). It therefore appears that cromakalim is capable of opening two types of K-channel, only one of which is permeable to 86Rb. Nakao et al. (1988) also found two components of the membrane hyperpolarization induced by cromakalim in guineapig mesenteric vein, only one of which was sensitive to Mn2+; of these, only the Mn2÷-insensitive

245

Potassium channel openers and smooth muscle relaxation component could be detected in guinea-pig mesenteric artery. These findings, together with the variety of K-channel types which have been shown to be opened by cromakalim, strongly suggest that the potassium channel openers are not selective for a single channel type. The different potencies of these agents in different tissues might therefore reflect the presence of different K-channel types or the varied relative densities of the specific K-channels which are the targets for the action of these substances. The recent discovery by Okabe and coworkers (1990) that cromakalim inhibits Ca ~+ currents in whole cells freshly isolated from rat portal vein, and evidence that pinacidil might also inhibit Ca 2+ channels in guinea-pig mesenteric arteries (Nakashima et al., 1990) implies that relaxant effects of potassium channel openers might not even be restricted to an action on K-channels. Furthermore the finding that pinacidil reduces C1- conductance in rat mesenteric arterioles (Videbaek et al., 1990), indicates a further property which could contribute to the total pharmacology of these agents.

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3.3. IN VITRO PHARMACODYNAMICS As a group, the potassium channel openers exert their inhibitory effects on vascular (and other) smooth muscles in a very characteristic fashion and this forms the basis for the screening of novel molecules by the pharmaceutical industry. Some of the in vitro characteristics of K-channel openers are illustrated in Fig. 8. 3.3.1. Effects on Mechanical Activity In portal veins, both cromakalim and pinacidil reduce the amplitude and frequency of spontaneous tension waves with final abolition of all mechanical activity (Hamilton et al., 1986; Southerton et al., 1988; Longman et al., 1988; Cook, N. S. et al., 1988a). In vascular tissues precontracted with KC1, the potassium channel openers show differential inhibitory effects, dependent on the concentration of added KC1. If low concentrations of KCI are used (

Potassium channel openers and vascular smooth muscle relaxation.

Potassium channel openers comprise a diverse group of chemical agents which open plasma-lemmal K-channels. They show selectivity for smooth muscle, al...
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