Calcium sensitisers: mechanisms of action and potential usefulness as inotropes

Time for primary review 28 days. Heart failure remains a common and lethal condition, with treatment options which are less than satisfactory. Epidemiological studies [1–5]indicate an overall prevalence in the population of 1–3%, while mortality for all grades of heart failure is approximately 25% at 2 years and 50% at 4 years. Severe heart failure has a 2 year mortality as high as 75%. Although some cases are caused by correctable problems such as hypertension or valve lesions, the majority result either from loss of functioning myocardium due to ischaemic damage or from ineffective contraction of partially ischaemic or otherwise functionally disabled myocardium. Therapy for heart failure thus has two main objectives [6–12]. Firstly, treatment of the failing pump and its main physiological consequences, fluid retention and vasoconstriction. Secondly treatment of the common associated problems which may be either primarily involved in causing heart failure (e.g. ischaemia) or which may be secondary complications (e.g. arrhythmias). Therapies which are currently available can reduce morbidity and have also achieved some reductions in mortality [13–18]. However, most of these treatments, including diuretics, angiotensin converting enzyme (ACE) inhibitors, vasodilators and beta-blockers, are aimed at alleviating the fluid retention and vasoconstriction associated with heart failure, rather than the primary cause. In principle therapy aimed at supporting the pump function of the heart might also be expected to have a major role in the treatment of heart failure. In practice the benefits of positive inotropic agents have been hard to demonstrate. This therapeutic approach has a long history, beginning with Withering's introduction of digitalis in 1785 [19]. But cardiac glycosides have multiple actions and there has been protracted controversy over whether these drugs are beneficial to patients with heart failure in sinus rhythm. Recent results do point to a benefit [20], even in the presence of treatment with ACE inhibitors [21], but glycosides remain drugs with a relatively narrow therapeutic window and significant side-effects. Other inotropes have fared worse: phosphodiesterase (PDE) inhibitors have been virtually withdrawn after being found to increase mortality [22, 23]. The recent disappointments with positive inotropic drugs have been so great that many now question whether they are likely to have a significant role in the treatment of chronic heart failure [24]. In this review we suggest that the deficiencies of currently available inotropes may be, at least in part, a consequence of their cellular mode of action and that new agents utilising fundamentally different cellular mechanisms to increase force may find a useful role in heart failure therapy in the future. The drugs known as “calcium sensitisers” are a diverse group of compounds which are beginning to access such novel cellular pathways. We provide an overview of the theoretical background and experimental data relating to the effects of calcium sensitisers on heart muscle and we comment on the potential therapeutic relevance of this information. Since most calcium sensitisers are still at the experimental stage of development, it is clearly not yet possible to give a firm answer to the question of whether this inotropic approach will actually turn out to be an improvement on those we already have. Nevertheless, there seem to be grounds for optimism and we suggest that this area of investigation deserves more attention. Appreciation of the advantages and disadvantages of various inotropic agents requires an understanding of heart muscle activation. This section briefly summarises some important points. Heart muscle contracts when an action potential activates an inward Ca2+ current across the surface membrane. The Ca2+ entry associated with this current is small, but it is sufficient to trigger a much larger Ca2+ release from the sarcoplasmic reticulum. The resulting short-lived increase in myoplasmic free Ca2+ concentration ([Ca2+]i) — known as the Ca2+ transient — causes Ca2+ binding to troponin C. This in turn causes conformational changes in various myofilament proteins, with the result that crossbridge attachment is stimulated and contraction occurs. A conventional view of the latter stages of this process is that Ca2+ binding alters the interactions of troponin C with troponin I and troponin T in such a way that tropomyosin moves from its position in the actin filament groove, allowing myosin heads (“crossbridges”) to interact with binding sites on the actin filament. In this steric hindrance model, developed force increases when [Ca2+]i rises because the exposed actin binding sites recruit more crossbridges. In recent years this mechanism has been challenged by an alternative idea which suggests that crossbridges are either weakly or strongly bound to actin. In this model, the main effect of increased [Ca2+]i is to change the rate at which myosin heads convert from a weak attachment (WXB in Fig. 1) to a strongly bound state which produces force (SXB). This is known as the turnover kinetics model of muscle activation [25]. The evidence for this model comes from biochemical [26], mechanical [27]and X-ray diffraction studies [28]showing that crossbridges are weakly bound to the actin site in resting muscle, a finding which is incompatible with the simple steric hindrance model. Brenner [29]devised a method for measuring conversion rate of weakly bound to strongly bound crossbridges (designated fapp — the apparent rate of attachment of strongly bound crossbridges) and showed that it was sensitive to [Ca2+]i. Thus in the turnover kinetics model the effect of Ca2+ binding to troponin-C is to accelerate the conversion of crossbridges from the weak to the strongly bound form (see Fig. 1). Schematic diagram of processes involved in activation of heart muscle. Key: Ca — intracellular calcium; Tr — troponin; WXB — weakly bound cross-bridges (non-force-producing); SXB — strongly bound cross-bridges (force-producing). fapp is the apparent rate constant for conversion of WXB to SXB; gapp is the corresponding reverse rate constant. (i)–(v) represent possible sites at which a calcium sensitiser might act (see text). In intact heart muscle the Ca2+ transient rises in 10–20 ms and declines over 200–400 ms [38]so that tension never reaches a steady state with [Ca2+]i. Nevertheless it would be useful to be able to determine n and Ca50 in intact preparations and there have been various experimental attempts to determine these values [39]. A simple method is to modify developed force by changing extracellular Ca2+ concentration ([Ca2+]o) and to determine the peak [Ca2+]i and force at each level of [Ca2+]o, (Fig. 2A). Peak [Ca2+]i is then plotted against peak force (Fig. 2B). Although this method does not give the correct absolute values of n and Ca50, it has allowed some important observations to be made. As [Ca2+]i is elevated, force initially rises steeply and then rises more slowly, with the general form shown in Fig. 2B. However, large increases in [Ca2+]i cause a fall of force, which would not be predicted from skinned fibre data. This is the phenomenon of “Ca2+ overload” (Fig. 2A and B) [40, 41]. Along with reduced force, several other phenomena are also seen in Ca2+ overload — spontaneous Ca2+ release [i.e. Ca2+ release not triggered by an action potential — indicated by the arrow in Fig. 2A (iv)], aftercontractions and triggered arrhythmias. All these phenomena are now known to be caused by a tendency for the sarcoplasmic reticulum to release Ca2+ spontaneously when its Ca2+ content becomes excessive [42, 43]. From the present perspective, the important point is that any intervention which has the capacity to sufficiently elevate [Ca2+]i — including all conventional positive inotropic agents — is likely to reduce force at high concentrations and also to cause arrhythmias. Furthermore since sympathetic activity itself raises [Ca2+]i, doses of inotropic agents which by themselves are within the therapeutic range, may contribute to Ca2+ overload when endogenous sympathetic activity occurs in the patient. Relation between [Ca2+]i and force in heart muscle during inotropic interventions. Panel A. [Ca2+]i transients and force in intact heart muscle at 4 levels of [Ca2+]o (modified from Ref. [40]). The arrow in (iv) identifies a spontaneous elevation of [Ca2+]i. These are characteristic of Ca2+ overloaded preparations. Panel B shows a typical relation between peak [Ca2+]i and peak force; the marked points correspond to the activation levels in A. Note that at the highest levels of [Ca2+]i, developed force starts to decline; this phenomenon is not observed when the steady state force-Ca2+ relation is determined in skinned fibres (e.g. Figs. 3 and 4). As is now well-known, all major classes of inotropes currently in use act by elevating systolic [Ca2+]i, i.e. by increasing the amplitude of the calcium transient [44]. However, although they produce a similar end result, a variety of cellular mechanisms are accessed by different drugs. Cardiac glycosides block the Na+ pump, the elevated [Ca2+]i being secondary to a rise in [Na+]i and involving the Na+/Ca2+ exchanger. β-adrenergic agents and inhibitors elevate and cause of intracellular sites on In the of of the causes increased the action of a and inhibitors reduce the activity of the rate of which are as a consequence of the increased the surface Ca2+ increases Ca2+ entry and sarcoplasmic reticulum Ca2+ and and — which accelerate the rate [Ca2+]i is elevated the sarcoplasmic reticulum becomes more with Ca2+ and more Ca2+ when triggered by an action potential (Fig. Although conventional inotropes increase force by increasing [Ca2+]i, there are in alternative of the end (see Fig. is to increase the force of which the muscle is This either the of crossbridges to force increased (e.g. the force crossbridge to increase mechanism in or crossbridges for a of the crossbridge mechanism which increase force would be for the to more Ca50 or force [25]. We will not this since there is evidence for its at In general an increase in n might be because activity then be on and with less Ca2+ This mechanism be an important for action in the as understanding of muscle activation force also be by increasing the of the to [Ca2+]i In this several different mechanisms be These are with to a such as that shown in Fig. A in Ca50 — that is an increase in Ca2+ — be in at least different increase in the of troponin C for increase in the with which the troponin increases the rate of crossbridge and (iv) A change in crossbridge force be by either an increased rate of attachment or a rate of These are in Fig. which shows a and two the effect of fapp or cause increased Ca2+ a of and also increased force a of with effect on A change in the between crossbridges and troponin C for increase in causes increased Ca2+ and also increases in recent years have a large of which are able to the changes in intracellular muscle troponin I myosin C and many drugs of which the known is Since these the force in to a [Ca2+]i they can be as the Ca2+ As in the it is to between which increase Ca2+ and those which increase force In practice many produce although or the other the conventional steric hindrance model of muscle activation a for these effects is that it two potential by which the force of cardiac contraction be of [Ca2+]i, and in some effects are observed However, as in the turnover kinetics model of activation effects may from a intervention Fig. is that the “Ca2+ is in the to compounds which increase Ca2+ force or in skinned i.e. as a for Ca2+ agents which force by increasing Ca2+ might problems associated with Ca2+ Thus there has been in inotropic drugs on this mechanism of The finding that it was possible to Ca2+ with compounds was a known The effect was in a skinned fibre the was to the Since that several other compounds with similar activity have been including and of the actions of these compounds in intact preparations and in is because the of the effects on Ca2+ and in intact heart muscle between different and also because has several effects in intact muscle These of calcium in heart muscle as as in of cellular mechanisms it is clearly to have compounds which increase force in intact muscle by the Ca2+ of the compounds currently — which does cause some in produces the most positive inotropic effect due to increased Ca2+ yet observed in intact cardiac muscle has also been in a variety of preparations. is a which has marked calcium effects causing effects on calcium in intact muscle are not yet In the we the evidence for the various potential mechanisms of action of agents which Ca2+ We also the of of and other to the potential of these agents in chronic heart failure. of the of the we on other compounds As the of an intervention which increases Ca2+ is that it causes a to the in the relation between force and [Ca2+]i, i.e. there is a in the [Ca2+]i which produces 50% force in Fig. Fig. 4 shows a of a relation from skinned muscle in the presence and of Note that Ca50, increases force and the of the from to The have been a model of the shown in Fig. to the data (see for was that the of troponin for Ca2+ was constant in the presence of and that the either fapp or The was by increasing fapp or by gapp by the In this section we the possible sites of action which produce these effects and which of the mechanisms in Fig. be of on the relation between force and points from skinned muscle, The are from the in Fig. 3 to the data points. the gapp was to and the then has the gapp and at the values and the shown with and was fapp was constant and gapp allowed to evidence suggests that available Ca2+ sensitisers do not increase the of troponin C for Ca2+ in Fig. this the of Ca2+ binding to either troponin C or to the in which of crossbridges was an increased Ca2+ binding with troponin C in the presence of sensitisers have not been to while Ca2+ binding to to increased binding in the presence of concentrations of Furthermore increased of troponin C for Ca2+ the mechanism of action it would be expected to cause a in the increased force at [Ca2+]i is is that a sensitiser might increase the force for a level of Ca2+ bound to troponin but in the of a change in the of fapp in Fig. of this are but an alternative approach is to whether sensitisers can interactions in the of troponin and that the rate of in the of troponin and was 2 in the presence of also that actin increased their of myosin in the presence of the in an in these that can interactions in the of troponin and tropomyosin and suggest that mechanism may be i.e. an increase in the of These have been in several either with [Ca2+]i so that Ca2+ bound to troponin is after a which troponin I and the to All these studies that the on interactions rather than on the A of studies have the possible of increased fapp or gapp or and the rate of tension after a and also the at which muscle was at a processes in the presence of This suggests that crossbridge are in the presence of that is The was also in this and was but at force levels of Since is to it was that the most likely effect of was to reduce the rate of in a skinned cardiac when Ca2+ was with a Ca2+ found that either effect or caused a of such would to be by This result suggests that gapp is by that has a site of action on crossbridge kinetics it is likely to increase the rate of tension in skinned fibres from a state with found that concentration increased the rate of force with an increase in the grounds that found change in they that might increase some of these results are and it is not yet possible to a on whether increases fapp or A of to the of these results with is by the recent finding that is itself Since many Ca2+ sensitisers are also inhibitors (see they may elevate and thus have effects similar to β-adrenergic including elevation of [Ca2+]i. is important to the effects of apparent Ca2+ sensitisers in intact cardiac preparations while measuring [Ca2+]i. can whether or all of the inotropic effect of a sensitiser is in to increased Ca2+ is also important to that a effect is in in intact as to skinned preparations. it is possible that a effect be in an intact the not the surface or effects in the intact results in intact muscle an inotropic effect which is due to increased calcium of the The effects of this be relatively in the intact muscle preparations since the Ca2+ transients reduced in the presence of the significant would be expected to increase the Ca2+ force is increased by [Ca2+]i, a of available crossbridges are this mechanism it is expected that more force is associated with since is by Ca2+ and by crossbridge Thus the for a novel inotropic is whether the force of is more or less than for a simple such as In the the increase of force the more a novel would be, since in ischaemic heart there is a capacity to increase the of and the heart produce measuring force of in may not be of effects on the the of many force for long with but the muscle or of the of an thus be on muscle which is force and However, studies to have this approach to inotropic drugs. important theoretical is that in a model of crossbridges (e.g. Fig. 1) force is to while is to Thus is to an increases the of proteins, the is that it that the of cross-bridges the are that in an intact heart this be since changes in may also Ca2+ some it may be possible to the force Although this is an for the the concentration of to this result in skinned muscle was much than to be in cardiac muscle. studies have shown that for intact in which was by drugs with activity increased the of muscle by with either or on skinned fibres have the although the increase in was and seen at or force levels Since associated with Ca2+ release and is in skinned fibres all the have been these results that the most important of increased in intact preparations from reduced Ca2+ Other studies have been in as an of In a was found between the by inotropic doses of Ca2+ or However, since with this result is likely to the mechanisms of with this a of has the as but is the with all the calcium activity and relatively a increase in developed force with change in A recent this experimental approach to intact so that any effects of on would be in muscle with crossbridge was area a and to of the increased force but not then exposed to several of which and was reduced In this a increase in this result is it that although this may not the of it can increase of myocardium by there is a for this finding in that studies on skinned cardiac indicated that the in by elevated of the main for of force in heart muscle Although these results the that Ca2+ sensitisers might be of increasing of in it is not yet to this improvement from Ca2+ as to crossbridge studies of effects on are since compounds which increase force while also increasing be useful in the A finding in intact preparations is that compounds which increase force by increasing calcium also the rate the to peak force is while shows high concentrations there can be of the of and even a failure of force to to resting levels in the on rate are significant since they may to mechanisms of action and also because of be in (see In of be caused by several of [Ca2+]i, of Ca2+ from or of crossbridges. that the Ca2+ transient was with mechanism skinned fibres in which Ca2+ was a Ca2+ these not it although this might have been caused by of the In a similar on and found change in and crossbridge in the presence of the Thus the most likely cause of of in the intact would to be mechanism even the overall binding of Ca2+ to troponin to be by sensitisers (see of Ca2+ from troponin be either a effect on troponin in or the result of interactions on the as The main causing cardiac in is the relatively between the and the Thus it is important that the be as possible during at elevated heart the of becomes for cardiac so it is important that be as as possible to these suggest that an which or increased would be since in some patients heart failure is associated with of this it has been that calcium inotropes might be rather than in heart failure However, potential effects on be against the other possible advantages of they an inotropic effect with a increase in and (see then the overall advantages be since is of primary in a of it is important to that concentrations of which useful effects (e.g. increase in have been found to have effect on the of large increases in force (e.g. are associated with a of a on showed that while there was a of with was apparent when the allowed to In in studies have that sensitisers are able to increase cardiac (see also be data is in of However, a increased which is the major to in many As many currently available sensitisers also have Since this effect causes of it is possible that a of the two be useful in it might be to use of sensitisers with drugs which the heart are of the inotropic effects by calcium sensitisers in cardiac muscle are to be useful effects in Although muscle, may be by or so it is important that an is beneficial at least not to these as of the inotropic effect to and is caused by of intracellular and the effects of several inotropes in cardiac muscle and in muscle exposed to or a the results similar for inotropes by increasing [Ca2+]i and for calcium sensitisers A sufficient inotropic by either of reverse the inotropic effect of inotropes this was achieved by increasing the Ca2+ transient the inotropic effect of was with a in Ca2+ transient amplitude In force is due to intracellular with a from classes of inotropes relatively ineffective in force in this that force by mechanisms the point of action of all the inotropes Thus sensitisers can reverse at least of the major mechanisms causing force in muscle and they do so in a way which is Ca2+ and less likely to arrhythmias. In many of a of is of or but in the heart even a can have Although can be caused by many mechanisms associated with of in the current [Ca2+]i is likely to be already in muscle conventional inotropes it still is thus not that have been a with positive inotropic therapy in sensitisers would be expected not to increase and thus might at least some of these However, in which to this point associated with increased in the heart increased by [Ca2+]o and by a variety of to the potential of inotropes in is clearly studies that it is possible to in and failing with increase in at least in the In can increase cardiac in in a model of heart failure Since also increased in these any effect due to of was clearly by the positive inotropic effect of the The was also in a model of heart failure, it and as as systolic function evidence thus suggests that current sensitisers can produce an inotropic effect in muscle and also have seem to be at least than conventional inotropes with to and theoretical suggest that they may advantages in this increased cardiac has been in heart failure evidence of and of damage would be the way to reduce the of heart failure and to its these effects are not yet it that such as and will that there will be of patients with or heart failure. In such cases it would be useful to have agents which of calcium a novel approach to this which has several theoretical has not yet been in the The of the that its activity is with of the mechanisms involved in cardiac muscle force remains although great are now being with the of current inotropes increase force by increasing [Ca2+]i, but some such as the sensitisers in this are beginning to access the for increasing force in the myofilament As we have there are for that this approach to positive may have than In the the to which therapy can an on heart failure will be determined by the to which it can be shown to beneficial changes to of and This in turn on and the stage at which a that treatment be Although there have been it is that we not the by which Ca2+