Excitation-contraction coupling in myocardium

INTRODUCTION — Excitation-contraction (E-C) coupling refers to the series of events that link the action potential (excitation) of the muscle cell membrane (the sarcolemma) to muscular contraction. Although E-C coupling in myocardium is similar in many ways to skeletal muscle and smooth muscle, there are also critical differences. The cyclical nature of cardiac contraction and the importance of myocardial relaxation to cardiac pump function requires that any discussion of E-C coupling also consider the events terminating the muscle twitch as an integral part of the subject. Modulation of muscular function is said to affect inotropy (the speed and strength of muscular contraction) or lusitropy (the ability of the muscle to relax). Increased knowledge about E-C coupling has been a key to understanding both the inotropic and lusitropic states, and it continues to be useful in developing improved therapy for heart failure and cardiogenic shock.

This is a brief review of cardiac excitation (the myocardial action potential) followed by a description of muscular contraction. E-C coupling is then presented as the transduction of a membrane signal (the action potential) to an intracellular effector (the contractile apparatus) by way of a second messenger (intracellular free calcium [Ca²⁺]).

MYOCARDIAL ACTION POTENTIAL — The resting membrane potential of the myocardial cell is cell interior negative (-90 mV), and is primarily determined by the ratio of intracellular-to-extracellular potassium as predicted by the Nernst equation. The action potential is a temporary depolarization of the membrane. It is caused by transient changes in membrane conductance of several charged ions, especially sodium, due to the opening and closing of ion-specific channels in the membrane. This process can be summarized as follows (figure 1 and movie 1) [1]. (See "Cardiac excitability, mechanisms of arrhythmia, and action of antiarrhythmic drugs".)

●Rapid depolarization (phase 0) occurs when the resting cell is brought to threshold, leading sequentially to activation or opening of voltage-dependent sodium channels, rapid sodium entry into the cells down a favorable concentration gradient, and a cell interior positive potential that can approach +45 mV. The marked depolarization results in voltage-dependent inactivation of the sodium channels and cessation of the inward sodium flux. Calcium channels also open during depolarization but the onset of the inward calcium flux is much slower.

●Phase 1 repolarization is primarily due to inactivation of the sodium channels with abolition of the inward sodium current.

●This is followed by a plateau in phase 2 in which continued slow calcium entry into the cell balances the electrical effect of potassium loss, as more voltage-dependent potassium channels open up. The voltage in the plateau phase is sufficient to maintain the sodium channels in the closed, inactive state.

●The cessation of the calcium current and a further increase in potassium current leads to rapid completion of repolarization with return to the resting potential (phase 3).

●Normal cell sodium and potassium activities are restored by the Na-K-ATPase pump, which extrudes the sodium that entered during depolarization and pumps in the potassium that was lost during repolarization.

The action potential at a specific region of membrane is conducted along the membrane and depolarizes neighboring regions, causing their thresholds to be reached, thereby generating another local action potential. This process conducts the action potential as an electrical signal along the muscle fibers of the heart. Linear propagation of the action potential occurs rapidly because the myocytes along muscle fibers are electrically continuous. An action potential can also cause the depolarization of a neighboring fiber, and the electrical impulse occurs as a "wave" through the heart.

The regulation and process of depolarization is somewhat different in pacemaker cells. Each normal heartbeat is initiated by pacemaker cells in the sinoatrial node, generating a slow, calcium ion-mediated action potential. These cells manifest continuous variability of their resting membrane potential in a cyclical fashion (ie, spontaneous automaticity), generating an action potential every time the resting potential reaches threshold (-40 mV). (See "Cardiac excitability, mechanisms of arrhythmia, and action of antiarrhythmic drugs".)

MUSCLE CONTRACTION — The contractile apparatus of myocytes is the myofibril, a cylindrical structure composed of thick and thin filaments arranged longitudinally parallel within the myofibril.

●The thin filament is composed of two strands of F-actin wound around each other. Each strand of F-actin is composed of globular G-actin monomers, polymerized in a linear pattern.

●The thick filament is composed of myosin, a molecule with a long "tail" section made up of two helices twisted around one another, each attached to a roughly globular "head." There is a hinge point between the tail and the head that allows the head segment of the molecule to fold back upon itself. The thick filament, then, is an association of all the tail portions of many myosin molecules with the head portions protruding toward the adjacent thin filaments.

●The sarcomere is the functional unit of the myofibril. Each sarcomere is approximately 2 micrometers in length, and there are many copies throughout the length of the myofibril. The sarcomere is bounded at each end by the Z-bands, which run perpendicular to the thick and thin filaments and to which the thin filaments are attached. In the relaxed state, the thin filaments extend from the Z-band toward the center of the sarcomere (figure 2). The gap in the center between the thin filaments is the H-zone. The thick filaments are interspersed between the thin filaments (parallel to them) in the center of the sarcomere, crossing the H-zone. During contraction, the thick filaments "pull" the thin filaments and the Z-bands toward the center of the sarcomere. Both the H-zone and the sarcomere itself shorten in length during contraction.

Mechanism — Muscular contraction is the shortening of the myofibrils that occurs when the thick and thin filaments slide past one another (figure 2). The sliding of the thick and thin filaments is thought to occur by repetitive binding of the myosin heads to actin, flexion of the myosin molecules at their hinge points, release of the binding between the molecules, and relaxation of the myosin molecule prior to binding the actin again. This repetitive motion is referred to as cross-bridge cycling. In this way, the thick filament is thought to "ratchet" itself past the thin filament.

The mechanism of cross-bridge cycling is complex, but may be usefully modeled by a four-step process requiring the presence of adenosine triphosphate (ATP) (figure 3) [1]. Beginning with actin and myosin in the dissociated state, the myosin head is bound to an ATP molecule.

●The first step is the hydrolysis of the high energy phosphate bond of ATP to form adenosine diphosphate (ADP) and Pi (inorganic phosphate), thereby energizing the myosin molecule.

●The second step is formation of the active actin-myosin complex (the cross-bridge).

●The third step is dissociation of ADP and Pi from the myosin head, which allows the flexion of the myosin at its hinge point. The myosin head remains bound and flexed until the fourth step.

●The fourth step is binding of ATP, which allows dissociation of the actin and myosin returning to the beginning of the cycle.

Thus, while the hydrolysis of ATP supplies the energy for the flexion of the myosin molecule, it is the binding of ATP that allows the dissociation of the myosin head from actin. (This is also true in skeletal muscle, accounting for the phenomenon of rigor mortis, the stiffening of skeletal muscle after death, due to lack of ATP to bind the myosin in the flexed state.)

Role of tropomyosin and troponins — In the resting state, the interaction of the thick and thin filaments is physically blocked by the presence of the protein tropomyosin, which lies in the groove of the thin filament formed by the two F-actin strands. Tropomyosin is closely associated with three other proteins, troponin I, troponin T, and troponin C. It is the troponin-tropomyosin complex that regulates the interaction of the thick and thin filaments and therefore muscular contraction. Troponin C has a calcium binding site that, when occupied by a calcium ion, causes a conformational change in the troponin-tropomyosin complex (figure 4). This change moves the tropomyosin molecule from its resting position, which makes the myosin binding site accessible to the myosin head. Cross-bridge cycling then begins and the myofibrils forcefully shorten until calcium is removed from troponin C and tropomyosin returns to its resting position, blocking actin and myosin binding.

The other troponin proteins also contribute to this process. Troponin I is an inhibitory protein that inhibits the ATPase of actomyosin and also modulates calcium binding to troponin C. Troponin T serves to attach the troponin complex to actin and tropomyosin; in the relaxed state, it blocks the actin-myosin binding site.

SIGNAL TRANSDUCTION — Excitation-contraction (E-C) coupling can be viewed as the transduction of a cell membrane signal (the action potential) to stimulate the action of an effector in the cell (myofibril contraction). The second messenger in this transduction system is calcium. High concentrations of calcium within the cell are very cytotoxic, stimulating proteases and lipases whose actions ultimately result in cell death. As a result, nearly all cells are highly adapted to maintain very low levels of intracellular free calcium.

The low cell calcium concentration has been put to use by several cell types, allowing rapid influx of calcium to serve as a trigger for the action of an intracellular effector [1]. The calcium transient must be fleeting in order to ensure the long-term health of the cell. In mammalian myocardial cells, for example, the resting intracellular calcium concentration is in the range of 100 to 300 nmol/L. With each action potential, intracellular free calcium rises rapidly by a factor of approximately 10,000 to the millimolar range. At this concentration, calcium binds to troponin C, allowing actin-myosin cross-bridge formation and myofibril shortening. The rapid removal of free calcium from the cytoplasm is also critical, not only to relaxation of the myocardium, but to the health of the cell.

The calcium transient — Thus, the generation, modulation, and termination of the intracellular calcium transient is the essence of E-C coupling. The calcium transient is initiated by the entry of calcium into the cell via voltage-activated calcium-specific membrane channels. These are among the channels that are opened transiently during the action potential. The favorable and large electrochemical gradient (low cell calcium concentration, cell interior electronegative) provides a strong driving force for the influx of calcium.

However, calcium influx through these sarcolemmal channels is not the major source of calcium producing the calcium transient. Instead, a relatively small influx of calcium triggers the release of a much larger quantity of calcium from the primary intracellular store of calcium, the sarcoplasmic reticulum (SR), through SR calcium release channels.

The sarcolemmal calcium channel and the SR calcium release channel are different entities. While the calcium channels of the sarcolemma (known as L-type calcium channels) are activated by voltage and blocked by the dihydropyridines (such as nifedipine), the SR calcium channels are stimulated to open by the binding of calcium itself on the cytoplasmic side of the membrane.

The rise in intracellular free calcium concentration is a very short-lived phenomenon, lasting approximately 100 msec. The excess calcium must be removed from the cytoplasm in order to stop the process of actin-myosin cross-bridge cycling and to permit the myocardium to relax. In the steady state, the same amount of calcium that entered the cell through the sarcolemma from the extracellular space must be extruded from the cell. Similarly, the amount of calcium that was released from SR must be re-sequestered in the SR. In both cases the transport of calcium is against the electrochemical gradient, requiring active, energy dependent transport processes, and ultimately adenosine triphosphate (ATP).

Calcium efflux to the extracellular space is mediated by two different transporters:

●The sarcolemmal calcium pump, which is a membrane protein that transports its substrate (calcium) against the electrochemical gradient using the energy obtained by hydrolysis of a high energy phosphate bond of ATP.

●The Na⁺/Ca²⁺ exchanger, a secondary active transporter that uses the inward energy in the electrochemical sodium gradient to drive the extrusion of calcium against its gradient. The sodium gradient is established by the action of the Na-K-ATPase pump, which maintains a low cell sodium concentration and a cell interior negative potential.

The Na⁺/Ca²⁺ exchange mechanism has a high capacity for calcium, but a low affinity. It is therefore most efficient at calcium efflux when the intracellular calcium is high. Conversely, the calcium pump has a high affinity for calcium but a low capacity. Its primary role is to remove the remaining calcium above the resting concentration after intracellular calcium has decreased from its peak, when the Na⁺/Ca²⁺ exchanger is working less efficiently.

Uptake of calcium into the SR is mediated by a calcium pump protein located in the SR membrane. The SR calcium pump is another member of the family of transporters that derives its energy from hydrolysis of ATP. Calcium is sequestered in the SR at high concentrations. Much of this calcium is bound to proteins, such as calsequestrin 2, which concentrate the calcium in the terminal cisternae of the SR. From this site, calcium is readily available for release into the cytoplasm during the next contraction cycle.

MODULATION OF MYOCARDIAL FUNCTION — In view of the central role of the calcium transient, modulation of either the calcium transient or the response of the contractile apparatus to the calcium transient will also affect contractility. Such modulation may occur as a response to a physiologic need for increased cardiac output. On the other hand, failure of proper control of excitation-contraction (E-C) coupling may be the etiology of pathologic states, a setting in which pharmacologic modification of E-C coupling may be an important component to therapy.

This section will discuss some of the ways in which modulation of E-C coupling can change myocardial function. Two figures are representations of the regulation of sarcolemma and sarcoplasmic reticulum function (figure 5 and figure 6).

Alteration of the calcium transient — The magnitude of the calcium transient may be increased in several ways. The simplest is making more calcium available to the cell from the extracellular space. It must be remembered that the extracellular calcium is not the primary source of the calcium transient. However, increased extracellular calcium raises the transsarcolemmal gradient, increasing the influx of calcium in each cycle of calcium channel opening. This change in intracellular calcium gradually increases the sarcoplasmic reticulum (SR) calcium store, and therefore increases the magnitude of the calcium transient until a new steady state between sarcolemmal influx and efflux is established. Increasing extracellular calcium is effective in vitro, but has little application in vivo. The sophisticated systems for regulating plasma free calcium make it difficult to significantly change the transsarcolemmal calcium gradient for an extended period of time.

Another way to increase cell calcium is to inhibit efflux by increasing the cell sodium concentration, thereby diminishing the activity of Na⁺/Ca²⁺ exchange. This is the mechanism by which digitalis, via inhibition of the Na-K-ATPase pump, affects contractility (see below) (figure 5).

The amount of calcium available for SR sequestration can also be increased by raising cycle frequency. The duration of contraction (systole) is much less flexible than the duration of relaxation (diastole). (It is a well-known phenomenon that, as the heart rate increases, diastole shortens much more than systole.) Thus, calcium efflux for each cycle cannot match the calcium influx as the rate increases. The amount of calcium in the cell increases until a new steady state is reached. The gradually increasing force of contractility with an increase in contraction frequency is known as the treppe or Bowditch positive staircase.

Extrasystoles have a similar effect. While the force of a premature systole may not be great, there is an "extra" opportunity for calcium to enter the cell. The contraction following an extrasystole is significantly more forceful than a contraction of typical cycle (a phenomenon called postextrasystolic potentiation). The extra action potential allows another opportunity for calcium influx via the voltage activated calcium channels (and also decreases calcium efflux via the Na⁺/Ca²⁺ exchanger). More intracellular calcium is now available to bind to troponin C and activate the myofibrils.

Digitalis — Cardiac glycosides (such as digoxin) are thought to act by indirectly increasing the availability of intracellular calcium [2]. The proposed mechanism begins with the primary action of these drugs, inhibition of the Na-K-ATPase pump. This transporter is the primary route of sodium efflux from the cell. Inhibition of the pump sequentially increases intracellular sodium, diminishes the activity of the Na⁺/Ca²⁺ exchanger (since there is a less favorable inward gradient for sodium), and, because of a lower rate of extrusion, enhances the amount of intracellular calcium available for sequestration and use in E-C coupling (figure 5).

Cyclic adenosine monophosphate — Cyclic adenosine monophosphate (AMP) is the most important modulator of intracellular calcium handling, making it the most important second messenger in E-C coupling after calcium itself. Cyclic AMP is formed from adenosine triphosphate (ATP) by the enzyme adenylyl cyclase. The activity of adenylyl cyclase is stimulated by the beta-adrenergic receptor (via a G-protein) when occupied by beta agonist (catecholamine) (figure 5). Cyclic AMP is metabolized in the cell by phosphodiesterases, inhibition of which can increase cardiac contractility. (See "Inotropic agents in heart failure with reduced ejection fraction".)

Cyclic AMP activates cyclic AMP-dependent protein kinases, which catalyze the phosphorylation of their specific target proteins and modify the function of these proteins. The effects of increasing cyclic AMP illustrate the importance of the interaction between inotropy and lusitropy in increasing stroke volume, which is the ultimate function of beta adrenergic stimulation of the heart [3].

●Cyclic AMP mediates the phosphorylation of the sarcolemmal calcium channel, which results in an increase in the conductance of the channel. As a result, more calcium can enter the cell and strengthen the force of contraction.

●Cyclic AMP stimulates protein kinase A, which phosphorylates the protein phospholamban, which, when not phosphorylated, inhibits the SR calcium pump [4]. Phosphorylation removes this inhibitory activity; this is an inotropic effect since it enhances intracellular calcium stores and increases the magnitude of the calcium transient. Experimental studies and preliminary human data suggest that gene therapy aimed at reducing phospholamban activity can reduce heart failure progression in association with increased SR calcium pump activity. (See "Investigational therapies for management of heart failure", section on 'Stem cell therapy'.)

Increasing SR calcium pump activity also has a lusitropic effect since the increased rate of sequestration of calcium into the SR shortens the duration of the calcium transient. This has the effect of removing calcium from its troponin C binding site and halting actin myosin cross-bridge cycling more quickly. Systole is shortened, leaving time in the cycle for the heart to relax and fill with blood in preparation for the next systole. Increased filling increases stroke volume and enhances the force of contraction via the Frank-Starling mechanism.

●Another lusitropic effect of cyclic AMP is achieved by phosphorylation of the Na-K-ATPase pump, which increases its turnover rate, thereby enhancing sodium efflux. The ensuing fall in cell sodium concentration increases the ability of the Na⁺/Ca²⁺ exchanger to extrude calcium from the cell. Once again, the effect is to shorten the duration and decrease the magnitude of the calcium transient and hasten myocardial relaxation.

Phosphatidylinositol pathway — Another second messenger pathway is involved in calcium handling. Inositol 4,5-triphosphate (IP3) and diacylglycerol (DAG) are the products of hydrolysis of phosphatidylinositol 4,5-biphosphate (PIP2) (figure 7). Hydrolysis of PIP2 is catalyzed by the enzyme phospholipase C. Phospholipase C activity is stimulated (via a G-protein) by the binding of an alpha-adrenergic agonist to the alpha adrenergic receptor. IP3 stimulates the calcium release channel of the SR, increasing the magnitude of the calcium transient, which in turn increases contractile force [5].

Angiotensin II — There are specific receptors for angiotensin II in myocardium. Angiotensin II is a potent vasoconstrictor that also acts directly on the heart. In addition, there is strong evidence for local secretion of renin and angiotensinogen by the heart itself (ie, for the local formation of angiotensin II) [6,7]. Angiotensin II affects heart rate and contractility and, over the long term, growth [6,8]. It has been proposed that angiotensin II is an important determinant of the development of left ventricular hypertrophy in patients with hypertension [9].

Most of the data suggest that angiotensin II enhances contractility by a cyclic AMP-mediated mechanism. However, not all studies have shown an inotropic effect of angiotensin II. An in vitro study of human myocardium, for example, found that angiotensin II increased contractility of atrial but not ventricular myocardium [10].

Alteration of the myofibril response to calcium — Modification of contractility without changing the calcium transient requires modification of the myofibril response to free calcium. The affinity of the troponin C binding site or the response of the myofibrils to a given level of calcium binding may be changed.

Cyclic AMP also plays a role in this type of modification of E-C coupling. Cyclic AMP mediates the phosphorylation of troponin I. This decreases the affinity of troponin C for calcium, which is a lusitropic effect. The lower affinity for calcium permits faster sequestration in the SR. Once again, diastolic relaxation and ventricular filling, stimulated by beta 1 and beta 2- adrenergic activity and mediated by cyclic AMP, may be seen as a means of increasing stroke volume and cardiac output [11]. Though the reduction in affinity for calcium has a negative inotropic effect, this is overcome in systole by the increase in calcium available. Cyclic AMP also has the positively inotropic effect of enhancing the rate of actin-myosin cross-bridge cycling [12].

The alpha adrenergic/PIP2 pathway also appears to affect the calcium affinity of the contractile apparatus. IP3 and DAG increase the binding of calcium to its receptor, a positive inotropic effect. This appears to be mediated by phosphorylation by protein kinase C, although the target for phosphorylation is not yet known [13].

Intracellular pH — It has long been known that acidosis has a negative inotropic influence on the heart [14,15]. An acid environment affects each stage of E-C coupling, including the action potential and the Ca²⁺ transient. An important site of action appears to be on the sensitivity of the contractile apparatus to calcium. Acidosis markedly decreases calcium binding to troponin C by increasing the inhibitory effect of troponin I and to a lesser degree by a direct effect on the affinity of troponin C for calcium [16].

Heart failure — Abnormal regulation of intracellular calcium plays a prominent role in heart failure. In an animal model of myocardial infarction, the calcium current influx is less able to trigger the release of calcium from the SR, an important factor contributing to the development of contractile dysfunction and heart failure [17].

Monitoring calcium transients in vitro has shown that patients with cardiomyopathies have abnormal regulation of intracellular calcium [18]. The reduced intracellular calcium transient is likely due to a decreased SR calcium uptake and content [19]. Impaired regulation of the calcium transient and of the maintenance of a low resting intracellular calcium can cause both poor contractile function in systole and poor relaxation in diastole [20,21]. It has been estimated, for example, that up to 40 percent of patients with heart failure have normal systolic function, suggesting diastolic dysfunction as the cause of failure [22]. (See "Heart failure with preserved ejection fraction: Clinical manifestations and diagnosis".)

One possible cause of abnormal calcium regulation in congestive heart failure is impaired production of intracellular cyclic AMP. This is suggested by the relatively poor response of diseased myocardium to inotropic drugs which, in normal myocardium, increase cyclic AMP levels by inhibiting its breakdown by phosphodiesterase. Furthermore, pharmacologic stimulation of cyclic AMP production via stimulation of adenylyl cyclase with forskolin can potentiate the effects of phosphodiesterase inhibitors [23,24].

Since intracellular calcium appears so important in both systolic and diastolic dysfunction, drugs that modify intracellular calcium are potential candidates for heart failure therapy. This is an area of active investigation, although the results at the clinical level have been somewhat disappointing. Inotropic agents may actually increase or, with digoxin, have no effect on mortality [25], while drugs that decrease cardiac work, particularly the angiotensin converting enzyme inhibitors, prolong survival by slowing the rate of deterioration of myocardial function. (See "Inotropic agents in heart failure with reduced ejection fraction" and "Overview of the management of heart failure with reduced ejection fraction in adults".)

Inotropic drugs that modulate calcium handling may be reasonably divided into two groups [20]:

●Those that act by increasing cyclic AMP, including the beta adrenergic agonists (such as isoproterenol, norepinephrine, epinephrine, dopamine, dobutamine), the phosphodiesterase inhibitors such as inamrinone and milrinone.

●Those that act more directly on calcium movements into and out of the sarcoplasm, such as digitalis. On the other hand, calcium channel blockers, such as nifedipine (and other dihydropyridines) and verapamil have negative inotropic effects.

Another potential mechanism for an inotropic agent is increasing the sensitivity of the myofibril response to the calcium transient, thereby increasing contractility at a given concentration of calcium. Pimobendan is an agent that sensitizes the contractile proteins to calcium and has been shown to have an inotropic effect as well as a positive clinical effect on patients with heart failure [26]. (See "Inotropic agents in heart failure with reduced ejection fraction".)

Familial hypertrophic cardiomyopathy — Studies in patients with familial forms of hypertrophic cardiomyopathy illustrate the importance of proper function of the different steps in E-C coupling. Defects in several different contractile protein genes have been identified in these patients. (See "Hypertrophic cardiomyopathy: Morphologic variants and the pathophysiology of left ventricular outflow tract obstruction" and "Hypertrophic cardiomyopathy: Gene mutations and clinical genetic testing".)

Aging — Advancing age is associated with alterations in the various steps of E-C coupling [27].

SUMMARY

●Linear propagation of the action potential occurs rapidly because the myocytes along muscle fibers are electrically continuous. An action potential can also cause the depolarization of a neighboring fiber, and the electrical impulse occurs as a "wave" through the heart. (See 'Myocardial action potential' above.)

●The contractile apparatus of myocytes is the myofibril, a cylindrical structure composed of thick and thin filaments arranged longitudinally parallel within the myofibril. (See 'Muscle contraction' above.)

●Muscular contraction is the shortening of the myofibrils that occurs when the thick and thin filaments slide past one another (figure 2). The sliding of the thick and thin filaments is thought to occur by repetitive binding of the myosin heads to actin, flexion of the myosin molecules at their hinge points, release of the binding between the molecules, and relaxation of the myosin molecule prior to binding the actin again. This repetitive motion is referred to as cross-bridge cycling. (See 'Mechanism' above.)

●In the resting state, the interaction of the thick and thin filaments is physically blocked by the presence of the protein tropomyosin, which lies in the groove of the thin filament formed by the two F-actin strands. (See 'Role of tropomyosin and troponins' above.)

●Excitation-contraction (E-C) coupling can be viewed as the transduction of a cell membrane signal (the action potential) to stimulate the action of an effector in the cell (myofibril contraction). The second messenger in this transduction system is calcium. (See 'Signal transduction' above.)

●The generation, modulation, and termination of the intracellular calcium transient is the essence of E-C coupling. (See 'The calcium transient' above.)

●Cyclic adenosine monophosphate (AMP) is the most important modulator of intracellular calcium handling, making it the most important second messenger in E-C coupling after calcium itself. Cyclic AMP is formed from adenosine triphosphate by the enzyme adenylyl cyclase. (See 'Cyclic adenosine monophosphate' above.)

●Modification of contractility without changing the calcium transient requires modification of the myofibril response to free calcium. (See 'Alteration of the myofibril response to calcium' above.)

●Abnormal regulation of intracellular calcium plays a prominent role in heart failure. (See 'Heart failure' above.)

●Since intracellular calcium appears so important in both systolic and diastolic dysfunction, drugs that modify intracellular calcium are potential candidates for heart failure therapy. (See 'Heart failure' above.)