**2. Acute antiarrhythmic mechanisms of melatonin**

Melatonin acts at multiple electrophysiological levels due to its receptordependent and receptor-independent mechanisms. In 1998, the seminal work of Tan et al. highlighted the antiarrhythmic properties of melatonin [159]. During the past two decades, our understanding of the pleiotropic action of melatonin increased significantly.

The antiarrhythmic effect of melatonin was first attributed to its notable antioxidant properties, mainly because melatonin results were better than those obtained with an ascorbic acid at concentration 10–500 times higher [159]. Numerous studies confirmed antiarrhythmic protection and related it to its remarkable antioxidant properties [160–167].

Our research group corroborated the antiarrhythmic effect of melatonin in isolated hearts of female rats, when administered continuously from the stage prior to the onset of myocardial ischemia [127]. Notably, the antiarrhythmic protection had a dose-dependent response, while the antioxidant capacity was the same for all the doses studied. The preventive effect on the shortening of the action potential that occurs between the 7th and 10th minute of the ischemia was another dose-dependent variable found in our study. This led us to think that the antiarrhythmic mechanism could be due to a lower heterogeneity in the repolarization of myocardial tissue that diminishes the possibility of reentry circuits being formed and maintained. As previously mentioned, the time- and dose-dependent responses could be due to melatonin inhibitory effect against swell-activated anionic currents [126–128].

We recently showed that melatonin reduces arrhythmias when administered during reperfusion, a useful timing for the clinical context of acute coronary syndromes, because most therapies can only start close to the reperfusion period [168]. Melatonin showed protective mechanisms when administered to isolated hearts of rats fed with fructose and spontaneously hypertensive rats. These animals show greater activity of the enzyme NADPH oxidase, which is one of the main systems for generating free radicals, and, therefore, higher levels of oxidative stress. The antiarrhythmic effect was not affected in the models with greater oxidative stress, and in all groups, it was accompanied by a temporary shortening of the duration of the action potential during the first 3–5 minutes of reperfusion. This result was interpreted as a reduction in the ability to generate early and late postdepolarizations. Self-limited arrhythmic events, such as ventricular extrasystoles, salvos, and even non-sustained ventricular tachycardia, occurred in all experimental groups. The main difference was that the hearts treated with melatonin did not show sustained forms of arrhythmias, either sustained ventricular tachycardia or ventricular fibrillation. These results (potential shortening and absence of sustained arrhythmia) are difficult to reconcile with the mechanisms postulated for reentry circuits.

The same year of our publication of the antiarrhythmic protection of melatonin administered in reperfusion, another group published that melatonin protects against arrhythmias, by increasing the threshold to electrically induce sustained ventricular fibrillation, by increasing the myocardial Cx43 by PKC in hypertensive rats [169]. Melatonin prevented myocardial abnormalities of connexin and improved cardiac conduction.

Based in these interesting results, we tested if melatonin could prevent hypokalemia-induced ventricular fibrillation by Cx43 preservation [83]. The acute administration of melatonin during low potassium perfusion reduced the incidence of ventricular fibrillation and improved the recovery of sinus rhythm in those hearts that, despite being treated with melatonin, developed sustained fibrillation. Protection was mediated by the activation of melatonin receptors and by the prevention of dephosphorylation and lateralization of Cx43.

**57**

*Melatonin for a Healthy Heart Rhythm*

*DOI: http://dx.doi.org/10.5772/intechopen.91447*

preservation of membrane potential.

related to Cx43 lateralization and dephosphorylation.

A brief explanation of the electrophysiological changes induced by hypokalemia will help to appreciate the relevance as antiarrhythmic. Severe hypokalemia induces changes in ventricular repolarization, such as lengthening the QT interval, prominent U waves, fusion of T and U waves associated with and increases risk of arrhythmic death [83, 170, 171]. Our experimental model confirmed the lengthening of the QT interval and correlated with an increase in the duration of the action potential [83]. Melatonin did not prevent the prolongation of the action potential induced by hypokalemia when measured at 90% of repolarization but maintained action potential duration at 50% of repolarization and made the membrane potential more stable, showing less after depolarization. Luzindole blunted both effects of melatonin, suggesting the involvement of melatonin receptor activation in the

Hypokalemia decreases NaK-ATPase activity and causes an intracellular Ca2+ overload that facilitates the development of delayed postdepolarizations through the transient inward currents [172–174]. Delayed postdepolarizations are considered triggers of arrhythmias because they can initiate an action potential in isolated cells. However, it is unlikely that an extra action potential can be initiated from a single cell in the tissue due to a mismatch between the current source from the cell and the current sink produced by the surrounding cells [175]. To overcome the source-sink mismatch, there must be a reduced sink through intercellular decoupling or an increase in the source through the synchronization of delayed postdepolarizations between several adjacent cells. Both situations could be assumed based on the results of anisotropic conduction studies and immunofluorescence imaging [83]. In fact, hypokalemia induces conduction abnormalities, increased amplitude and duration of the P wave, a slight prolongation of the PR interval, atrioventricular block, increased QRS duration, and cardiac arrest [173, 176]. We found all these electrocardiographic disorders during our experimental model of hypokalemia [83]. Melatonin prevented the widening of the QRS and delayed activation of the potential for epicardial action. The latter could be considered as a substitute for conduction velocity in complex tissues such as ventricles, assuming unknown routes from endocardial activation points that indicate the onset of QRS to epicardial myocytes recorded with microelectrodes. These improvements in ventricular conduction were

The lateralization of connexins has been detected in chronic atrial fibrillation, cardiac hypertrophy, heart failure, and after myocardial infarction [21, 177–179]. An increase in the fraction of lateral connexins that form functional channels improves transverse conduction velocity and contributes to the spread of the arrhythmogenic impulse. High side-by-side lateralization can favor conduction blockage due to mismatches between the source and the sink [175, 180]. A unidirectional block can lead to reentry circles that result in tachycardia or ventricular fibrillation [181]. Therefore, the acute lateralization induced by hypokalemia is an important arrhythmogenic factor [83]. It is noteworthy that melatonin prevented acute lateralization of Cx43. Connexin 43 phosphorylation could lead to better coupling or uncoupling depending on the target amino acid, but dephosphorylation is clearly associated with uncoupling [21, 177, 182]. It is not yet known whether the dephosphorylation of Cx43 during low potassium is the result of increased phosphatase activity and/or an increase in phosphokinase or what are the intracellular mechanisms that prevented dephosphorylation when treated with melatonin. Dramatic reductions in intercellular communication due to the loss of phosphorylated Cx43 and the accumulation of nonphosphorylated Cx43 were previously reported in other experimental models [177]. Our results could be relevant mainly in those situations in which acute hypokalemia can be anticipated as in dialysis [183, 184]. Both QT interval and the QT dispersion increase after dialysis. We propose that melatonin could make the heart

#### *Melatonin for a Healthy Heart Rhythm DOI: http://dx.doi.org/10.5772/intechopen.91447*

*Melatonin - The Hormone of Darkness and Its Therapeutic Potential and Perspectives*

Melatonin acts at multiple electrophysiological levels due to its receptordependent and receptor-independent mechanisms. In 1998, the seminal work of Tan et al. highlighted the antiarrhythmic properties of melatonin [159]. During the past two decades, our understanding of the pleiotropic action of melatonin

The antiarrhythmic effect of melatonin was first attributed to its notable antioxidant properties, mainly because melatonin results were better than those obtained with an ascorbic acid at concentration 10–500 times higher [159]. Numerous studies confirmed antiarrhythmic protection and related it to its remarkable antioxidant

Our research group corroborated the antiarrhythmic effect of melatonin in isolated hearts of female rats, when administered continuously from the stage prior to the onset of myocardial ischemia [127]. Notably, the antiarrhythmic protection had a dose-dependent response, while the antioxidant capacity was the same for all the doses studied. The preventive effect on the shortening of the action potential that occurs between the 7th and 10th minute of the ischemia was another dose-dependent variable found in our study. This led us to think that the antiarrhythmic mechanism could be due to a lower heterogeneity in the repolarization of myocardial tissue that diminishes the possibility of reentry circuits being formed and maintained. As previously mentioned, the time- and dose-dependent responses could be due to melatonin

We recently showed that melatonin reduces arrhythmias when administered during reperfusion, a useful timing for the clinical context of acute coronary syndromes, because most therapies can only start close to the reperfusion period [168]. Melatonin showed protective mechanisms when administered to isolated hearts of rats fed with fructose and spontaneously hypertensive rats. These animals show greater activity of the enzyme NADPH oxidase, which is one of the main systems for generating free radicals, and, therefore, higher levels of oxidative stress. The antiarrhythmic effect was not affected in the models with greater oxidative stress, and in all groups, it was accompanied by a temporary shortening of the duration of the action potential during the first 3–5 minutes of reperfusion. This result was interpreted as a reduction in the ability to generate early and late postdepolarizations. Self-limited arrhythmic events, such as ventricular extrasystoles, salvos, and even non-sustained ventricular tachycardia, occurred in all experimental groups. The main difference was that the hearts treated with melatonin did not show sustained forms of arrhythmias, either sustained ventricular tachycardia or ventricular fibrillation. These results (potential shortening and absence of sustained arrhythmia) are difficult to reconcile with the mechanisms postulated for reentry circuits. The same year of our publication of the antiarrhythmic protection of melatonin

administered in reperfusion, another group published that melatonin protects against arrhythmias, by increasing the threshold to electrically induce sustained ventricular fibrillation, by increasing the myocardial Cx43 by PKC in hypertensive rats [169]. Melatonin prevented myocardial abnormalities of connexin and

Based in these interesting results, we tested if melatonin could prevent hypokalemia-induced ventricular fibrillation by Cx43 preservation [83]. The acute administration of melatonin during low potassium perfusion reduced the incidence of ventricular fibrillation and improved the recovery of sinus rhythm in those hearts that, despite being treated with melatonin, developed sustained fibrillation. Protection was mediated by the activation of melatonin receptors and by the

prevention of dephosphorylation and lateralization of Cx43.

inhibitory effect against swell-activated anionic currents [126–128].

**2. Acute antiarrhythmic mechanisms of melatonin**

increased significantly.

properties [160–167].

**56**

improved cardiac conduction.

A brief explanation of the electrophysiological changes induced by hypokalemia will help to appreciate the relevance as antiarrhythmic. Severe hypokalemia induces changes in ventricular repolarization, such as lengthening the QT interval, prominent U waves, fusion of T and U waves associated with and increases risk of arrhythmic death [83, 170, 171]. Our experimental model confirmed the lengthening of the QT interval and correlated with an increase in the duration of the action potential [83]. Melatonin did not prevent the prolongation of the action potential induced by hypokalemia when measured at 90% of repolarization but maintained action potential duration at 50% of repolarization and made the membrane potential more stable, showing less after depolarization. Luzindole blunted both effects of melatonin, suggesting the involvement of melatonin receptor activation in the preservation of membrane potential.

Hypokalemia decreases NaK-ATPase activity and causes an intracellular Ca2+ overload that facilitates the development of delayed postdepolarizations through the transient inward currents [172–174]. Delayed postdepolarizations are considered triggers of arrhythmias because they can initiate an action potential in isolated cells. However, it is unlikely that an extra action potential can be initiated from a single cell in the tissue due to a mismatch between the current source from the cell and the current sink produced by the surrounding cells [175]. To overcome the source-sink mismatch, there must be a reduced sink through intercellular decoupling or an increase in the source through the synchronization of delayed postdepolarizations between several adjacent cells. Both situations could be assumed based on the results of anisotropic conduction studies and immunofluorescence imaging [83].

In fact, hypokalemia induces conduction abnormalities, increased amplitude and duration of the P wave, a slight prolongation of the PR interval, atrioventricular block, increased QRS duration, and cardiac arrest [173, 176]. We found all these electrocardiographic disorders during our experimental model of hypokalemia [83]. Melatonin prevented the widening of the QRS and delayed activation of the potential for epicardial action. The latter could be considered as a substitute for conduction velocity in complex tissues such as ventricles, assuming unknown routes from endocardial activation points that indicate the onset of QRS to epicardial myocytes recorded with microelectrodes. These improvements in ventricular conduction were related to Cx43 lateralization and dephosphorylation.

The lateralization of connexins has been detected in chronic atrial fibrillation, cardiac hypertrophy, heart failure, and after myocardial infarction [21, 177–179]. An increase in the fraction of lateral connexins that form functional channels improves transverse conduction velocity and contributes to the spread of the arrhythmogenic impulse. High side-by-side lateralization can favor conduction blockage due to mismatches between the source and the sink [175, 180]. A unidirectional block can lead to reentry circles that result in tachycardia or ventricular fibrillation [181]. Therefore, the acute lateralization induced by hypokalemia is an important arrhythmogenic factor [83]. It is noteworthy that melatonin prevented acute lateralization of Cx43.

Connexin 43 phosphorylation could lead to better coupling or uncoupling depending on the target amino acid, but dephosphorylation is clearly associated with uncoupling [21, 177, 182]. It is not yet known whether the dephosphorylation of Cx43 during low potassium is the result of increased phosphatase activity and/or an increase in phosphokinase or what are the intracellular mechanisms that prevented dephosphorylation when treated with melatonin. Dramatic reductions in intercellular communication due to the loss of phosphorylated Cx43 and the accumulation of nonphosphorylated Cx43 were previously reported in other experimental models [177].

Our results could be relevant mainly in those situations in which acute hypokalemia can be anticipated as in dialysis [183, 184]. Both QT interval and the QT dispersion increase after dialysis. We propose that melatonin could make the heart

#### **Figure 1.**

*Acute antiarrhythmic mechanisms of melatonin. The red arrows indicate stimulation, and the interrupted blue lines indicate blockage.*

more resistant to arrhythmic events triggered by rapid changes in plasma electrolyte concentrations, regardless of a lack of effects on the ECG. In addition, those dialysis patients also suffer from disorders in the circadian rhythms and low levels of melatonin [185]. However, clinical translations of our results should be done with caution, mainly because we use a high dose of melatonin administered directly to the heart. Based on melatonin's pharmacokinetics in humans, to achieve a similar concentration in plasma to the one tested *ex vivo*, a dose 10 times higher the highest intravenous dose tested until now should be administered [186, 187].

Melatonin has a remarkable antiarrhythmic activity that is carried out based on actions dependent on and independent of receptor activation. To summarize we propose that the antiarrhythmic effect of melatonin is mediated by receptor activation beyond its outstanding antioxidant actions (**Figure 1**). The shortening of the action potential could be associated with the activation of MT1 melatonin receptors, since they can regulate specific ion channels such as Kir3.1 channel. MT1 and MT2 receptors could indirectly modulate other electrophysiological effects through intracellular signaling such as decreased cyclic adenosine monophosphate, increased phospholipase C, and PKC activation.

## **3. Chronic antiarrhythmic mechanisms of melatonin**

Endogenous melatonin would be an intrinsically protective factor with therapeutic potential [188, 189]. Melatonin is a promising treatment for cardiovascular diseases such as myocardial ischemia/reperfusion injury, hypertension, and heart failure. It has been shown that melatonin levels were reduced in patients with acute myocardial infarction and in patients undergoing primary coronary angioplasty [190]. These findings suggest that melatonin could play an important role in preventing ischemia/reperfusion heart injury. Indeed, reperfusion arrhythmias increase in pinealectomized animals, suggesting a protective role of endogenous physiological melatonin levels [163, 189].

Chronic melatonin supplementation, either in physiological or pharmacological ranges, protects against arrhythmias [8, 21, 97, 163, 169, 189, 191, 192]. Beyond the reported antioxidant properties of melatonin, it reduces severe ventricular

**59**

**Figure 2.**

*marks direct mitochondrial protection.*

*Melatonin for a Healthy Heart Rhythm*

*DOI: http://dx.doi.org/10.5772/intechopen.91447*

arrhythmias by antifibrotic mechanisms, electrical remodeling, direct mitochondrial protection, myocardial Cx43 preservation via PKC signaling, and vitamin D-HSP70/AT1 counterbalance (**Figure 2**). Its cardioprotective properties persist in relevant cardiovascular risk factor models like hypertensive, obese, and nephropathic rats. The latter is interesting because most of the therapeutic interventions

A preventive approach would be of great value in the face of unpredictable acute arrhythmic events, especially if the intervention manages to avoid the most severe and potentially lethal arrhythmias such as ventricular tachycardia and fibrillation. Numerous efforts have been made in that direction. In the last quarter of the twentieth century, several antiarrhythmic drugs were tried, but most of them showed a proarrhythmogenic profile or failed to reduce mortality [193–196]. A time of great progress was appreciated with the introduction of implantable cardiodefibrillators. However, surgical intervention and high cost limit its population efficiency. A strategy to improve the availability of preventive interventions is to select potential beneficiaries based on their risk of serious events. This would compensate for potential side effects and optimize the investment of resources. Other strategies, such as vaccines, are based on achieving the greatest possible scope with the least number of interventions that attenuate the severity of diseases. In the case of arrhythmias, we still have no clear "antiarrhythmic vaccine." Therefore, risk-oriented strategies would be an acceptable approach. From a preventive point of view, the pleiotropic protection mechanisms of melatonin could effectively limit the arrhythmic complications associated with hypertension [21, 169]. Arterial hypertension causes vascular deterioration, overloads the heart, and predisposes to a greater number of arrhythmic events. More than five decades ago, it was reported that surgical removal of the pineal gland, a procedure that essentially eliminates circulating levels of melatonin, was followed by a slow but persistent increase in blood pressure in rats [197]. This finding has been confirmed in several subsequent studies [11]. In addition, daily treatment of pinealectomized rats with melatonin attenuated the elevation in blood pressure

*Antiarrhythmic mechanisms of chronic melatonin administration. Extracellular lines represent reduced fibrosis after melatonin treatment. The red arrows indicate stimulation and blue ones show blockage. The green arrow* 

postulated so far fail to be reproduced under risk factor conditions.

### *Melatonin for a Healthy Heart Rhythm DOI: http://dx.doi.org/10.5772/intechopen.91447*

*Melatonin - The Hormone of Darkness and Its Therapeutic Potential and Perspectives*

more resistant to arrhythmic events triggered by rapid changes in plasma electrolyte concentrations, regardless of a lack of effects on the ECG. In addition, those dialysis patients also suffer from disorders in the circadian rhythms and low levels of melatonin [185]. However, clinical translations of our results should be done with caution, mainly because we use a high dose of melatonin administered directly to the heart. Based on melatonin's pharmacokinetics in humans, to achieve a similar concentration in plasma to the one tested *ex vivo*, a dose 10 times higher the highest

*Acute antiarrhythmic mechanisms of melatonin. The red arrows indicate stimulation, and the interrupted blue* 

Melatonin has a remarkable antiarrhythmic activity that is carried out based on actions dependent on and independent of receptor activation. To summarize we propose that the antiarrhythmic effect of melatonin is mediated by receptor activation beyond its outstanding antioxidant actions (**Figure 1**). The shortening of the action potential could be associated with the activation of MT1 melatonin receptors, since they can regulate specific ion channels such as Kir3.1 channel. MT1 and MT2 receptors could indirectly modulate other electrophysiological effects through intracellular signaling such as decreased cyclic adenosine monophosphate,

Endogenous melatonin would be an intrinsically protective factor with therapeutic potential [188, 189]. Melatonin is a promising treatment for cardiovascular diseases such as myocardial ischemia/reperfusion injury, hypertension, and heart failure. It has been shown that melatonin levels were reduced in patients with acute myocardial infarction and in patients undergoing primary coronary angioplasty [190]. These findings suggest that melatonin could play an important role in preventing ischemia/reperfusion heart injury. Indeed, reperfusion arrhythmias increase in pinealectomized animals, suggesting a protective role of endogenous

Chronic melatonin supplementation, either in physiological or pharmacological ranges, protects against arrhythmias [8, 21, 97, 163, 169, 189, 191, 192]. Beyond the reported antioxidant properties of melatonin, it reduces severe ventricular

intravenous dose tested until now should be administered [186, 187].

increased phospholipase C, and PKC activation.

physiological melatonin levels [163, 189].

**3. Chronic antiarrhythmic mechanisms of melatonin**

**58**

**Figure 1.**

*lines indicate blockage.*

arrhythmias by antifibrotic mechanisms, electrical remodeling, direct mitochondrial protection, myocardial Cx43 preservation via PKC signaling, and vitamin D-HSP70/AT1 counterbalance (**Figure 2**). Its cardioprotective properties persist in relevant cardiovascular risk factor models like hypertensive, obese, and nephropathic rats. The latter is interesting because most of the therapeutic interventions postulated so far fail to be reproduced under risk factor conditions.

A preventive approach would be of great value in the face of unpredictable acute arrhythmic events, especially if the intervention manages to avoid the most severe and potentially lethal arrhythmias such as ventricular tachycardia and fibrillation. Numerous efforts have been made in that direction. In the last quarter of the twentieth century, several antiarrhythmic drugs were tried, but most of them showed a proarrhythmogenic profile or failed to reduce mortality [193–196]. A time of great progress was appreciated with the introduction of implantable cardiodefibrillators. However, surgical intervention and high cost limit its population efficiency. A strategy to improve the availability of preventive interventions is to select potential beneficiaries based on their risk of serious events. This would compensate for potential side effects and optimize the investment of resources. Other strategies, such as vaccines, are based on achieving the greatest possible scope with the least number of interventions that attenuate the severity of diseases. In the case of arrhythmias, we still have no clear "antiarrhythmic vaccine." Therefore, risk-oriented strategies would be an acceptable approach.

From a preventive point of view, the pleiotropic protection mechanisms of melatonin could effectively limit the arrhythmic complications associated with hypertension [21, 169]. Arterial hypertension causes vascular deterioration, overloads the heart, and predisposes to a greater number of arrhythmic events. More than five decades ago, it was reported that surgical removal of the pineal gland, a procedure that essentially eliminates circulating levels of melatonin, was followed by a slow but persistent increase in blood pressure in rats [197]. This finding has been confirmed in several subsequent studies [11]. In addition, daily treatment of pinealectomized rats with melatonin attenuated the elevation in blood pressure

#### **Figure 2.**

*Antiarrhythmic mechanisms of chronic melatonin administration. Extracellular lines represent reduced fibrosis after melatonin treatment. The red arrows indicate stimulation and blue ones show blockage. The green arrow marks direct mitochondrial protection.*

that accompanies pinealectomy [198, 199]. Potentially related to these experimental findings are those observational studies in humans that document an age-related gradual increase in blood pressure [7]. Of special interest is that the ability of the pineal gland to produce melatonin is compromised during aging so that the levels of melatonin in the blood at night gradually decrease [28, 30, 110]. An implication of these findings is that the loss of melatonin during aging can contribute to gradual hypertension and arrhythmias.

The structural remodeling of the myocardium that follows hypertension (mainly cardiomyocyte hypertrophy and fibrosis) is accompanied by changes in the expression, distribution and function of the ionic channels of the cell membrane and the intercellular channels constituted by Cx43 [21, 191, 200]. Remodeling predisposes to life-threatening ventricular tachycardia and ventricular fibrillation by early or late postdepolarization and reentry. Melatonin prevents changes in ventricular redistribution of Cx43 and reduces arrhythmia inducibility [8, 21, 147, 191].

Another chronodisruptor that increases arrhythmic risk are kidney diseases. Chronic kidney diseases (CKD) alter the nocturnal secretion of melatonin [185, 201]. Melatonin levels correlate negatively with the intrarenal activity of reninangiotensin II-aldosterone system (RAAS) [202]. Melatonin improves intrarenal RAAS in the 5/6 nephrectomy rat model and reduces blood pressure, oxidative stress, and interstitial fibrosis in the remaining kidneys [203].

Renal diseases cause cardiovascular and electrolytic remodeling that increases the risk of arrhythmias [204–206]. Cardiovascular events occur more frequently in patients with chronic kidney disease. Ventricular arrhythmias are particularly prevalent among patients with CKD, even when those patients do not suffer from any electrolyte imbalance [207]. The risk of mortality also increases in patients with CKD who suffer from an acute coronary syndrome [208]. We demonstrated that unilateral ureteral obstruction caused a cardiac remodeling that was accompanied by an increase in reperfusion arrhythmias [209].

The electrophysiological properties of chronic melatonin deserve attention, due to their relevance for cardiorenal situations with high arrhythmic risk and lack of treatments. We recently confirmed the antifibrotic, antiapoptotic, and antioxidant effects of melatonin and linked them to an HSP 70-VDR/AT1 counterbalance which prevents kidney damage and arrhythmogenic remodeling of the heart [97].

In renal and myocardial tissue, melatonin increased HSP 70 and VDR and decreased AT1 and fibrosis. Melatonin increases HSP 70 and protects the liver of rats exposed to toluene from cytotoxicity induced by oxidative stress [100]. HSP 70 regulates antioxidant responses to cellular oxidative stress and reduces NADPH oxidase activity and expression [210]. We demonstrated a myocardial increase in HSP 70 in rats treated with melatonin. HSP 70 induces VDR and facilitates intracellular localization of active vitamin D metabolites and transactive VDRs [209, 211, 212]. Nuclear melatonin receptors, as members of retinoid-related orphan receptors, may interact and prevent degradation of VDR [97, 103]. Expression of myocardial VDR links chronic kidney disease with cardiovascular disease due to the reduction in VDR that amplifies the effects of angiotensin [212]. Melatonin decreases renal and myocardial overexpression of AT1 [97]. It is well documented that the AT1 pathway leads to myocardial fibrosis during CKD [97]. As previously suggested, the low expression of AT1 through VDR induction could be a consequence of HSP 70-mediated cellular protection [213]. Angiotensin II exerts a tonic modulation of melatonin synthesis by influencing the activity of tryptophan hydroxylase through AT1 supporting the postulated feedback (or reciprocal regulation) between AT1 and melatonin [97, 202].

Additionally, the mitochondrial dynamics relates to the RAAS. We show that melatonin prevents mitochondrial edema, high activity of NADPH oxidase, and

**61**

effects of melatonin.

chronic antiarrhythmic.

**Acknowledgements**

**Conflict of interest**

The authors declare no conflict of interest.

**4. Conclusions**

*Melatonin for a Healthy Heart Rhythm*

*DOI: http://dx.doi.org/10.5772/intechopen.91447*

apoptosis. In this sense, the reduction of mitochondrial damage melatonin could be related to the negative regulation of AT1. The induction of HSP 70 through melatonin is compatible with an additional mechanism related to Tom 70. Furthermore, Tom 70 regulates melatonin-induced protection against myocardial infarction [115, 116]. All these data allow us to assume that the induction of HSP 70 by melatonin and the reduction of AT1 are critical components of the cellular stress response. We attribute the higher vulnerability to ventricular fibrillation during reperfusion in the kidney disease rat model to the prooxidative and profibrotic changes that accompanied the increase in AT1 and the decrease in HSP 70 [97, 209]. Myocardial oxidative stress—particularly in the mitochondria—and fibrosis are well-known proarrhythmic substrates [55, 71]. Free radicals act as triggers for the beginning of arrhythmic events. The persistence of high-frequency rhythms requires reentry circuits [214]. Altered conduction and shortening of the action potential contribute

to the complex reentry mechanisms involved in ventricular fibrillation.

Melatonin protection against myocardial remodeling induced by kidney disease is one of the factors that protect against ventricular fibrillation. Chronic melatonin prolongs the action potential duration and hyperpolarizes the cardiomyocytes. These changes are the first report of myocardial action potential modifications by chronic administration of melatonin. Opening of Kir3.x channels by melatonin receptor activation could explain hyperpolarization [131]. The action potential lengthening is harder to explain because melatonin activates currents involved in the action potential repolarization and the only inhibitory effect of melatonin against outward potassium currents was described in neurons [90, 129, 130, 215–217]. As previously mentioned, the downregulation of volume-activated anionic currents can explain attenuated response to action potential shortening induced by ischemia [126].

A synthesis of the mechanisms of protection of chronic treatment of melatonin cardiovascular complications is outlined in **Figure 2**. We focus our attention on the preventive effects of melatonin against the alteration of Cx43, mitochondrial oxidant capacity, and membrane potentials. In addition, modulation of the AT1 and VDR receptors related to the increase of HSP 70 contributes to the cardioprotective

Melatonin is the rhythmic protector of healthy heart rhythm and a promising preventive agent against ventricular fibrillation, the most lethal and disorganized heart rhythm. Pleotropic effects of melatonin make it an exceptional acute and

This work was supported by grant by ESC research grant funded by the European Society of Cardiology, by ERC-2014-StG 638284 funded by the European Research Council, by project DPI2016-75458-R funded by MINECO (Spain) and FEDER, and by Reference Group BSICoS T39-17R and project LMP124-18 funded by Gobierno de Aragón and FEDER 2014-2020 "Building Europe from Aragón."

#### *Melatonin for a Healthy Heart Rhythm DOI: http://dx.doi.org/10.5772/intechopen.91447*

*Melatonin - The Hormone of Darkness and Its Therapeutic Potential and Perspectives*

hypertension and arrhythmias.

that accompanies pinealectomy [198, 199]. Potentially related to these experimental findings are those observational studies in humans that document an age-related gradual increase in blood pressure [7]. Of special interest is that the ability of the pineal gland to produce melatonin is compromised during aging so that the levels of melatonin in the blood at night gradually decrease [28, 30, 110]. An implication of these findings is that the loss of melatonin during aging can contribute to gradual

The structural remodeling of the myocardium that follows hypertension (mainly cardiomyocyte hypertrophy and fibrosis) is accompanied by changes in the expression, distribution and function of the ionic channels of the cell membrane and the intercellular channels constituted by Cx43 [21, 191, 200]. Remodeling predisposes to life-threatening ventricular tachycardia and ventricular fibrillation by early or late postdepolarization and reentry. Melatonin prevents changes in ventricular redistribution of Cx43 and reduces arrhythmia inducibility [8, 21, 147, 191].

Another chronodisruptor that increases arrhythmic risk are kidney diseases. Chronic kidney diseases (CKD) alter the nocturnal secretion of melatonin [185, 201]. Melatonin levels correlate negatively with the intrarenal activity of reninangiotensin II-aldosterone system (RAAS) [202]. Melatonin improves intrarenal RAAS in the 5/6 nephrectomy rat model and reduces blood pressure, oxidative

Renal diseases cause cardiovascular and electrolytic remodeling that increases the risk of arrhythmias [204–206]. Cardiovascular events occur more frequently in patients with chronic kidney disease. Ventricular arrhythmias are particularly prevalent among patients with CKD, even when those patients do not suffer from any electrolyte imbalance [207]. The risk of mortality also increases in patients with CKD who suffer from an acute coronary syndrome [208]. We demonstrated that unilateral ureteral obstruction caused a cardiac remodeling that was accompanied

The electrophysiological properties of chronic melatonin deserve attention, due to their relevance for cardiorenal situations with high arrhythmic risk and lack of treatments. We recently confirmed the antifibrotic, antiapoptotic, and antioxidant effects of melatonin and linked them to an HSP 70-VDR/AT1 counterbalance which

Additionally, the mitochondrial dynamics relates to the RAAS. We show that melatonin prevents mitochondrial edema, high activity of NADPH oxidase, and

prevents kidney damage and arrhythmogenic remodeling of the heart [97]. In renal and myocardial tissue, melatonin increased HSP 70 and VDR and decreased AT1 and fibrosis. Melatonin increases HSP 70 and protects the liver of rats exposed to toluene from cytotoxicity induced by oxidative stress [100]. HSP 70 regulates antioxidant responses to cellular oxidative stress and reduces NADPH oxidase activity and expression [210]. We demonstrated a myocardial increase in HSP 70 in rats treated with melatonin. HSP 70 induces VDR and facilitates intracellular localization of active vitamin D metabolites and transactive VDRs [209, 211, 212]. Nuclear melatonin receptors, as members of retinoid-related orphan receptors, may interact and prevent degradation of VDR [97, 103]. Expression of myocardial VDR links chronic kidney disease with cardiovascular disease due to the reduction in VDR that amplifies the effects of angiotensin [212]. Melatonin decreases renal and myocardial overexpression of AT1 [97]. It is well documented that the AT1 pathway leads to myocardial fibrosis during CKD [97]. As previously suggested, the low expression of AT1 through VDR induction could be a consequence of HSP 70-mediated cellular protection [213]. Angiotensin II exerts a tonic modulation of melatonin synthesis by influencing the activity of tryptophan hydroxylase through AT1 supporting the postulated feedback (or reciprocal regulation) between AT1

stress, and interstitial fibrosis in the remaining kidneys [203].

by an increase in reperfusion arrhythmias [209].

**60**

and melatonin [97, 202].

apoptosis. In this sense, the reduction of mitochondrial damage melatonin could be related to the negative regulation of AT1. The induction of HSP 70 through melatonin is compatible with an additional mechanism related to Tom 70. Furthermore, Tom 70 regulates melatonin-induced protection against myocardial infarction [115, 116]. All these data allow us to assume that the induction of HSP 70 by melatonin and the reduction of AT1 are critical components of the cellular stress response.

We attribute the higher vulnerability to ventricular fibrillation during reperfusion in the kidney disease rat model to the prooxidative and profibrotic changes that accompanied the increase in AT1 and the decrease in HSP 70 [97, 209]. Myocardial oxidative stress—particularly in the mitochondria—and fibrosis are well-known proarrhythmic substrates [55, 71]. Free radicals act as triggers for the beginning of arrhythmic events. The persistence of high-frequency rhythms requires reentry circuits [214]. Altered conduction and shortening of the action potential contribute to the complex reentry mechanisms involved in ventricular fibrillation.

Melatonin protection against myocardial remodeling induced by kidney disease is one of the factors that protect against ventricular fibrillation. Chronic melatonin prolongs the action potential duration and hyperpolarizes the cardiomyocytes. These changes are the first report of myocardial action potential modifications by chronic administration of melatonin. Opening of Kir3.x channels by melatonin receptor activation could explain hyperpolarization [131]. The action potential lengthening is harder to explain because melatonin activates currents involved in the action potential repolarization and the only inhibitory effect of melatonin against outward potassium currents was described in neurons [90, 129, 130, 215–217]. As previously mentioned, the downregulation of volume-activated anionic currents can explain attenuated response to action potential shortening induced by ischemia [126].

A synthesis of the mechanisms of protection of chronic treatment of melatonin cardiovascular complications is outlined in **Figure 2**. We focus our attention on the preventive effects of melatonin against the alteration of Cx43, mitochondrial oxidant capacity, and membrane potentials. In addition, modulation of the AT1 and VDR receptors related to the increase of HSP 70 contributes to the cardioprotective effects of melatonin.

## **4. Conclusions**

Melatonin is the rhythmic protector of healthy heart rhythm and a promising preventive agent against ventricular fibrillation, the most lethal and disorganized heart rhythm. Pleotropic effects of melatonin make it an exceptional acute and chronic antiarrhythmic.
