Arrhythmias in the developing heart

18 Pages·2014·1.26 MB·English

ActaPhysiol2014
REVIEW
Arrhythmias in the developing heart
D. Sedmera,1,2 R. Kockova,2,3 F. Vostarek2 and E. Raddatz4
1 Institute ofAnatomy,First Faculty ofMedicine, Charles University, Prague, CzechRepublic
2 Institute ofPhysiology,Academy ofSciencesofthe Czech Republic,Prague, Czech Republic
3 Department ofCardiology,Institute ofClinicalandExperimental Medicine, Prague, Czech Republic
4 Department ofPhysiology,Faculty ofBiologyandMedicine, University ofLausanne, Lausanne, Switzerland
Received13August2014, Abstract
revisionrequested8September Prevalence of cardiac arrhythmias increases gradually with age; however,
2014,
specific rhythm disturbances can appear even prior to birth and markedly
revisionreceived1October
affect foetal development. Relatively little is known about these disorders,
2014,
chiefly because of their relative rarity and difficulty in diagnosis. In this
accepted23October2014
Correspondence:D.Sedmera, review, we cover the most common forms found in human pathology, spe-
AcademyofSciencesofthe cifically congenital heart block, pre-excitation, extrasystoles and long QT
CzechRepublic,Instituteof syndrome. In addition, we cover pertinent literature data from prenatal
Physiology,Videnska1083,14220
animal models, providing a glimpse into pathogenesis of arrhythmias and
Prague4,CzechRepublic.
possible strategies for treatment.
E-mail:[email protected]
Keywords anti-arrhythmicdrugs,cardiacdevelopment,chickembryo,con-
ductionsystem,hypoxia,mouse.
using the ‘heart failure score’ presented by Huhta
Disturbances of cardiac rhythm in the human
(2005).
foetus
Using all three standard echocardiographic modali-
During routine obstetric examination, foetal rhythm ties (B-mode, M-mode and Doppler), we can assess
disturbances may be detected in at least 2% of preg- atrial andventricular contraction frequencies and their
nancies (Copel et al. 2000, Jaeggi & Nii 2005). Foetal timerelations.AnequivalentforPwaveonelectrocar-
arrhythmias account for about 10–20% of referrals diogram is the A wave detected by pulse wave Dopp-
for foetal cardiology assessment (Srinivasan & Stras- ler in mitral inflow or atrial wall motion detected by
burger 2008). Due to a number of limitations, a foetal M-mode. Similarly, the beginning of retrograde flow
electrocardiogram (cardiotocogram) is not the ideal in the superior vena cava indicates the beginning of
method for assessment of arrhythmias. A relatively atrial systole. Atrioventricular (AV) valve closure,
novel and efficient method for foetal heart electrical semilunar valve opening and positive Doppler flow in
activity recording is foetal magnetocardiography the aorta are equivalents of the beginning of QRS
(Strasburger et al. 2008, Strasburger & Wakai 2010). complex. Simultaneous Doppler recording in the supe-
However, this method is not widely available and is rior vena cava and the aorta shows the time correla-
preferred only after the 20th week of gestation tion between the atrial and ventricular systole – times
because it is less reliable in the earlier stages of preg- corresponding to the P wave andQRS complex on the
nancy. Thus, echocardiography remains the principal ECG. These parameters allow us to calculate the
method in evaluation of heart rhythm disturbance in heart rate, AV delay and diagnose different types of
the foetus. In addition to heart rhythm analysis, echo- arrhythmias by measuring the mechanical response of
cardiography may reveal other signs associated with the heartchambers to theelectrical stimulus.
prolonged or persistent foetal rhythm disturbances, The relationship between foetal arrhythmia and
such as hydrops (pleural or pericardial effusion, asci- structural heart disease is not clearly established.
tes) in its early as well as more advanced stages. The Stewart and Copel found no clear relationship
severity of foetal heart failure can be then monitored between foetal arrhythmia and structural heart disease
©2014ScandinavianPhysiologicalSociety.PublishedbyJohnWiley&SonsLtd,doi:10.1111/apha.12418 1
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Foetalcardiacarrhythmias DSedmeraetal. ActaPhysiol2014
(Stewart et al. 1983, Copel et al. 2000). In the obser- (a)
vational study published by Stewart and associates,
only two foetuses from 17 with documented ectopic
beats had structural heart disease. No structural heart
disease was found in five foetuses with tachycardia
(heart rate over 180 bpm), but four of eight foetuses
with documented bradycardia had severe structural
heart disease. Copel and colleagues reported only two
of 10 foetuses diagnosed with significant arrhythmia –
one with supraventricular tachycardia and one with a
second-degree AV block – associated with structural
heart disease of all 614 foetuses with irregular heart
rhythm. On the other hand, there is evidence that foe-
(b)
tal arrhythmia may be associated with structural heart
disease. Schmidt et al. (1991) reported that 53% of
foetuses (of a total of 55) with complete AV block
had concomitant structural heart disease (left atrial
isomerism, discordant AV connection). Vergani et al.
(2005) reported structural heart anomalies in five of
six foetuses with bradycardia from a total cohort of
114 infants with foetal arrhythmias. Only two of four
foetuses with AV block survived. Eronen reported 12
foetuses (three supraventricular and three ventricular
ectopic activities, four AV blocks and two sinus
bradycardias) with significant arrhythmia associated
with structural heart disease from a total of 125 foe-
Figure1 Epidemiologyoffoetalarrhythmiasinhumans.(a)
tuses with significant arrhythmia (Eronen 1997). She
Incidenceofvarioustypesofarrhythmiasinnon-selected
also found 95% survival in foetuses with sole signifi-
population(N=406,collatedfromreferences(Copeletal.
cant arrhythmia compared to a 75% mortality in
2000),(Verganietal.2005)).(b)Incidenceinhighlyselected
those with arrhythmia associated with structural heart population(N=591,collatedfromreferences(Stewartetal.
disease. Interestingly, the total mortality in the group 1983),(Reedetal.1990),(Eronen1997),(Verganietal.
of foetuses with structural heart disease was only 2005),(Zhaoetal.2006)).
67%. Based on these two observational studies, it
could be speculated that bradyarrhythmias are more
frequently associated with structural heart disease None of these were associated with structural heart
and have a worse outcome than tachyarrhythmias disease.
or irregular heart rhythm, which are frequently cur- Prolonged foetal tachycardia is usually a serious
able or might resolve spontaneously during develop- condition often leading to foetal hydrops or even
ment. death. Simpson & Sharland (1998) reported hydrops
For simplicity, we may divide foetal arrhythmias occurrence in 41% of 127 foetuses diagnosed with
into the three groups (Fig. 1): ectopic beats, mostly tachycardia. Seventy-five non-hydropic foetuses from
originating in atrial ectopic foci; tachyarrhythmias, this cohort responded well to transplacental treatment
which are defined as heart rates over 180 bpm; and (mostly with digoxin) with an excellent survival to
bradyarrhythmias, defined as heart rates below birth (96%). Conversely, only two-thirds of hydropic
110 bpm(Jaeggi &Nii 2005). foetuses with tachycardia responded to transplacental
Of these three types, extrasystoles typically have the treatment, and of these, only 73% survived till birth.
best outcomes (Reed 1989). Vergani et al. (2005) Thus, foetal hydrops is a negative prognostic sign sug-
reported that 38% of cases with extrasystoles (in 87 gesting severe hemodynamic consequences from the
foetuses)resolvedinuteroand49%atbirth.Onlyone underlying causes – for example, arrhythmia and/or
neonate required postnatal therapy, and in nine neo- structural heartdisease.
nates, the arrhythmia was still present at 1-year fol- Sustained or prolonged bradycardia (heart rates
low-up without need for therapy. Two foetuses with <100 bpm) or tachycardia (heart rates over 180 bpm)
extrasystolesconvertedtosupraventriculartachycardia are of clinical significance and might have a significant
in utero and were successfully treated pharmacologi- impact on further foetal development in utero; even
cally with no impact on their further development. later postnatal development might be affected. Jaeggi
2 ©2014ScandinavianPhysiologicalSociety.PublishedbyJohnWiley&SonsLtd,doi:10.1111/apha.12418
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ActaPhysiol2014 DSedmeraetal. Foetalcardiacarrhythmias
and Nii reported foetal tachycardia as the most fre- limited. For significant number of foetuses with
quent arrhythmia in the foetus and was present in complete heart block caused by maternal autoantibod-
57% of 66 foetuses examined with proven serious ies, transplacental treatment with beta-receptor-stimu-
arrhythmia (Jaeggi & Nii 2005). Supraventricular lating agents, corticosteroids or immunosuppressives is
tachycardia was present in 40% of cases, atrial flutter recommended. In principle, foetal pacemaker implan-
accounted for 11%, and sinus tachycardia was present tation (Liddicoat et al. 1997) should be considered
in6%. using minimally invasive techniques (Sydorak et al.
2001,Eghtesadyet al.2011,Nicholsonet al.2012).
During sinus bradycardia, there is 1 : 1 AV cou-
Diagnosis, classification and management of
pling with a slow frequency of atrial contractions
foetal arrhythmias
(<100 bpm). Simple sinus bradycardia may be caused
The12-leadECGthatissousefulinnewbornoradult by foetal distress with episodes of hypoxia and blood
cardiology suffers from major limitations in the foe- flow redistribution, while brain and heart are sup-
tus. Echocardiography is typically the only way to plied preferentially. Sinus bradycardia can be a mani-
diagnose tachyarrhythmia in the foetus, and it is not festation of foetal long QT syndrome, and all
easy to differentiate between different types of tach- newborns with a history of foetal heart rate below
yarrhythmias. Supraventricular tachycardia with the 3rd percentile should be assessed for this entity
mostly 1 : 1 AV conduction can be distinguished from early after birth (Mitchell et al. 2012). Sinus brady-
atrial flutter, with mostly 2 : 1 AV conduction block, cardia may be a rare manifestation of sinus node dys-
due to excessive atrial frequency in flutter(about 440– function. Supraventricular bigeminy or trigeminy with
480 bpm) translating into a 220–240 bpm ventricular AV block must always be excluded when assessing
rate. In AV re-entry, the time interval between the the foetus for bradycardia. The telltale sign would be
ventricular and atrial activity would be short, while in an atrial frequency above that of the ventricle and an
atrial tachycardia originating from ectopic foci, this irregular heart rhythm. The outcome is usually
time interval is usually prolonged. Ventricular benign and this arrhythmia mostly does not require
tachycardia with typical dissociation of ventricular treatment.
and atrial rhythm or conducted 1 : 1 from ventricles
to atria is extremely rare in the foetus as most
Foetal AV block
tachyarrhythmias originate in the atria. In such
cases, it is clear that only an experienced physician The most frequent cause of bradycardia is congenital
trained in echocardiography can make the correct AV block. First-degree AV block is characterized by
diagnosis. prolonged AV conduction with 1 : 1 AV coupling. It
The most frequent foetal tachyarrhythmia is supra- is necessary to realize that AV conduction time
ventricular tachycardia represented by three different increases during gestation and the exact numbers
types:AVre-entranttachycardia,permanentjunctional also differ for various ECHO modalities. Normal
reciprocating tachycardia and atrial ectopic tachycar- values for 30–34 weeks of gestational stage are
dia. The second most frequent foetal tachyarrhythmia 122.7 (cid:1) 11.1 ms by left ventricle inflow/outflow
is atrial flutter caused by a macro-re-entry circuit Doppler method, 116.5 (cid:1) 8.8 ms by Doppler method
located in the atria. The final differentiation is often in the superior vena cava/aorta, 142.4 (cid:1) 14.2 by
made only after birth when the arrhythmia persists or atrial contraction/ventricular systole as measured via
reoccurs, or a delta wave typical for the accessory Tissue Doppler Imaging (TDI) of the basal right ven-
pathwayispresentonthe12-leadECG.Thetreatment tricularfree wall (Niiet al. 2006).
strategyformosttypesoftachyarrhythmiasisbasedon We distinguish two types of second-degree AV
transplacental digoxin administration in non-hydropic block. Wenckebach type (Mobitz I) second-degree AV
foetuses. Sotalol, flecainide or amiodarone is mostly block is characterized by the gradual lengthening of
reserved for hydropic foetuses or more resistant AV conduction time terminated by a dropped ventric-
tachyarrhythmias.Treatment isrequiredforpuresinus ular contraction. Mobitz type (Mobitz II) of second-
tachycardia with typical heart rates of 180–200 bpm degree AV block is typified bysudden loss of ventricu-
usually caused by foetal distress, foetal thyrotoxicosis, lar contraction, while AV conduction time remains
anaemiaetc. unchanged. A specific type of Mobitz II AV block is
Sustained or prolonged bradycardia is present in 2 : 1 conduction when every second atrial beat is not
43% of significant foetal arrhythmia cases, as pre- conductedto the ventricles.
sented by Jaeggi & Nii (2005). Complete AV block The third-degree AV block (complete heart block)
accounts for38%,andonly5%manifestassinus bra- has the most serious impact on further foetal develop-
dycardia cases. The treatment of foetal bradycardia is ment leading frequently to foetal demise. Atrial and
©2014ScandinavianPhysiologicalSociety.PublishedbyJohnWiley&SonsLtd,doi:10.1111/apha.12418 3
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Foetalcardiacarrhythmias DSedmeraetal. ActaPhysiol2014
ventricular electrical and mechanical activities are
Importance of the cardiac conduction system
completely independent in this type of AV block. This
for the origin of arrhythmias
always leads to significant and prolonged bradycar-
dia. The only physiological pathway to compensate It is widely recognized in clinical practice that the car-
the decrease in cardiac output caused by bradycardia diac conduction system (CCS) can be a focal point of
is the Frank-Starling mechanism which might be lim- arrhythmogenesis (Braunwald et al. 2001). This pro-
ited at early stages according to data from animal pensity was extensively analysed from developmental
experiments (Kockova et al. 2013). When increased perspective by Jongbloed and associates (Jongbloed
stroke volume fails to compensate severe foetal bra- et al. 2004) using CCS-LacZ transgenic mouse model.
dycardia, heart failure occurs leading to foetal Detailed analysis of the developing CCS was per-
hydrops. formed on hearts at embryonic day (ED) 9.5–15.5
Foetal complete heart block occurs more frequently stained for beta-galactosidase activity and co-stained
in conjunction with various congenital structural heart with the myocardial marker HHF35 followed by
diseases (Stewart et al. 1983, Jaeggi & Nii 2005, Ver- three-dimensional reconstruction. CCS-lacZ expres-
gani et al. 2005). Schmidt reported that 53% of foe- sion detected by X-gal staining was observed in the
tuses diagnosed with complete heart block had sinoatrial node, left and right venous valves, septum
associated complex congenital heart disease (Schmidt spurium, right and left AV ring, His bundle, bundle
et al. 1991). Another major reason for congenital branches, moderator band, Bachmann’s bundle, left
complete heart block is maternal autoimmune disease atrial posterior wall surrounding the pulmonary
such as lupus erythematosus, Sjo€gren syndrome, rheu- venous orifice and later on in the pulmonary vein
matoid arthritis or unclassified systemic rheumatoid wall. These data supported the idea that areas derived
disease. Elevated titres of anti – Ro/SSA and anti – from the developing CCS may form the arrhythmo-
La/SSB antibodies are typically found in mothers genicsubstrate inadult hearts.
affected by the above-mentioned autoimmune dis- A comparative study between patients with left
eases. The risk of developing foetal complete heart atrial tachycardia originating from the junction of
block in pregnant women with positive anti-Ro/SSA mitral annulus and aortic ring and mouse embryos
antibodies is about 2% (Brucato et al. 2001). These demonstrated the presence of the developing special-
antibodies cause myocardial inflammation specifically ized conduction system in this region starting at
affecting the AV node leading to various degrees of embryonic age11.5(Gonzalez et al. 2004).
AV conduction impairment, which usually occurs Particular attention was focused on the develop-
around 20–24 gestational weeks. This might also pres- mental origin of pulmonary vein myocardium (Mom-
ent as endomyocardial fibrosis in the foetus or new- mersteeg et al. 2007a), which is derived from the
born. Because complete heart block has been shown second heart field. The area around the pulmonary
to be associated with very highmortalityrates ranging veins entranceis in humansa frequent site of origin of
between 18% and 43% (Jaeggi & Nii 2005), there atrial fibrillation, so its electrical insulation by cathe-
has been a major effort to prevent this autoimmune ter intervention is a frequent procedure during clinical
disease. Corticosteroids were administrated to preg- intervention for ablation of this increasingly prevalent
nant women with positive titres of autoantibodies human arrhythmia. A recent study based on HCN4-
intravenously or orally (Reinisch et al. 1978, Fried- Cre mouse line with LacZ or eGFP reporter (Liang
man et al. 2009), but major side effects were noticed et al. 2013) precisely delineated relative contributions
afterwards including oligohydramnion, foetal adrenal of first and second heart lineages to the CCS and pro-
suppression, intrauterine grow retardation and so on. vided a time line of developmental expression of this
Corticosteroid treatment is recommended only for CCS marker in concert with other markers during its
advanced heart block with significant and prolonged formation.
bradycardia with a high risk of hydrops development.
Isolated prolongation of AV conduction only rarely
Genetic and epigenetic determination of the
leads to progressive AV block, as shown by Jaeggi
CCS
et al. (2011) in anti-Ro and anti-La positive mothers,
and corticosteroid treatment is therefore not recom- To better appreciate the developmental potential of
mended. Recently, current recommendations of the CCS to generate arrhythmias, one needs to consider
American Heart Association regarding diagnosis and themechanismsgoverningitsinductionandpatterning
management of foetal heart disease, including prenatal (reviewed in (Gourdie et al. 2003), (Christoffels et al.
arrhythmias, were summarized in a form of Scientific 2010). Lineage tracing experiments performed by the
Statement (Donofrioet al.2014). Mikawa lab have shown that cardiac pacemaker cells
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ActaPhysiol2014 DSedmeraetal. Foetalcardiacarrhythmias
are physically segregated and molecularly pro- and intracellular calcium cycling is of lesser impor-
grammed in a tertiary heart field prior to the onset of tanceat thisearlystage.
cardiac morphogenesis, and this process depends on From the functional side, endothelin signalling was
Wnt signalling (Bressan et al. 2013). Recently, the shown to be necessary not for specification like in
genetic cascade governing specification of cardiac birds (Gourdie et al. 1998), but normal function of
pacemaking tissues was elucidated by the Amsterdam the embryonic pacemaker in mammals (Karppinen
group (Mommersteeg et al. 2007b). Restricted expres- et al. 2013). Stimulation with endothelin-1 increased
sion pattern of the homeodomain transcription factor beatingfrequency of ED9–ED11cardiomyocytes. Inhi-
Shox2 in the sinus venosus myocardium, including the bition by receptor antagonist tezosentan led to dose-
sinoatrial nodal region and the venous valves, was dependent bradycardia in vitro as well as in utero, but
found to be important for the recruitment of these only during the early (ED12.5) and not late (ED18.5)
cells to the pacemaking fate (Blaschke et al. 2007). embryonic stages. Irregular rhythm was also observed,
The authors furthermore demonstrated aberrant and use of specific antagonists indicated that the
expression of gap junction proteins connexin 40 and effects are mediated viaendothelin receptorB.
43 as well as the transcription factor Nkx2.5 specifi- Location of thefirst activation site inthe ratembry-
cally within the sinoatrial nodal region, leading to onic heart was investigated by the Kamino group (Hi-
embryonic lethality between ED11.5 and ED13.5 in rota et al. 1985). At the time of heartbeat initiation,
Shox2(cid:3)/(cid:3) mice. Finally, they showed that Shox2 defi- the first pacemaking activity was located in the left
ciency interferes with pacemaking function in embry- side of the sinus venosus, but within a few hours
onic zebrafish in vivo. Particular attention was also migrated to the right side, where the definitive pace-
devoted to specification of pulmonary venous myocar- maker islocated. Asimilar situation wasreported also
dium (Mommersteeg et al. 2007a), which is a signifi- in avian embryos; remnants of this initial left-sided
cant source of atrial fibrillation. Genetic labelling activity were reported in a small proportion of normal
reveals that atrial cells do not contribute to this spe- avian hearts at later stages of development (Sedmera
cific population, characterized by Nkx2.5 expression et al. 2006); under normal conditions, no such left-
distinguishing it from the systemic venous return. sided activity was reported in a large series of embry-
MaintenanceofthisphenotypeisdependentonPitx2c, onic mice (Leaf et al. 2008, Ammirabile et al. 2012,
which prevents it from adopting the Cx40-negative, Benes et al. 2014).
Hcn4-positive pacemaking phenotype of the right- Considerably less is known about the mechanisms
sided sinoatrial node. regulating specification of the remaining components
Embryonic pacemaking differs in details from the of the CCS. Neuregulin was proposed as a factor
mechanisms operating in the adult sinoatrial node. influencing differentiation of the ventricular myocytes
The early stages are crucially dependent on the cal- towards the conduction phenotype (Rentschler et al.
cium clock, as demonstrated by Wakimoto et al. 2002), but this secreted molecule has many important
(2000) who studied the functional importance of functions in the embryonic cardiomyocytes, such as
sodium-calcium exchanger (NCX) for heartbeat initia- their survival (Liu et al. 2010). The neuregulin/Erb
tion and maintenance. To address this question, they signalling cascade could function in concert with
generated Ncx1-deficient mice by gene targeting to endothelin signalling, which was shown to be impor-
determine the in vivo function of the exchanger. The tant in Purkinje fibre differentiation in the chick
hearts of Ncx(cid:3)/(cid:3) embryonic deficiency in Ncx, (Gourdie et al. 1998, 2003, Takebayashi-Suzuki et al.
embryos did not beat, and cardiomyocytes frequently 2000, Sedmera et al. 2008). Other important factors
underwent apoptosis leading to embryonic lethality participating in formation of the His bundle and its
between ED9 and ED10. branches include Nkx2.5 (Jay et al. 2004), Irx3
To study cardiac physiology near the onset of the (Zhang et al. 2011) and T-box transcription factors
heartbeat in embryonic mouse hearts, Chen and asso- (Jerome & Papaioannou 2001, Moskowitz et al.
ciates performed dual optical mapping of membrane 2004, Hoogaars et al. 2007, Aanhaanen et al. 2009,
voltage and intracellular calcium (Chen et al. 2010). Frank et al. 2012).
Action potentials and calcium transients were detected Differentiation of embryonic myocytes into the con-
in approx. 50% of mouse embryo hearts at ED8.5 ducting phenotype is governed also by the epigenetic
and 100% at E9.0, indicating that the heartbeat starts factors, of which mechanical loading is of the most
between ED8 and ED9. Cardiac activity was abol- criticalimportance.Invitrounloadingofchickembry-
ished by calcium channel blocker nifedipine and the I onic hearts (Sankova et al. 2010) led to de-differentia-
(f) blocker ZD7288, suggesting that both HCN4 and tion of the ventricular conduction system that could
voltage-dependent calcium channels are important for be rescued by ventricular stretching using a droplet of
embryonic pacemaking. The role of sodium channels silicone oil. These experiments resolved the issue
©2014ScandinavianPhysiologicalSociety.PublishedbyJohnWiley&SonsLtd,doi:10.1111/apha.12418 5
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Foetalcardiacarrhythmias DSedmeraetal. ActaPhysiol2014
arisingfrompreviousinvivostudiesusingalteredhae- increased after isoproterenol stimulation with 6%
modynamics models (Reckova et al. 2003, Hall et al. incidence of ventricular ectopic rhythms. Addition of
2004) that showed that increased hemodynamic load- carbachol after isoproterenol caused dissociated ante-
ing accelerated, while reduced ventricular preload grade and retrograde AV ring conduction in almost
inhibited ventricular CCSdifferentiation byattributing 10% of ED10.5–ED11.5 hearts. Re-entry persisting
the stimulus to myocyte stretching, rather than to for multiple beats was also observed, but none
shear stress-induced signallingfrom theendocardium. occurred at ED9.5. Rare cases of irregular rhythm
Studies on chick embryos in vivo showed that (sinoatrial and AV block, alternating patterns of ven-
hypoxia can accelerate maturation of the AV junction tricular activation) were also observed in our large
andleadtoearlierappearanceofmature(apex-to-base) mouse series (Sankova et al. 2012), while ectopics
ventricular activation patterns (Nanka et al. 2008), originating from the outflow tract myocardium were
possibly through increased apoptosis of the AV myo- seen exclusively in ED10.5–ED11.5 hearts cultured
cardium. Another player in developing proper fibrous for 24 h (Vostarek et al. 2014). AV re-entry was
insulation of the AV junction is the developing epicar- observedas a rarity inoneED4 chickheart (Fig. 2).
dium (Kolditz et al. 2007, 2008), and perturbations of
this process may lead to ventricular pre-excitation.
Genetic mouse models of arrhythmias
ElectricalinsulationoftheHisbundleisalsodependent
onimmigratingcardiacneuralcrestcells(Gurjarpadhye As noted above, spontaneous arrhythmias are very
et al.2007). rare in normal embryonic hearts, thus facilitating
analysis of results in experimental perturbation mod-
els. The importance of catecholaminergic signalling in
Spontaneously occurring arrhythmias in
development and function of the CCS was recently
embryos
reported by Steve Ebert’s group (Baker et al. 2012).
This area of embryonic arrhythmias is not well inves- These results are in good agreement with previous
tigated for numerous reasons. First, there are the studies showing lethality of mouse embryos deficient
methodological difficulties inherent to all observa- in a component of adrenergic signalling, beta-adrener-
tionalstudiesofmammalianembryosthatareshielded gicreceptorkinase (Jaber et al. 1996).
in utero by maternal tissues. The most significant StudiesbyCollinPhoonvalidatedultrasoundbiomi-
breakthrough in this respect was availability of high- croscopy as a prime tool for in vivo identification of
resolution ultrasound (Phoon et al. 2002, Phoon abnormal mouse embryonic heart function, including
2006, Nomura-Kitabayashi et al. 2009, Lo et al. arrhythmias.Usingthistechnique,theystudiedlongitu-
2010), paralleling the advances in human embryonic dinally embryonic ED10.5–ED14.5 NFATc1(cid:3)/(cid:3)
echocardiography (Maeno et al. 1999, Pedra et al. embryos and control littermates (Phoon et al. 2004).
2002). The second obstacle is the relative rarity of Thenullembryos,lackingtheoutflowvalves,dieprior
such events (compare with the situation in clinical set- to completion of ventricular septation from presumed
tings, discussed above) necessitating the examination heartfailure.Theauthorsshowedthatabnormalblood
of large numbers of embryos. Therefore, most ar- flowwaspresentatE12.5whenoutflowvalvesnormally
rhythmias detected in the embryonic hearts could be firstdevelop.Reducedcardiacoutputanddiastolicdys-
at least in part be due to ‘gentle’ alterations of physio- function contributed to heart failure, but contractile
logical conditions, as it is close to impossible to moni- function remained unexpectedly normal. The only
tor the embryonic mammalian heart in a completely arrhythmiadetectedpriortoembryonicdemisewaspro-
non-invasive manner. gressive bradycardia, indicating that embryonic heart
Various arrhythmias in the isolated mouse embry- failureoccursrapidlyinthismousemodel.
onic heart were revealed using simultaneous voltage Mutations in TBX3 cause congenital anomalies in
and calcium optical mapping (Valderrabano et al. patients with ulnar-mammary syndrome (Frank et al.
2006). The focus of this study was on AV conduction 2012). Data from both mice and humans suggest mul-
during transition from immature base-to-apex to tiple roles of this transcription factor in morphogene-
mature apex-to-base ventricular activation pattern. sis and function of the CCS. Disruption of Tbx3
The authors hypothesized that after this transition, the function in different regions of the developing heart
remnants of the myocardial AV ring remain tran- caused discrete phenotypes and lethal arrhythmias.
siently able to conduct, providing a possible substrate Sinus pauses (normally present at low frequency in
for arrhythmias. They noted that arrhythmias were adult mice) and bradycardia indicated sinoatrial node
rare under normal conditions, with only occasional dysfunction; pre-excitation and AV block revealed
AV blocks (4%) and junctional rhythms in four of problems in the AV junction. These arrhythmias were
309embryonicheartsanalysed.Thefrequencynotably accompanied by perturbed expression of several ion
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ActaPhysiol2014 DSedmeraetal. Foetalcardiacarrhythmias
Figure2 Atrioventricularre-entryinED4chickembryonicheart.Toppanelsshowtheembryonicheartfromthebackandtime
courseofcalciumtransients.Thefirstderivativepanelshowsbothprogradeactivation(fromtheatriumtotheventricletothe
outflowtract,orangearrow)andretrogradeactivationfromtheventriclebacktotheAVcanalandatrium(greyarrow).Activa-
tionmapsinthebottomdepictthisphenomenonatdifferenttemporalscales.Notethattheactivationpatternoftheatriumdif-
fersbetweenprogradeandretrogradeactivation.A,atrium;AV,atrioventricularcanal;V,ventricle;OT,outflowtract.For
bettervisualizationoftheactivationsequence,seetheMovieS1.
channel components (e.g. upregulation of Kcne3, of prescribed medicines. This does not only impact
Chac1, Kcnj4 and downregulation of Scn7a), despite overt morphological anomalies, but also more subtle
normal expression of previously identified CCS mark- functional alterations, such as mild neurological
ers, raising the possibilityof functional disturbances in defects, or indeed, embryonic arrhythmias that in
apparently morphologically normalCCS. extreme casescan lead toembryonic orfoetaldeath.
The Notch signalling cascade was found to be As mentioned above, propensity to arrhythmia
important in regulation of AV conduction in the depends considerably on developmental stage. At the
mouse (Rentschler et al. 2011), and activation of earliest stages, where the heart is small and conduc-
Notch signalling during development consistently led tion generally slow, the only ‘allowed’ arrhythmias
to accessory AV pathways and pre-excitation. On the are alterations of heart rate of which bradycardia is
other hand, inhibition of this cascade led to AV node the most dangerous as it can lead to reduced cardiac
hypoplasia and loss of expression of slowly conduct- output and embryonic death. Once the cardiac cham-
ing connexin 30.2 gap junction channels, resulting in bers are formed (Moorman et al. 2010), alternating
shortened AVdelay. regions of fast and slow conduction develop, creating
a heterogeneity in conduction that can lead to unidi-
rectional or bidirectional blocks, re-entry and more
Drug-induced arrhythmias in mammalian
complex arrhythmias (Valderrabano et al. 2006).
models
With further development, the heart becomes more
A significant worry of every clinician taking care of complexwiththeestablishmentofthecoronaryvascu-
women of childbearing age is the teratogenic potential lar network and autonomic innervation (Hildreth
©2014ScandinavianPhysiologicalSociety.PublishedbyJohnWiley&SonsLtd,doi:10.1111/apha.12418 7
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Foetalcardiacarrhythmias DSedmeraetal. ActaPhysiol2014
et al. 2009). In humans, sensitivity to bradycardia in phenytoin inhibits the inward rectifier potassium cur-
premature infants suggests that the heart rate response rent. The authors attributed these effects to reactive
to cholinergic stimulation may change during develop- oxygen species (ROS) generated at reoxygenation
ment (Maurer 1979). This hypothesis was tested on after resumption of normal rhythm, as an antioxidant
isolated intact foetal mouse hearts (ED13–ED22). agent alpha-phenyl-N-tert-butyl-nitrone showed pro-
Acetylcholine led to a marked ((cid:3)50%) heart rate tective effects. A similar mechanism was proposed for
decrease in the micromolar range in ED13–ED14 another teratogenic anti-epileptic drug, trimethadione
hearts, but the decrease was progressively blunted anditspharmacologicallyactivemetabolitedimethadi-
with increasing age with a mere 3% drop at ED21– one.Thesamegroupofinvestigatorsthenfollowedup
ED22 with the same dose. Physostigmine significantly by showing that these effects were exacerbated in
enhanced the cholinergic response in older hearts, sug- combination with several anti-epileptics (phenytoin,
gesting that the effect is at least in part due to increas- phenobarbital,dimethadioneandcarbamazepine),sup-
ing intrinsic cholinesterase activity with gestational porting the idea that the increased risk for malforma-
age. tions following polytherapy is linked to an increased
Unlike the adult heart, whose energy needs are risk for cardiac rhythm disturbance (Danielsson et al.
mostly met by fatty acid oxidation, the developing 2007).
myocardium relies mostly on glycolysis. Chen et al. Almokalant, a class III anti-arrhythmic drug, caused
(2007) investigated how inhibition of glycolysis affects embryotoxicity in the mouse (Skold & Danielsson
membrane voltage and calcium transients in embry- 2000), most likely secondary to its adverse effects on
onic mouse hearts. Glycolysis inhibition by 10 mM the embryonic heart, as dose-dependent bradycardia
2-deoxyglucose or 0.1 mM iodoacetate decreased sig- and periods of cardiac arrest were observed in whole
nificantly heart rate and induced (unspecified) arrhyth- embryo culture at ED10. Thus, all drugs capable of
mias in over 50% of the treated hearts. Similar effects causing embryonic bradycardia should be regarded as
were noted when oxidative phosphorylation was potentially embryotoxic and used during pregnancy
blocked by 500 nM p-(trifluoromethoxy)phenylhydraz- with extreme caution.
one. During experiments aimed at elucidation of pace-
making mechanisms in early mouse heart (Chen et al.
Chick embryonic model
2010), the investigators observed various arrhythmias,
including AV re-entry induced by adenosine (ADO). The cardiac electrical activity in the chick embryo has
This occurred at stages that had already differentiated been investigated in pioneering works, in vivo
fast-conducting atrial and ventricular chamber myo- (Van Mierop 1967, Rajala et al. 1984, Tazawa et al.
cardiumand slowlyconducting AVcanal. 1989, Sugiyama et al. 1996), in the intact embryo
Anti-epileptic drugs frequently act on ion channels (Hoff & Kramer 1939, Paff et al. 1964), the dissected
regulating membrane potential in excitable tissues; heart (Paff et al. 1968, Paff & Boucek 1975, Kasuya
someofthesechannelsarepresentalsointhedevelop- et al. 1977, Hirota et al. 1987), isolated cardiac
ingheart.Thiscouldbeoneexplanationfortheknown chambers (Boucek et al. 1959, Arguello et al. 1986)
teratogenic potential of these substances. Danielsson and in cultured cardiomyocytes (Shrier & Clay 1982).
et al. (1997) investigated the capacity of phenytoin, a In particular, ECG of the whole heart displays
hERGchannelblockerinhibitingtheIKrthatiscritical characteristic P, QRS and T components which allows
for embryonic heart function, to induce embryonic assessment of the beating rate from PP or RR interval,
hypoxia via adverse effects on the embryonic heart AV conduction from PR interval, duration of the ven-
using a whole embryo culture model. In these mouse tricular activation from QT interval and intraventricu-
embryo studies, phenytoin caused a concentration- lar conduction from of the QRS complex width. The
dependent decrease in embryonic heart rate, with spatio-temporal interpretation of the ECG is facili-
temporary or permanent cardiac arrest at the highest tated by the fact that ventricular activation occurs in
dosage. The exact concentration, as well as incidence a ‘base-to-apex’ fashion, and there is no differentiated
of other arrhythmias, was strain-dependent. Similar conduction system at early developmental stages
resultswereobtainedinratembryos. (Chuck et al. 1997,Reckova et al.2003).
Arrhythmogenicpropertiesofphenytoinwereexam- The principal types of arrhythmias observed in the
ined in mouse (Azarbayjani & Danielsson 2002). validated 4-day-old embryonic chick heart model
Between ED9 and ED13, a dose-dependent bradycar- under various stresses (e.g. anoxia–reoxygenation) or
dia and other unspecified arrhythmias such as AV exposed to pharmacological agents are transient atrial
block were observed at maternal plasma concentra- tachycardia (range 180–300 bpm) and bradycardia
tions in the micromolar range. Patch-clamp recording (range 110–140 bpm), atrial ectopy, first-degree atrio-
on HERG-transfected cells demonstrated that ventricular blocks (AVB), second-degree AVB (2 : 1 to
8 ©2014ScandinavianPhysiologicalSociety.PublishedbyJohnWiley&SonsLtd,doi:10.1111/apha.12418
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ActaPhysiol2014 DSedmeraetal. Foetalcardiacarrhythmias
8 : 1), Wenckebach phenomenon (Mobitz type I), collaborators described heart blocks after digoxin
third-degree AVB (ventricular escape beats) and bursts treatment (Paff et al. 1964), defining the stage of
of irregular activity followed by intermittent atrial chamber formation as critical for induction of conal
arrest (cardioplegia) as previously documented (Sarre (40 h of incubation) and AV block (42 h). Before
et al. 2006) and presented in Figure 3. Some of these these stages, the only reaction of the embryonic heart
arrhythmias resemble those observed in the human to drug treatment was complete cardiac arrest. The
foetus (Strasburger& Wakai2010). authors noted similarity between this AV block and
Effects of drugs inducing arrhythmias can be conve- the situation in humans, including the Wenckebach
niently studied in the chick embryo in vivo. Paff and phenomenon. The Rochester group studied effects of
(a)
(b)
(c)
(d)
(e)
(f)
Figure3 Examplesofmajortypesofar-
rhythmiasobservedinthe4-day-old
embryonicchickheart.ECGoftheiso-
latedheartdisplayscharacteristicP
wave,QRScomplexandTwavecompo-
nents.(a)Atrialectopy,(b)2:1AVB,
(c)atrialectopy+3:1AVB,(d)ven- (g)
tricularescapebeats(third-degreeAVB),
(e)Wenckebachphenomenon(Mobitz
typeI),(f)episodeofheart
block+Wenckebach,(g)third-degree (h)
AVB+burstingactivityand(h)intermit-
tentsinoatrialarrest(cardiople-
gia)+burstingactivity.AVB,
atrioventricularblock.Asteriskindicates
atrialprematurebeat.
©2014ScandinavianPhysiologicalSociety.PublishedbyJohnWiley&SonsLtd,doi:10.1111/apha.12418 9
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Foetalcardiacarrhythmias DSedmeraetal. ActaPhysiol2014
various cardiac drugs on the developing cardiovascu- mechanical delay (reflecting excitation-contraction
lar system. Isoproterenol at a teratogenic dose induced (EC)coupling)significantlyincrease,whereasatrialEC
increased vascular resistance and reduced cardiac coupling remains unchanged. Ventricular contractility
output (Clark et al. 1985), suggesting the presence at the apex and intraventricular conduction are also
of a functional adrenergic signalling system in the significantlyreducedbyanoxia–reoxygenation(Fig. 4),
4-day-old embryonic heart. On the other hand, butnofibrillations,nore-entryandnoventricularecto-
chronic verapamil (calcium antagonist) infusion pic beats are observed. At reoxygenation, arrhythmias
decreasedbothcardiacandembryonicgrowththrough and conduction disturbances are associated with a
decreased cardiac performance (Clark et al. 1991, burst of ROS production (Sarre et al. 2005, Raddatz
Sedmera et al. 1998) and led to delayed ventricular et al. 2011) and reduced by the antioxidant ascorbic
morphogenesis (increased trabeculation, decreased acid (Fig. 5). Although the presence of glucose at the
compact layer thickness). Recently, Kockova and physiological concentration of 8 mM prolongs cardiac
colleagues studied the effects of beta blockers and activity during anoxia, it enhances the reoxygenation-
ivabradine on cardiacfunction andembryonic survival induced ROS production and arrhythmias relative to
(Kockova et al. 2013). High doses led to mortality glucose-free conditions (Tran et al. 1996, Raddatz
through decreased cardiac output, based upon brady- et al. 2011). This observation underscores the role
cardia and insufficient Frank-Starling compensation. that alterations of glycolytic activity may play in ar-
Partial AV blocks were also observed in both early rhythmogenesis associated with ROS. Nitric oxide
(day4) andlater (day8)embryos. (NO) at supraphysiological concentration delays post-
anoxic recovery of AV propagation, and sinoatrial
Arrhythmias during anoxia–reoxygenation pacemaker cells are less responsive to NO (Terrand
et al.2003).AnNOsynthaseinhibitor(L-NAME)pro-
Inthe4-day-oldembryonicchickheartmodel(Raddatz longs the ventricular electromechanical delay during
et al. 1997, Sarre et al. 2006), the chrono-, dromo- anoxia and delays its recovery during reoxygenation,
and inotropic disturbances and the ultrastructural while an NO donor (DETA-NONOate) has opposite
modifications(e.g.mitochondrialandnuclearswelling) effects (Maury et al. 2004). Thus, a NO-dependent
induced by 30-min anoxia followed by 60-min reoxy- pathwayappearstocontributetoregulationofventric-
genationarereversiblewithinaperiodoftimedepend- ular excitation–contraction coupling in the anoxic–
ing on the developmental stage; the older the embryo, reoxygenatedembryonicheart.
the lower the reversibility (Sedmera et al. 2002). It should also be mentioned that a cycle of cooling
Anoxia induces bradycardia, atrial ectopy, first-, sec- (4 °C, 30 min)/rewarming (37 °C, 60 min) under
ond-,andthird-degreeAVBandtransientcardioplegia. normoxia is less arrhythmogenic than anoxia (30 min)
Reoxygenation provokes also the Wenckebach phe- followed by reoxygenation (60 min). However,
nomenon and ventricular escape beats. At the onset of between 15 and 20 min of rewarming, when tempera-
reoxygenation, PR, QT and ventricular electro- ture rises from 27 to 31 °C, the beating rate
Figure4 Ina4-day-oldchickembry-
onicheart,heartrate,atrioventricular
(AV)propagation(PRinterval),QT
duration,atrialandventricularexcita-
tion-contraction(EC)coupling,contrac-
tility(apicalventricularshortening)and
intraventricularconduction(QRSwidth)
aremarkedlyalteredduringanoxiaand
reoxygenation,butfullyrecoverafter
30–40min.Mean(cid:1)SEM;n=4;
n=20forQRSdetermination;bpm,
beatsperminute.*P<0.05vs.prea-
noxicvalues.
10 ©2014ScandinavianPhysiologicalSociety.PublishedbyJohnWiley&SonsLtd,doi:10.1111/apha.12418