© 2000 American Heart Association, Inc.
Integrative Physiology |
Abnormal Cardiac Conduction and Morphogenesis in Connexin40 and Connexin43 Double-Deficient Mice
From the Universität Bonn (S.K., E.T., O.K., K.W.), Institut für Genetik, Abt. Molekulargenetik, Bonn, Germany; Department of Anatomy and Embryology (J.-S.K., W.H.L.), University of Amsterdam, Amsterdam, the Netherlands; and Universität Leipzig (A.H.), Medizinische Klinik I, Leipzig, Germany.
Correspondence to Dr Klaus Willecke, Institut für Genetik, Abt Molekulargenetik, Römerstr 164, 53117 Bonn, Germany. E-mail genetik{at}uni-bonn.de
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Abstract |
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Abstract—Connexin40-deficient (Cx40-/-/Cx43+/+) and connexin43-heterozygous knockout mice (Cx40+/+/Cx43+/-) are viable but show cardiac conduction abnormalities. The ECGs of adult double heterozygous animals (Cx40+/-/Cx43+/-) suggest additive effects of Cx40 and Cx43 haploinsufficiency on ventricular, but not on atrial, conduction. We also observed additive effects of both connexins on cardiac morphogenesis. Approximately half of the Cx40-/-/Cx43+/+ embryos died during the septation period, and an additional 16% died after birth. The majority of the latter mice had cardiac hypertrophy in conjunction with common atrioventricular junction or a ventricular septal defect. All Cx40-/-/Cx43+/- progeny exhibited cardiac malformations and died neonatally. The most frequent defect was common atrioventricular junction with abnormal atrioventricular connection, which was more severe than that seen in Cx40-/-/Cx43+/+ mice. Furthermore, muscular ventricular septal defects, premature closure of the ductus arteriosus, and subcutaneous edema were noticed in these embryos. Cx40+/-/Cx43-/- embryos showed the same phenotype (ie, obstructed right ventricular outflow tract) as reported for Cx40+/+/Cx43-/- mice. These findings demonstrate that Cx43 haploinsufficiency aggravates the abnormalities observed in the Cx40-/- phenotype, whereas Cx40 haploinsufficiency does not worsen the Cx43-/- phenotype. We conclude that the gap-junctional proteins Cx40 and Cx43 contribute to morphogenesis of the heart in an isotype-specific manner.
Key Words: mice, transgenic • defects • electrocardiography • myocardium • conduction
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Introduction |
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Gap junctions are intercellular channels that allow the exchange of ions, second-messenger molecules, and other metabolites between adjacent cells. In mammalian cardiomyocytes, gap junctions mediate electrical coupling. To date, 15 different murine connexin isoforms (ie, the subunit proteins of gap junctions) have been described.1 2 3 4 Cx40 and Cx43 proteins and mRNA were detected in vascular smooth muscles, endothelial cells, and a subpopulation of cardiomyocytes in the vertebrate cardiovascular system.5 In the rodent heart, the expression of both connexins is developmentally regulated.6 7 8 9 Cx40 expression is upregulated in a posterior-to-anterior fashion during cardiac development, reaching a maximum at embryonic day (ED) 14. During subsequent development, Cx40 expression becomes restricted to the atria and the ventricular conduction system. Cx43 is also gradually upregulated in both atria and ventricle during early cardiac development but reaches its highest level only at weaning. Cx43 is the main connexin isoform of the ventricular myocardium, but it is absent from the proximal ventricular conduction system in rodents.6 7
Mice heterozygous for Cx43 (Cx40+/+/Cx43+/-) are viable and show, apart from a mild dilatation of the right ventricular chamber in some cases,10 no defects in cardiac morphogenesis, whereas Cx43-deficient (Cx40+/+/Cx43-/-) mice die shortly after birth due to obstruction of the right ventricular outflow tract.11 12 There are conflicting results on the conduction velocity in the myocardium of Cx40+/+/Cx43+/- mice.13 14 Cx40-deficient (Cx40-/-/Cx43+/+) mice are viable and show reduced velocity of impulse conduction in the atria and the ventricular conductive myocardium.15 16 17 Spontaneous as well as inducible dysrhythmias were observed in Cx40-/-/Cx43+/+ mice,18 but until now, no morphological abnormalities were associated with this genotype. In the present study, we observed, however, some Cx40-/-/Cx43+/+ embryos with small septational defects.
We were interested to establish the extent to which Cx40 and Cx43 can complement one another in cardiac morphogenesis and in heart physiology. Thus, we interbred Cx40- and Cx43-mutant mice and characterized the genotypically different progeny through histology and ECG. We observed that haploinsufficiency of Cx40 and of Cx43, although coexpressed only in the distal part of the ventricular conductive myocardium, has additive effects on ventricular conduction in Cx40+/-/Cx43+/- mice and that haploinsufficiency of Cx43 increased the frequency and severity of cardiac malformations in Cx40-/- mice, whereas haploinsufficiency of Cx40 did not affect the Cx43-/- phenotype.
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Materials and Methods |
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Breeding and Preparation of Embryos
Cx40-/-/Cx43+/+ mice were obtained by interbreeding Cx40+/-/Cx43+/+ mice of mixed 129/Sv-C57BL/6 background.15 Cx40-/-/Cx43+/+ mice were mated with Cx40+/+/Cx43+/- mice, which were also of mixed 129/Sv-C57BL/6 background.11 Double heterozygous animals (Cx40+/-/Cx43+/-) were mated with each other or with Cx40-/-/Cx43+/+ mice. The morning on which the presence of the vaginal plug was noticed was considered ED0.5. Embryos were dissected and genotyped with polymerase chain reaction or Southern blot hybridization (Figure 1
) as described
previously.15 19 For histological studies,
the embryos were fixed in a solution of ice-cold methanol/acetone/water
(2:2:1) for 4 hours or overnight. All experiments were carried out in
accordance with the German law on animal protection.
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Histological Analysis
Embryos were embedded in Paraplast plus (Sherwood Medical Co),
serially sectioned at 7-µm thickness, and mounted onto
polylysine-coated glass slides. The sections were stained with
hematoxylin and azophloxine. Correlations between the normal and
abnormal development of cardiac structures were evaluated with the
2 test. A probability value of <0.05 was
considered statistically significant.
ECG Recordings
ECG recordings were performed as previously
described.18 All data obtained were expressed as
mean±SEM. The effect of each connexin was tested with a
multivariate 2-factor ANOVA (factors Cx40 and Cx43,
each with two levels: +/+ and +/-, respectively). Because no
significant interaction was found between the connexins, the
differences between the different genotypes could be tested
with a Student-Newman-Keuls test. Correlations of the effects of
connexins on separate parts of the ECG were determined per
genotype. A probability value of <0.01 was considered
statistically significant.
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Results |
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Embryonic Lethality and Structural Heart Defects in Cx40-Deficient Mice
Cx40-/-/Cx43+/+ mice are viable and fertile, but we found fewer Cx40-/-/Cx43+/+ animals after interbreeding of Cx40+/-/Cx43+/+ mice than anticipated for mendelian inheritance (Table
; compare Reference 15 ).
We observed the expected number of
Cx40-/-/Cx43+/+ embryos
at ED11.5 but only half of the expected number of
Cx40-/-/Cx43+/+ embryos
after ED13.5 (Table). This indicates that Cx40 plays an
important role in the period during which septation of the heart takes
place. We examined morphologically the hearts of five
Cx40-/-/Cx43+/+ mice at
ED12.5 but found them to be structurally normal except for the
mesenchymal cap on the free rim of the primary atrial septum, which was
incompletely formed in two of the hearts (Figure 2A).
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Sixteen percent of the newborn
Cx40-/-/Cx43+/+ mice (17
of 106) died shortly after birth. The surviving animals developed
normally, but some of the
Cx40-/-/Cx43+/+ animals
died as young adults at 4 to 8 weeks of age. When we analyzed
the hearts of three newborn and two young adult mice, four showed
myocardial hypertrophy. Three of these hearts had a small
septum primum defect, considered to be the mildest form of the common
atrioventricular junction (Figure 2B
; see also
Figure 1 online [data supplement available at
http://www.circresaha.org]). In addition, one of them had a persisting
interventricular foramen. Furthermore, two of the three
hearts exhibited a persistent foramen ovale. In the fourth hypertrophic
heart, only a ventricular septal defect was detected
(Figure 2C; see also Figure 2 online [data supplement available
at http://www.circresaha.org]).
Breeding and Electrophysiological
Characterization of Cx40+/-/Cx43+/- Double
Heterozygous Mice
To establish whether Cx40 and Cx43 have additive effects on the
function of the heart, we generated
Cx40+/-/Cx43+/- mice and
analyzed the conduction properties of their hearts through limb
lead surface ECG measurements (Figure 3).
No change in the duration of the RR interval was observed in any of the
genotypes, showing that the function of the sinus node is not
affected by Cx40 or Cx43 heterozygosity. The duration of the P wave was
increased in both
Cx40+/-/Cx43+/+ and
Cx40+/+/Cx43+/- mice,
but the measurements of
Cx40+/-/Cx43+/- showed
the effect was not additive. The duration of the PQ interval was not
significantly different in any of the genotypes. The duration
of the QRS complex was similar in
Cx40+/+/Cx43+/+,
Cx40+/-/Cx43+/+, and
Cx40+/+/Cx43+/- animals
but significantly increased in double heterozygotes, showing that the
small increases in the duration of the QRS complex due to Cx40 and Cx43
heterozygosity add up to a significant effect. The duration of the
QTmax interval was increased in
Cx40+/-/Cx43+/+ and
Cx40+/+/Cx43+/- animals
relative to Cx40+/+/Cx43+/+
animals. Measurement of the QTmax interval in
Cx40+/-/Cx43+/- animals
showed that these effects were also additive. Thus, decreased amounts
of Cx40 and Cx43 protein in the heart resulted in slowed atrial
depolarization and ventricular depolarization and
repolarization. These effects were additive in the ventricles but not
in the atria. Furthermore, the duration of the P wave always correlated
with the duration of the PQ interval, and the duration of the QRS
complex always correlated with the duration of the
QTmax interval, except in double heterozygotes.
Only in Cx40+/-/Cx43+/+
animals was a correlation observed between atrial and
ventricular parameters, except for the duration
of the P wave and the QS interval.
Cx40+/-/Cx43+/- animals
did not show any correlation among the ECG parameters.
There was no correlation between the cycle length (RR interval) and the
other parameters in any genotypes we investigated
(data not shown). This analysis shows that changes in the
length of the P wave are correlated with those of the PQ interval and
that changes in the duration of the QRS complex are correlated with
those in the QTmax interval in all
genotypes except the double heterozygotes.
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Morphological Abnormalities in Embryonic Hearts of
Cx40-/-/Cx43+/- or
Cx40+/-/Cx43-/- Mice
We interbred
Cx40+/-/Cx43+/- mice to
determine whether Cx40 and Cx43 have additive functions during cardiac
morphogenesis. As anticipated, we did not obtain
Cx43-/- mice after birth (compare Reference
11 ) and found fewer than half of the expected
Cx40-/-/Cx43+/+ mice
(Table). In addition, we did not find
Cx40-/-/Cx43+/- animals
at the age of 3 weeks (0 of 180 instead of 
20 of 180 expected).
Similarly, interbreeding of
Cx40+/-/Cx43+/+ and
Cx40+/-/Cx43+/- animals
yielded no animals of the
Cx40-/-/Cx43+/-
genotype in 124 mice analyzed at 3 weeks of age.
Genotyping of embryos at different developmental stages showed that the
Cx40-/-/Cx43+/- animals
died during the first two postnatal days.
The morphology of the embryonic hearts from 21 Cx40-/-/Cx43+/- mice and 4 Cx40+/-/Cx43-/- mice from ED12.5 to ED18.5 was analyzed histologically.
Cx40-/-/Cx43+/- Mice
The most characteristic defects were seen at the
atrioventricular junction. These malformations were due
to a graded deficiency in normal development, indicating that the
normally occurring growth of the right ventricle, the associated
rightward expansion of the atrioventricular junction,
and septation were inhibited to variable degrees. At ED12.5, the
youngest stage investigated, the two available
Cx40-/-/Cx43+/- embryos
showed unaltered cardiac morphology, except that the mesenchymal cap on
the free rim of the primary atrial septum was absent. At ED13.5, one of
the two embryos analyzed had a persisting foramen primum of the
atrium and missed the mesenchymal cap on the primary atrial septum,
whereas the other was normal. From ED14.5 on, when the
interventricular foramen was closed in the wild type, all
Cx40-/-/Cx43+/-
embryonic hearts (n=17) revealed a persisting
interventricular foramen (Figure 4). Thirteen of these embryos (76%) had,
in addition, a persisting foramen primum, indicating the presence of a
common atrioventricular junction. The common
atrioventricular junction was guarded by a common valve
with a single orifice in 10 of the 13 embryos (77%, Figure 5A) and by two valves in the remaining
three cases (Figure 5B). If only a single orifice was
present, both atrioventricular cushions had not
fused, whereas the presence of two orifices showed that they did but
not with the primary atrial septum. In the other four embryos, the
foramen primum was closed so that right and left
atrioventricular junctions had a myocardial boundary
except at the location of the future membranous septum. The pattern of
the atrioventricular connection ranged from absent
rightward expansion of the atrioventricular connection
(double inlet left ventricle, 3 of 17 cases, 18%, Figure 5A),
via a small right-sided connection (6 cases, 35%, Figure 5B),
to the normally balanced connection (8 cases, 47%). All embryos with
double-inlet left ventricle or small right connection had a common
atrioventricular junction. In all embryos with
double-inlet left ventricle, the atrioventricular valve
was common and the interventricular septum was malaligned
with the primary atrial septum. In these hearts, the right ventricle
communicated with the atria via the interventricular
foramen, and its was smaller than the left ventricle. In five of the
six hearts (83%) with a small right atrioventricular
connection, the atrioventricular junction was guarded
by a common valve, whereas the septa were malaligned in 50% of these
hearts. In these cases, the right ventricle has direct access from the
right atrium but was small compared with the left ventricle. Only four
of the eight cases with balanced connection had a common
atrioventricular junction. In these four hearts, the
valve was either common (50%) or separate. Septal alignment was
normal, and the sizes of both ventricles were comparable. In 10 of 13
hearts with a persisting foramen primum (77%), the primary atrial
septum was not covered on its lower free rim with the mesenchymal cap
(Figure 5A; see also Figure 3 online [data supplement available
at http://www.circresaha.org]) that normally occupies this position.
The absence of this structure was related to the presence of a common
atrioventricular valve (P=0.03) (see Figure 3 online; data supplement available at http://www.circresaha.org), a
lack of development of the right atrioventricular
connection (P=0.03), and septal malalignment
(P=0.01). At later stages, especially at ED18.5, the venous
valves appeared short. This occurred because their attachment to the
base of the heart was more caudal than that of control embryos. This
feature was not related to the absence of a mesenchymal cap on the free
rim of the primary atrial septum (P=0.63), the presence of a
common atrioventricular valve (P=0.63), a
lack of development of the right atrioventricular
connection (P=0.62), or septal malalignment
(P=0.35).
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A defect in the muscular interventricular septum (Figures 5B and 5C) was observed in two of three ED18.5 embryos
analyzed but not in younger embryos. However, some of these
latter embryos showed a loose, random arrangement of the septal
myocardium with deep canals lined by endocardium in the
region where the muscular defect was found at the later stage (Figure 5E).
All of the embryos of the
Cx40-/-/Cx43+/-
genotype displayed the usual atrial arrangement (ie, the
presence of a normal atrial situs). Furthermore, none of them had
defects in the arterial pole of the heart. The great
vessels were also inconspicuous, but surprisingly, the ductus
arteriosus was fully constricted in all ED18.5 embryos (Figure 5D). Most embryos showed subcutaneous edema between ED13.5 and
ED15.5, mainly in the neck and back (Figure 5F). The extent of
edemas had no apparent relation to the severity of cardiac defects, and
it was not found in any of the embryos after ED16.5.
Cx40+/-/Cx43-/- Mice
The four mice of this genotype that were investigated
exhibited similar features as
Cx40+/+/Cx43-/-
mice.12 The ED12.5 embryo showed the typical A-loop: the
right ventricle was located anteromedially relative to the left
ventricle, whereas the outflow tract occupied a leftward position and
turned toward the branchial arches with an acute bend (Figure 6A). Its atrioventricular
canal connected both atria to the left ventricle, and the
atrioventricular cushions were positioned on the left
and right sides of the canal rather than on the superior and
inferior side, respectively (Figure 6B). At ED15.5
and ED17.5, the
Cx40+/-/Cx43-/- hearts
showed the typical features of
Cx40+/+/Cx43-/- hearts at
the junction of the right ventricle and the outflow tract. At this
junction, several interconnected or blind pouches separated by thick
trabeculae were present (Figures 6C and 6D).
None of the ED15.5 or ED17.5 embryos had septal defects.
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Discussion |
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The Cx40 and Cx43 connexin isoforms are both expressed in the heart, showing a distinct and at some sites overlapping expression pattern. In the electrophysiological portion of the present study, we did observe additive effects of Cx40 and Cx43 heterozygosity on ventricular, but not atrial, conduction. Furthermore, cardiac morphogenesis was more severely impaired in Cx40-/-/Cx43+/- animals than in Cx40-/-/Cx43+/+ animals. However, the finding that Cx43 haploinsufficiency worsened the morphological phenotype of Cx40 deficiency, whereas Cx40 haploinsufficiency did not seem to alter the morphological phenotype of Cx43 deficiency, showed that Cx40 and Cx43 functions in cardiac morphogenesis were not mutually exchangeable.
Decreased Expression of Cx43 and Cx40 Genes in
Cx40+/-/Cx43+/- Mice Results in Decreased
Atrial and Ventricular Conduction
As previously shown,15 18 ECG parameters
in Cx40 heterozygous mice were only mildly affected. In fact, we
observed only a 10% increase in the duration of the P wave and the
QTmax interval in
Cx40+/-/Cx43+/+ mice
compared with wild-type control controls. However, substantial
increases in the duration of the P wave (>50%) and PQ, QS, and QT
intervals (20%) are present in
Cx40-/-/Cx43+/+
mice.18 In
Cx40+/+/Cx43+/- mice, the
durations of the P wave and the QTmax interval
were significantly increased compared with those of wild-type control
animals, but the PQ interval and the duration of the QRS complex were
not. Our data on ventricular conduction are similar to
those reported by Morley et al.14 However, as a result of
the smaller number of animals, increases in the QT interval did not
reach significance in the study by Morley et al.14
Ventricular depolarization was not significantly slowed in
Cx40+/+/Cx43+/- mice,
although different findings have been reported.13 Our
findings support the contention that the Cx43 heterozygous mice do not
experience major changes in intraventricular
conduction velocity. Ventricular depolarization and
repolarization are significantly slower in
Cx40+/-/Cx43+/- mice
compared with either
Cx40+/-/Cx43+/+,
Cx40+/+/Cx43+/-, or
wild-type mice. Because Cx40 and Cx43 are coexpressed only in the
distal conductive myocardium of the ventricles, these
findings were probably due to the additive effects of slightly lower
conduction velocities in the ventricular conduction system
(where Cx40 is expressed8 ) and in the working
myocardium of the ventricles (where Cx43 is
expressed6 7 ). Conceptually, it could be regarded as a
result of two increasing serial resistances. In mouse atria, Cx40 and
Cx43 are coexpressed.6 7 8 Heterozygosity for either Cx40
or Cx43 has an effect on the duration of the P wave, but intriguingly,
these effects were not additive. Such findings can in principle be
explained by a model in which the gene dosages of Cx40 and Cx43 are
responsible for different resistors in parallel.
Under anesthesia with avertin, correlation of the cycle length with any other ECG parameters was not detected in wild-type, Cx40+/-/Cx43+/+, Cx40+/+/Cx43+/-, or Cx40+/-/Cx43+/- animals. However, in wild-type, Cx40+/-/Cx43+/+, or Cx40+/+/Cx43+/- mice, the duration of the P wave correlated with the PQ interval and the duration of the QRS complex correlated with the QTmax interval. This suggests that adaptational changes in atrioventricular conduction velocities and in the time necessary for repolarization correlate with changes in atrial and ventricular conduction velocities, respectively. Cx40+/-/Cx43+/+ and Cx40+/+/Cx43+/- mice did not differ in this respect from wild-type animals. Interestingly, these correlations are both lost in Cx40+/-/Cx43+/- mice. This finding suggests that the coordination of atrial and atrioventricular conduction, on the one hand, and that of ventricular conduction and repolarization, on the other hand, are influenced by gap-junctional communication.
Cx43 Haploinsufficiency Aggravates the Cardiac Morphological
Phenotype in Cx40-Deficient Mice
Cx40-/-/Cx43+/+ mice
showed an increased incidence of prenatal death between ED11.5 and
ED13.5. Because we did not observe overt morphological defects in the
hearts of these embryos, the cause of embryonic death in
Cx40-/-/Cx43+/+ mice is
probably functional rather than structural. The expression of the
connexin genes in the embryonic heart is developmentally
regulated.8 9 In the developing heart, Cx40 is upregulated
in a posteroanterior manner. Cx40 mRNA is first detected in the
embryonic atrium and left ventricle at ED9.5 and, after ED11.5, also in
the embryonic right ventricle. Cx43 is expressed early in the
ventricles but not before ED12.5 in the atria.20 Because
Cx45 is expressed between ED8.5 and ED10.5 and is downregulated
thereafter,21 there may be a temporary lack of
gap-junctional communication in the hearts of some Cx40-deficient
embryos, resulting in embryonic death.
Another "window of sensitivity" is around birth, when 16% of the Cx40-/-/Cx43+/+ animals and all of the Cx40-/-/Cx43+/- animals die. The septational defects might hinder the efficient establishment of the pulmonary circulation at birth (ie, the animals cannot cope with changes in cardiac workload and finally die). The importance of Cx40 for the mechanical function of the heart is underscored by the myocardial hypertrophy that was found in the majority of Cx40-/-/Cx43+/+ animals that died postnatally.
Some Cx40-deficient mice showed a mild form of common atrioventricular junction or ventricular septal defects, indicating that the septational process is sensitive to Cx40 deficiency. The cardiac malformations found in Cx40-/-/Cx43+/- mice are more severe forms of the defects observed in Cx40-/-/Cx43+/+ animals. This group of cardiac malformations appears to arise as a result of a developmental arrest between ED12 (onset of cardiac septation) and ED14 (completion of cardiac septation). It is conceivable that the fusion defects developed as secondary effects due to a disturbed contractile pattern in the Cx40-/-/Cx43+/- hearts (eg, altered shear stress). However, if that were the case, it would be difficult to explain why Cx40+/-/Cx43-/- animals did not show similar septation defects but instead showed only abnormalities in the downstream portion of the heart.11 12 Alternatively, the morphological changes could be directly due to decreased intercellular gap-junctional communication.
The process of atrioventricular septation is impaired in Cx40-deficient embryos, and this impairment is worsened by haploinsufficiency for Cx43. Atrioventricular septation involves, in addition to the myocardium of the atrioventricular canal, several developmental primordia, including the endocardial cushions, the mesenchymal rim on the free edge of the primary atrial septum, the right pulmonary ridge, and the valves of the sinus venosus. The only other genetic model that is associated with defects of the atrioventricular junction is trisomy 16.22 23 Webb et al22 hypothesized that a failing development of the spina vestibuli underlies the maldevelopment of the atrioventricular junction in these mice. In agreement with this hypothesis, we found in Cx40-/-/Cx43+/- embryos that the absence of a mesenchymal rim on the free edge of the primary atrial septum was associated with the most pronounced abnormalities of the atrioventricular connection.
Cx40- and Cx43-mediated gap-junctional communication appears to be
crucial for the complex morphogenetic process of
atrioventricular septation. Nevertheless, we observed a
wide spectrum in the extent to which development of the
atrioventricular canal was affected. Some animals, for
example, suffered only from a ventricular septal defect
(compare Figure 4). The development of such a spectrum of
effects would be expected, if septational defects in the
atrioventricular canal of Cx40-deficient embryos arise
as a result of a disturbed heart function (see earlier). On the other
hand, small differences in the genetic background could also modulate
the phenotypic appearance of individual animals.
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Acknowledgments |
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This work was supported by grants from the German Research Association (SFB 284, C1), the BioMed Program of the European Community, and the Fonds der Chemischen Industrie to Dr Willecke. We are grateful to Drs Robert H. Anderson (London) and Diego Franco (Amsterdam) for helpful discussion and critical reading of the manuscript.
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Footnotes |
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1 Both authors contributed equally to this study.
Received January 31, 2000; revision received July 6, 2000; accepted July 7, 2000.
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References |
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1. Goodenough A, Goliger JA, Paul DL. Connexins, connexons and intercellular communication. Annu Rev Biochem. 1996;65:475–502.[Medline] [Order article via Infotrieve]
2. Condorelli DF, Parenti R, Spinella F, Salinaro AT, Belluardo N, Cardile V, Cicirata F. Cloning of a new gap junction gene (Cx36) highly expressed in mammalian brain neurons. Eur J Neurosci. 1998;10:1202–1208.[Medline] [Order article via Infotrieve]
3. Söhl G, Degen J, Teubner B, Willecke K. The murine gap junction gene connexin36 is highly expressed in mouse retina and regulated during brain development. FEBS Lett. 1998;428:27–31.[Medline] [Order article via Infotrieve]
4.
Manthey D, Bukauskas F, Lee CG, Kozak CA, Willecke K.
Molecular cloning and functional expression of the mouse gap junction
gene connexin-57 in human HeLa cells. J Biol Chem. 1999;274:14716–14723.
5.
Little TL, Beyer EC, Duling BR. Connexin 43 and
connexin 40 gap junctional proteins are present in arteriolar
smooth muscle and endothelium in vivo. Am J
Physiol. 1995;268:H729–H739.
6.
Van Kempen MJ, Fromaget C, Gros D, Moorman AF, Lamers
WH. Spatial distribution of connexin43, the major cardiac gap junction
protein, in the developing and adult rat heart. Circ Res. 1991;68:1638–1651.
7. Gourdie RG, Green CR, Severs NJ, Thompson RP. Immunolabelling patterns of gap junction connexins in the developing and mature heart. Anat Embryol. 1992;185:363–378.[Medline] [Order article via Infotrieve]
8. Delorme B, Dahl E, Jarry-Guichard T, Marics I, Briand JP, Willecke K, Gros D, Théveniau-Ruissy M. Developmental regulation of connexin40 gene expression in mouse heart correlates with the differentiation of the conduction system. Dev Dyn. 1995;204:358–371.[Medline] [Order article via Infotrieve]
9.
Delorme B, Dahl E, Jarry-Guichard T, Briand J-P,
Willecke K, Gros D, Théveniau-Ruissy M. Expression pattern of
connexin gene products at the early developmental stages of the
mouse cardiovascular system. Circ Res. 1997;81:423–437.
10. Huang GY, Wessels A, Smith BR, Linask KK, Ewart JL, Lo CW. Alteration in connexin 43 gap junction gene dosage impairs conotrucal heart development. Dev Biol. 1998;198:32–44.[Medline] [Order article via Infotrieve]
11.
Reaume, AG, de Sousa PA, Kulkarni S, Langille BL, Zhu
D, Davies TC, Juneja SC, Kidder GM, Rossant J. Cardiac
malformation in neonatal mice lacking connexin43. Science. 1995;267:1831–1834.
12.
Ya J, Erdtsieck-Ernste EB, de Boer PA, van Kempen MJ,
Jongsma H, Gros D, Moorman AFM, Lamers WH. Heart defects in
connexin43-deficient mice. Circ Res. 1998;82:360–366.
13.
Thomas SA, Schuessler RB, Berul CI, Beardslee MA,
Beyer EC, Mendelsohn ME, and Saffitz JE. Disparate effects of
deficient expression of connexin43 on atrial and
ventricular conduction. Circulation. 1998;97:686–691.
14. Morley GE, Vaidya D, Samie FH, Lo C, Delmar M, Jalife J. Characterization of conduction in the ventricles of normal and heterozygous Cx43 knockout mice using optical mapping. J Cardiovasc Electrophysiol. 1999;10:1361–1375.[Medline] [Order article via Infotrieve]
15. Kirchhoff S, Nelles E, Hagendorff A, Krüger O, Traub O, Willecke K. Reduced cardiac conduction velocity and predisposition to arrhythmias in connexin40-deficient mice. Curr Biol. 1998;8:299–302.[Medline] [Order article via Infotrieve]
16. Simon AM, Goodenough DA, Paul DL. Mice lacking connexin40 have cardiac conduction abnormalities characteristics of atrioventricular block and bundle branch block. Curr Biol. 1998;8:295–298.[Medline] [Order article via Infotrieve]
17. Verheule S, van Batenburg CA, Coenjaerts FE, Kirchhoff S, Willecke K, Jongsma HJ. Cardiac conduction abnormalities in mice lacking the gap junction protein connexin40. J Cardiovasc Electrophysiol. 1999;10:1380–1389.[Medline] [Order article via Infotrieve]
18.
Hagendorff A, Schumacher B, Kirchhoff S, Lüderitz
B, Willecke K. Conduction disturbances and increased atrial
vulnerability in connexin40 deficient mice analyzed by
transesophageal stimulation. Circulation. 1999;99:1508–1515.
19. Houghton FD, Thönnissen E, Kidder GM, Naus CCG, Willecke K, Winterhager E. Doubly mutant mice, deficient in connexin32 and -43, show normal prenatal development of organs where the two gap junction proteins are expressed in the same cells. Dev Genet. 1999;24:5–12.[Medline] [Order article via Infotrieve]
20. Yancey SB, Biswal S, Revel J-P. Spatial and temporal patterns of distribution of the gap junction protein connexin43 during mouse gastrulation and organogenesis. Development. 1992;114:203–212.[Abstract]
21.
Alcoléa S, Théveniau -Ruissy M,
Jarry-Guichard Th, Marics I, Tzouanacou E, Chauvin JP, Briand J-P,
Moorman AFM, Lamers WH, Gros DB. Down regulation of connexin45 gene
products during mouse development. Circ Res. 1999;84:1365–1379.
22.
Webb S, Anderson RH, Lamers WH, Brown NA. Mechanism of
deficient septation in the mouse with trisomy 16. Circ Res. 1999;84:897–905.
23.
Webb S, Brown NA, Anderson RH. Cardiac morphology at
late fetal stages in the mouse with trisomy 16: consequences for
different formation of the atrioventricular junction
when compared to humans with trisomy 21. Cardiovasc Res. 1997;34:515–524.
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