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or ß1
Division of Endocrinology and Metabolism (B.G., S.U.T., W.F.B., E.A.S., W.H.D.), University of California, San Diego, La Jolla, California 92093; Departments of Physiology and Biophysics and Medicine (R.C., R.W., K.M.J., W.G.), University of Calgary, School of Medicine, Calgary, Alberta T2N 4N1, Canada; and Laboratoire de Biologie Moleculaire et Cellulaire (O.C., J.S.), Centre Nationale de la Recherche Scientifique, Ecole Normale Superieure de Lyon, 69364 Lyon, France
Address all correspondence and requests for reprints to: Wolfgang H. Dillmann, Division of Endocrinology and Metabolism, University of California, San Diego, 9500 Gilman Drive, La Jolla, California 92093-0618. E-mail: wdillman{at}ucsd.edu
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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and T3Rß. However,
which isoform contributes to specific, T3-induced
alterations of cardiac function remains unclear. Here, we used
individual T3R isoform knockout (KO) mice to study the
effects of T3R and T3Rß in the heart. Our
findings indicate that potassium channel genes that code for
K+ channels involved in action potential repolarization,
like KV 4.2 and minK, are T3R targets. Both are markedly
regulated by thyroid status. The recently identified cyclic
nucleotide-gated channels, HCN2 and HCN4, are targets of
T3R and are unchanged in a euthyroid T3Rß
KO. However, these transcripts respond markedly to altered
T3 signaling concomitant with bradycardia in
T3R KO and hypothyroid animals, as well as tachycardia
in hyperthyroid T3Rß KO mice. SERCA2a and myosins are
T3 regulated and were also targets of T3R,
and the papillary muscles of KO animals showed a slowed rate of
force development. Because of the absence of significant cardiac
effects in euthyroid T3Rß KO mice, we determined
messenger RNA levels for both T3R and
T3Rß in the heart. We found that T3Rß is
present at a 1:3 ratio to T3R1. We conclude that
the cardiac phenotype regulated by T3 is predominantly
mediated by T3R and that the lack of T3R
cannot be compensated by T3Rß in the heart. |
Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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gene and increases expression of the MHC ß gene (4, 5). These changes result in a decrease in myosin V1 and an
increase in the amount of myosin V3 (which has a lower ATPase activity)
and a decreased speed of systolic contraction (6, 7). The
molecular basis of alterations in electrophysiological function caused
by changes in thyroid hormone status has not been completely explored.
However, some specific alterations in ion channel expression or
function have been identified. For example, hypothyroidism has been
shown to decrease the messenger RNA (mRNA)’s coding for KV 1.5, KV
4.2, and KV 4.3 in rats (8, 9, 10). The calcium-independent
transient outward potassium current Ito is
carried primarily by the K+ channel KV 4.2 and KV
4.3 in adult rats; and in hypothyroid animals, the transient outward
current Ito is diminished (11). Due
to the complexity of ion channel expression in different species and
region-specific heterogeneity of ion channel expression in the heart,
no complete picture has emerged of the thyroid hormone influences on
different currents or the genes encoding the respective -subunits.
Furthermore, the molecular basis of the well-known alterations in
chronotropic function leading to thyroid hormone-induced alterations in
heart rate have not been studied in detail. Recently, two genes coding
for the hyperpolarization-activated current If,
which contributes to pacemaker activity, have been cloned (12, 13). They are termed HCN2 and HCN4 for
hyperpolarization-activated, cyclic nucleotide-gated current genes 2 or
4. The HCN2 gene product encodes channels responsible for the fast
component of the If current, and the HCN4 gene
encodes channels responsible for the slow component (14).
This report describes, for the first time, changes induced in the HCN2
and HCN4 gene expression by alterations in T3
receptor (T3R) isoform expression in the mouse
heart.
Thyroid hormone action in the heart is largely mediated by the binding
of T3 to two nuclear thyroid hormone receptors,
T3 receptor 1 (T3R1)
and T3 receptor ß1
(T3Rß1), which are encoded by two separate
genes (15). The T3R locus also
encodes a separate splice variant, the T3R2
gene, which does not bind T3 and has a weak
dominant negative effect (16). From the
T3Rß locus, a separate splice variant, the
T3Rß2 gene, in addition to the
T3Rß1 gene, is expressed. Immunohistochemistry
revealed T3Rß in most tissues; however, in
heart and muscle, the T3Rß isoform is expressed
at low levels (17). The T3Rß2
isoform is predominantly expressed in the pituitary and the central
nervous system, but some reports have identified
T3Rß2 receptor protein in the heart
(18). Until recently, it has been unclear whether
T3 action in the heart can be mediated by either
T3 binding to the T3R1
or the T3Rß1 receptor or whether certain
distinct effects are exclusively mediated by T3
binding to either T3R1 or
T3Rß1. Recently published reports indicate that
mice with deletion of T3R1 (i.e.
only expressing the T3Rß gene) have bradycardia
(19, 20). In this study, we have used a different line of
T3R knockout (KO) mice with complete
elimination of the T3R gene products,
T3R1 and T3R2,
generated by deletion of exon 2 from the T3R
locus (21). Our results show that, in these mice,
bradycardia also occurs, and this is linked to decreased levels of HCN2
and HCN4 gene expression. In addition, T3R KO
mice, in contrast to T3Rß KO mice, have
decreased contractile function. Although further details of
T3R vs.
T3Rß function need to be elucidated, it seems
that T3 action in the heart is dominated by
binding of T3 to the
T3R1 receptor. Identifying specific
T3R- and T3Rß-mediated
effects in the heart may provide a rational basis for the generation of
novel thyroid hormone analogues with preferred binding to only one
receptor isoform, resulting in selective alterations of specific
T3 effects in the cardiovascular system.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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T3R KO animals were generated in the
laboratory of Jacques Samarut, by either deleting most of exon 2 or
exons 5, 6, and 7 (including introns 5, 6, and 7) of the
T3R gene (21, 23) Both lines of
T3R KO animals showed the same changes in ion
channel expression. T3Rß KO animals were also
generated in the laboratory of Jacques Samarut, by deletion of exons 4
and 5 of the T3Rß gene (24).
RNA extraction and ribonuclease (RNase) protection
analysis
Hearts from control animals and experimentally altered mice were
dissected after deep anesthesia, and the atrium was removed, before the
heart was frozen in liquid nitrogen. Isolation of tissue RNA was
performed as described by Chomczynski (25).
Ten micrograms of total RNA were used for hybridization with specific
probes that were generated by PCR from mouse complementary DNAs (cDNAs)
of KV 4.2, minK, and calsequestrin. The probe for the HCN2 channel was
generated by PCR from mouse genomic DNA with primers internal to the
second exon of the gene. The positions of the primers in the published
mouse cDNA are: 629–649 for the sense primer, and 833–854 for the
antisense primer. The probe for the HCN4 channel was derived from a
plasmid containing a partial cDNA from the mouse HCN4 clone (a kind
gift from Bina Santoro, Columbia University, NY). A PstI/Aat
II fragment, spanning positions 770–938 in the published sequence, was
subcloned into pBKS II-. To generate probes for
T3R1, T3R2, and
T3Rß, PCR primer pairs were designed that could
be used with mouse genomic DNA as a template. The
T3R1/2 probe hybridizes in exon 9 from cDNA
position 992-1133, which yields protected fragments of 141
nucleotides (nt) for 1 and 122 nt for 2 in length. The
T3Rß probe hybridizes in exon 4 from cDNA
position 605–790 that yields a protected fragment of 185 nt in
length.
Hybridization conditions, generation of radiolabeled antisense RNA transcripts, and processing of the RNA/RNA hybrids were essentially as described in Current Protocols (26). Usually 2–3 ion channel probes and a control transcript, calsequestrin, were analyzed in one hybridization reaction. The protected fragments were separated on 6% acrylamide sequencing gels. Densitometry of the autoradiographs yielded digital values for each band, which were normalized to the calsequestrin signal. Data from at least three independent experiments were statistically evaluated to give the percent increases or decreases of the message levels for these ion channels.
Isolated papillary muscle experiments
Contractile parameters of papillary muscles were measured as
previously described (27). Briefly, left ventricular
papillary muscles, from the hearts of six T3R
KO mice and four wild-type mice or four T3Rß KO
mice and four wild-type mice, were excised under oxygenated Tyrode
solution (in mM: 136 NaCl, 5.4 KCl, 1
MgCl2, 0.33
NaH2PO4, 10 HEPES, and 10
glucose, pH 7.40) containing 30 mM 2,3-butanedione monoxime
and 2.5 mM CaCl2. They were inserted
into 
-shaped clamps made from strips of platinum foil, tied with 6.0
braided silk suture, and mounted on hooks of platinum wire in a 0.5-ml
muscle chamber.
Muscles were perfused with 2.5 mM Ca2+ Tyrode solution at 37 C and stimulated at 2 Hz and 6 Hz through the platinum clamps (5 V, 0.25-msec duration). Force was measured with an isometric force transducer (OPT1L, Scientific Instruments, Heidelberg, Germany) and recorded on a strip chart recorder. Muscles were stretched, over 30–60 min, to the length at which active force development was maximal. Forces (in mN) were normalized by the muscle cross-sectional areas, to yield stresses (in mN/mm2). The cross-sectional area was calculated, for each muscle, as the ratio of muscle volume (determined by weighing) and muscle length at the length at which active force development was maximal.
The time to peak tension was determined as the time from 10% of tension development to the peak of contraction. Relaxation time was determined as the time from the peak of contraction to 50% of maximum developed stress during relaxation. Data are expressed as mean ± SEM. Statistical comparisons were made by unpaired Student’s t tests.
Electrophysiological measurements in the mouse heart
For electrocardiogram (ECG) measurements, 6
T3R KO or 6 T3Rß KO
mice, 10 hypothyroid mice, and 5 age-matched control mice were
analyzed. The described procedures were performed in accordance with
the guidelines established by the Committee on Animal Research at the
University of California, San Diego. When mice were sedated after ip
injection of a Ketamine-Xylazine Cocktail, they were kept in a supine
position by four limb restrainers. Four needles were placed sc on each
limb, close to the trunk, and an ECG with the leads I, II, III, AVR,
AVL, and AVF (V, unipolar lead; R, right arm; L, left arm; F, left
foot; A, augmented) was obtained. The ECG was acquired, with an
analog-to-digital conversion at 2000 Hz, on an IBM-compatible 486
computer using Windaq software (Data Instruments, Akron, OH). The
following ECG parameters were measured: 1) duration of atrial
depolarization; 2) duration of excitation progression from the atrium
to the ventricle; 3) duration of depolarization of the ventricles, and
4) duration of excitation and repolarization of the ventricles.
Parameters were measured in each lead and averaged.
Voltage-clamp analysis in right ventricular myocytes
Myocytes were isolated from the right ventricles of control and
hypothyroid adult mice (age, 4–6 weeks) using standard methods.
Whole-cell, voltage- and current-clamp recordings were made from
myocytes, using the following solutions: 1) external: HEPES-buffered
Tyrode solution containing (mM): NaCl (140), KCl (5),
CaCl2 (1), MgCl2 (1), HEPES
(10), glucose (5.5), pH adjusted to 7.4 with NaOH; 2) pipette solution
containing (mM): K+ aspartate (110),
KCl (20), NaCl (8), CaCl2 (1),
MgCl2 (), EGTA (10), K2ATP
(4), HEPES (10), pH adjusted to 7.2 with KOH. Pipette series resistance
in the whole-cell mode was in the range of 4–8 M; 80–90%
series resistance compensation was always applied.
Whole-cell recordings were made with a patch-clamp amplifier (EPC7; List Electronics, Darmstadt, Germany). Recorded membrane potentials were corrected, by -10 mV, in software, to compensate for the patch pipette-bath liquid junction potential. Action potentials were evoked by injection through the patch pipette of 3- to 5-millisecond (ms) current steps (0.7 nA) at 1 Hz. A paired-step voltage clamp protocol was used to isolate the transient outward K current, Ito. This consisted of a pair of 750-ms steps to +30 mV, one with a preceding 100-ms step to -40 mV (to inactivate Ito), and the other without the so-called inactivating prepulse. Ito was isolated by subtraction of membrane currents with and without the inactivating prepulse. Membrane currents were normalized to cell capacitance (pA/pF) to allow averaging of values from different myocytes. All measurements were made at room temperature (21–23 C).
Statistical analysis of the data reported here was done using the Microsoft Corp. (Redmond, WA) Excel program’s Student’s t test function, with an unpaired setting.
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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).
Recent evidence indicates that elimination of expression of
T3R1 in mice, even when the
non-T3 binding T3R2
isoform is still present, results in bradycardia (28).
These mice (with deletion of the T3R1
receptor) are euthyroid. We also observed bradycardia in a separate set
of mice, in which both T3R1 and
T3R2 were eliminated. In these mice, the
T3R locus was disrupted by deleting exon 2,
resulting in elimination of T3R1 and the
T3R2 gene products. Although thyroid hormone
levels were in the normal range, they exhibited marked bradycardia. In
contrast, in mice in which expression of the
T3Rß gene was eliminated by deletion of exons 4
and 5, no bradycardia was observed. This confirms previous findings in
mice with deletion of a functional T3Rß (by
elimination of exon 3), which also did not show a decrease in heart
rate (29). Animals with deletion of exons 4 and 5 of the
T3Rß locus exhibit hyperthyroidism, and these
mice have tachycardia. Placing these animals on PTU and a low-iodine
diet and subsequent treatment with physiological doses of
T3 restores euthyroid status and normal heart
rate. To further explore the molecular mechanisms contributing to the
bradycardia induced by hypothyroidism or deletion of the
T3R gene, we determined the level of the mRNA
coding for the recently identified so-called pacemaker channel genes
HCN2 and HCN4 (12, 13). RNA was prepared from whole hearts
(n = 3). In hypothyroid animals, a decrease of HCN2 mRNA by
61 ± 4.5% (P = 0.0001) was observed; and in
hyperthyroid animals, this parameter increased by 106 ± 12%
(P = 0.0001). In addition, in euthyroid mice with the
deletion of T3R isoform, HCN2 levels were
markedly decreased, by 50 ± 5.5% (P = 0.0001)
(Fig. 1A). In contrast, in mice with
deletion of T3Rß receptor in which a euthyroid
status was restored by PTU and T3 treatment, HCN2
mRNA levels were indistinguishable from those in wild-type control
animals. Mice with a deletion of T3Rß, but not
treated with PTU and T3, are hyperthyroid; and in
these mice, HCN2 mRNA levels are increased by 64 ± 12.5%
(P = 0.0001). The hyperpolarization-activated cation
current If is encoded in heart by two separate
gene products, the HCN2 and the HCN4 gene. Quantification of HCN4 mRNA
levels revealed changes similar to those observed for HCN2. That is,
HCN4 levels were markedly decreased in hypothyroid (45 ± 4%,
P = 0.001) and T3R KO mice
(43 ± 8%, P = 0.0008) and markedly
increased in hyperthyroid (60 ± 13%, P = 0.001)
and T3Rß KO mice (44 ± 6%,
P = 0.0002), as shown in Fig. 1B.
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). In principle this could be caused by a decrease of HCN2
expression. We therefore have analyzed the thyroid hormone regulation
of the pacemaker channels HCN2 and HCN4, in total RNA from the atrium,
by RNase protection (Fig. 1C). We found that HCN2 is regulated by
thyroid hormone in the atrium being reduced by 48 ± 4.5%
(P = 0.005) in hypothyroid mice (Fig. 1C). In contrast,
the mRNA for the HCN4 channel was not significantly changed in the
atrium of hypothyroid animals.
Potassium channel regulation in the ventricle
In addition to modulation of heart rate, changes in thyroid status
can have marked influences on different phases of the action potential
(in particular, action potential repolarization). Accordingly,
whole-cell current clamp recordings of action potentials from the right
ventricle of control and hypothyroid adult mice were compared. These
measurements showed that the durations of action potentials at 50% and
90% repolarization were not significantly different, but the maximum
rate of repolarization (i.e. minimum rate of change of
membrane potential; dV/dt) of action potentials from hypothyroid
mice was significantly smaller than that from controls (Table 2). Because dV/dt is a direct measure of
the net ionic currents that flow during action potential
repolarization, we used whole-cell voltage clamp measurements to gain
further insight into which K+ current may have
contributed to slowed repolarization in ventricular myocytes from
hypothyroid mice. The Ca2+-independent,
transient outward current Ito, which contributes
significantly to early repolarization in mouse ventricular myocytes
(30), was found to be about 50% smaller in right
ventricular myocytes from hypothyroid mice, compared with controls
(Table 2). In line with this decrease in the Ito
current, the level of the mRNA coding for KV 4.2 was significantly
decreased in hypothyroid hearts (12 ± 6.5%, P =
0.003). In T3R KO mice, KV 4.2 mRNA levels
showed a significant decrease (24 ± 5.5%, P =
0.001), whereas KV 4.2 mRNA levels were unchanged from the control
levels in T3Rß KO mice and hyperthyroid mice
(Fig. 2).
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KO mice
(251 ± 25%, P = 0.0001) (Fig. 3). In contrast, a marked decrease in
minK mRNA occurs in hyperthyroid hearts (74 ± 7%,
P = 0.0001) and hearts of T3Rß
KO mice (59 ± 7.5%, P = 0.0001). When
T3Rß KO mice are made euthyroid, minK is
decreased to a much lesser degree (Fig. 3). It is of interest to note
that KV LQT1 is also slightly increased in T3R
KO mice (15 ± 5%, P < 0.05) but is unchanged in
hyperthyroid and T3Rß KO mice (data not shown).
The mRNA level of Kir2.1, which underlies the inward rectifier
K+ current IK1, was not
influenced by thyroid status or deletion of
T3R or T3Rß. Previous
reports have indicated that IK1 current and the
corresponding Kir2.1 channel are not influenced by thyroid status
(31).
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(Table 3).
In contrast, the contractile function of papillary muscle from mice
with deletion of T3Rß was very similar to that
of control animals. The decreased contractile function of cardiac
muscle from T3R mice is accompanied by
decreased expression of the genes coding for some contractile proteins.
The level of MHC mRNA was significantly decreased, by 22 ±
3% (P < 0.01), in the heart of
T3R KO mice (data not shown). In the hearts of
normal wild-type mice, MHC ß mRNA is not present; but in the hearts
of T3Rß KO mice, MHC ß mRNA was clearly
detectable. In contrast, in hearts of euthyroid
T3Rß KO mice, MHC mRNA and MHC ß mRNA
levels were similar to those in the hearts of euthyroid wild-type mice.
In addition, SERCA2 mRNA levels showed a marked decrease in
T3R KO hearts. In contrast, in the hearts of
euthyroid T3Rß KO mice, no significant change
in SERCA2 mRNA occurred (data not shown).
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and
T3Rß in the mouse heart
and T3Rß KO animals, with respect to cardiac
parameters, were different, we determined the expression of the
T3R and T3Rß genes by
RNase protection with total heart RNA from wild-type mice and also from
T3R and T3Rß KO mice.
When the protected bands for T3R1,
T3R2, and
T3Rß1/T3Rß2 were
quantified and the expression of the T3R1/2
was set to 100%, the relative expression of
T3Rß in wild-type mice was 11.5 ± 0.5%
(P < 0.005), as shown in Fig. 4. The expression of
T3Rß, relative to the
T3R1 isoform alone, was 33 ± 4%, shown
in Fig. 4. The predominance of the T3-activated
isoform T3R1 was about equal to
T3R2 mRNA coding for a receptor isoform that
is not activated by T3. On the assumption that
both mRNAs are translated into protein, the marked predominance of
T3R1 over T3Rß could,
in part, explain the different phenotype of the KO mice and lend
support to the hypothesis that the T3R1
isoform is functionally predominant in the heart.
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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and T3Rß can make
distinct contributions to these thyroid hormone-mediated effects and/or
cause changes in gene expression. In principle,
T3R and T3Rß could
exert equal and overlapping effects, and deletion of one of the
receptors may not then result in functional changes or an altered
phenotype. Alternatively, the cross-over or interactions between the
two types of receptors could only be partial, resulting in an absolute
requirement on only one T3R isoform for some
specific T3 effects. Results recently reported
from mice with deletion of only T3R1 (with the
T3R2 receptor still present) demonstrate
bradycardia, suggesting a preferred action of
T3R1 in events related to myogenic pacemaker
activity, e.g. chronotropic action (19, 20).
Alternatively, mice with elimination of T3Rß1
and ß2 are deaf as a result of functional deficiencies in selected
K+ channel expression in the hair cells of the
inner ear (29, 33).
Our results show that homozygous mice with deletion of exon 2 of the
T3R locus, leading to elimination of
T3R1 and T3R2, have a
decreased heart rate. In addition, we show, for the first time, that a
markedly decreased expression of the two genes, termed HCN2 and HCN4,
which code for the If (a so-called cardiac
pacemaker current) occurs in euthyroid T3R KO
mice. The decrease in HCN2 and HCN4 mRNA levels is of similar magnitude
to that seen in hypothyroid wild-type mice. In contrast, mice with
deletion of T3Rß1/2 are hyperthyroid, and they
have an increased heart rate and significantly increased levels of HCN2
and HCN4. When these T3Rß KO mice are made
euthyroid, both heart rate and levels of HCN2 and HCN4 mRNA return to
the normal range. The expression of other ion channel genes, which
are thyroid hormone-responsive, show a similar pattern. For
example, mRNA for KV 4.2, which codes for the
Ca2+-independent transient outward
K+ current Ito, is
decreased by 13% in hypothyroid mice and 24% in euthyroid
T3R KO mice. The magnitude of
Ito was reduced by about 50% in ventricular
myocytes from hypothyroid mice, and reduction of this current
contributes to the reduced rate of repolarization of action potentials
in ventricular myocytes of hypothyroid mice (Table 2). In contrast,
mRNA levels for minK are markedly increased, by 45%, in hypothyroid
mice and by an even larger increase of 251% in euthyroid
T3R KO mice. The minK gene product, together
with KV LQT1, forms the slow delayed rectifier current
IK,s. This current seems to have little role in
action potential repolarization in adult mouse ventricle
(34). Expression of minK is significantly reduced in
T3Rß KO mice because of their hyperthyroid
state but is minimally decreased in T3Rß KO
mice that we made euthyroid. The detailed mechanisms that are
responsible for the marked alterations of the expression of ion channel
genes in the T3R KO mice remain unclear. Two
principal mechanisms could be invoked. One possibility is that
T3R1 is preferentially expressed in cardiac
myocytes, which form the central region of the sinus node where
pacemaker depolarization and action potential initiation occur.
Alternatively, T3R1 receptors might exert
preferential effects on the expression of specific genes attributable,
for example, to preferred binding to thyroid hormone response elements
of a specific configuration or by association with other specific
nuclear proteins involved in the transcription of these genes.
Differences in the protein structure of T3R1
vs. T3Rß1, especially at the
N-terminal end, might provide for such T3 isoform
specific interactions. Currently, it is not possible to distinguish
between these mechanisms or determine their relative importance. Some
data are, however, compatible with the hypothesis that sinus node
myocytes in the right atrium predominantly express
T3R1. Patients with the resistance to thyroid
hormone syndrome, presenting with elevated thyroid hormone levels and,
frequently, a marked tachycardia, seem to express the mutant
T3Rß, which exerts a dominant negative action
on normal thyroid hormone function in myocytes of the right atrium
(35). T3R1 exclusively or
predominantly expressed in the myocytes of sinus nodes would be
unopposed in its activation by the T3Rß mutant.
In addition, increased T3 levels will lead to
increased occupancy of T3R1, resulting in
increased HCN2 and HCN4 expression and, partly for this reason,
elevated heart rate. T3Rß may be expressed at
significant levels, equal to those of T3R in
other parts of the heart. Previous findings in the rat heart indicate
roughly equal levels of T3R and
T3Rß gene products (18). Our
findings in mouse hearts indicate almost 10-fold higher levels of
T3R1 and T3R2 gene
expression, at the mRNA level, than T3Rß gene
expression. The more marked cardiac phenotype in
T3R KO mice, compared with
T3Rß KO mice, would be compatible with a much
higher T3R predominance. The mRNA that we used
for most of our studies, to quantify the predominance of ion channel
gene expression, is derived from myocytes of the left ventricle,
because this part of the heart makes the major contribution to total
cardiac RNA. The mRNA coding, for example, for minK, which is part of
the delayed IK,s channel derived primarily from
such myocytes, also showed distinct changes in
T3R KO hearts. These findings may indicate
that, in addition to preferred expression of
T3R in myocytes of the sinus node, in left
ventricular myocytes, T3R may exert
preferential influence on the expression of specific genes, like the
gene coding for minK. Electrical impulse generation occurs in
structures of the cardiac atrium. We found HCN2 mRNA markedly decreased
in the atrium of hypothyroid mice, which corresponds to the observed
bradycardia.
The contractile studies that we performed on papillary muscle of
T3R KO and T3Rß KO
mice also point to a predominance of T3R
action in the left cardiac ventricle. Contractile function in muscle
from T3R KO mice was significantly diminished,
relative to force development and force decay. In contrast, papillary
muscle function was normal in muscle obtained from
T3Rß KO mice. Corresponding changes occurred in
the mRNA for MHC isoform coding proteins that are linked to force
generation and in SERCA2 mRNA that is linked to calcium lowering during
diastole. Decreased levels of MHC mRNA and SERCA2 mRNA may present
the cause for slowed force development and prolonged relaxation
time.
Previous findings by Johanssen (19) show that bradycardia
occurs in T3R1 KO mice and is not related to
altered sympathetic or parasympathetic innervation. Our findings of
decreased papillary muscle function in isolated muscle strips, which is
independent of the sympathetic parasympathetic innervation status,
makes it very likely that the observed effects occurring in
T3R1 KO mice are primarily related to
decreased T3 action in the cardiac myocyte and
are largely independent of the innervation status. The shortened life
span of the T3R KO mice and their inability to
reproduce presented an obstacle to our studies. This prevented the
breeding of homozygous T3R KO mice in which
long-term and more detailed studies could have been performed.
In summary, our findings demonstrate that T3R1
exerts a predominant effect on cardiac electrophysiological phenomena
like impulse generation and mechanical functions, e.g.
systolic force generation and diastolic relaxation. The molecular basis
for the predominance of T3R action in cardiac
myocytes needs to be explored in further detail but is compatible with
our finding of a significantly higher predominance of
T3R encoding mRNA over
T3Rß encoding mRNA in the normal mouse
heart.
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Acknowledgments |
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Footnotes |
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2 A Medical Scientist of the Alberta Heritage Foundation for Medical
Research.
Received August 3, 2000.
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 S. J. Swoap, C. Li, J. Wess, A. D. Parsons, T. D. Williams, and J. M. Overton Vagal tone dominates autonomic control of mouse heart rate at thermoneutrality Am J Physiol Heart Circ Physiol, April 1, 2008; 294(4): H1581 - H1588. [Abstract] [Full Text] [PDF]
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 C R Liu, L Y Li, F Shi, X Y Zang, Y M Liu, Y Sun, and B H Kan Effects of hyper- and hypothyroid on expression of thyroid hormone receptor mRNA in rat myocardium J. Endocrinol., December 1, 2007; 195(3): 429 - 438. [Abstract] [Full Text] [PDF]
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 L. Quignodon, S. Vincent, H. Winter, J. Samarut, and F. Flamant A Point Mutation in the Activation Function 2 Domain of Thyroid Hormone Receptor {alpha}1 Expressed after CRE-Mediated Recombination Partially Recapitulates Hypothyroidism Mol. Endocrinol., October 1, 2007; 21(10): 2350 - 2360. [Abstract] [Full Text] [PDF]
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S. A. Grandy, V. Trepanier-Boulay, and C. Fiset Postnatal development has a marked effect on ventricular repolarization in mice Am J Physiol Heart Circ Physiol, October 1, 2007; 293(4): H2168 - H2177. [Abstract] [Full Text] [PDF]
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 I Stoykov, H C van Beeren, A F M Moorman, V M Christoffels, W M Wiersinga, and O Bakker Effect of amiodarone and dronedarone administration in rats on thyroid hormone-dependent gene expression in different cardiac components Eur. J. Endocrinol., June 1, 2007; 156(6): 695 - 702. [Abstract] [Full Text] [PDF]
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 D. D. Belke, B. Gloss, E. A. Swanson, and W. H. Dillmann Adeno-Associated Virus-Mediated Expression of Thyroid Hormone Receptor Isoforms-{alpha}1 and -{beta}1 Improves Contractile Function in Pressure Overload-Induced Cardiac Hypertrophy Endocrinology, June 1, 2007; 148(6): 2870 - 2877. [Abstract] [Full Text] [PDF]
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F. Flamant, K. Gauthier, and J. Samarut Thyroid Hormones Signaling Is Getting More Complex: STORMs Are Coming Mol. Endocrinol., February 1, 2007; 21(2): 321 - 333. [Abstract] [Full Text] [PDF]
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I Stoykov, B Zandieh-Doulabi, A F M Moorman, V Christoffels, W M Wiersinga, and O Bakker Expression pattern and ontogenesis of thyroid hormone receptor isoforms in the mouse heart. J. Endocrinol., May 1, 2006; 189(2): 231 - 245. [Abstract] [Full Text] [PDF]
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 M. G. Trivieri, G. Y. Oudit, R. Sah, B.-G. Kerfant, H. Sun, A. O. Gramolini, Y. Pan, A. D. Wickenden, W. Croteau, G. Morreale de Escobar, et al. Cardiac-specific elevations in thyroid hormone enhance contractility and prevent pressure overload-induced cardiac dysfunction PNAS, April 11, 2006; 103(15): 6043 - 6048. [Abstract] [Full Text] [PDF]
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P. J. O'Shea, J. H. D. Bassett, S. Sriskantharajah, H. Ying, S.-y. Cheng, and G. R. Williams Contrasting Skeletal Phenotypes in Mice with an Identical Mutation Targeted to Thyroid Hormone Receptor {alpha}1 or {beta} Mol. Endocrinol., December 1, 2005; 19(12): 3045 - 3059. [Abstract] [Full Text] [PDF]
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Y. Hu, S. V. P. Jones, and W. H. Dillmann Effects of hyperthyroidism on delayed rectifier K+ currents in left and right murine atria Am J Physiol Heart Circ Physiol, October 1, 2005; 289(4): H1448 - H1455. [Abstract] [Full Text] [PDF]
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 Y. Sato, R. Nakamura, M. Satoh, K. Fujishita, S. Mori, S. Ishida, T. Yamaguchi, K. Inoue, T. Nagao, and Y. Ohno Thyroid Hormone Targets Matrix Gla Protein Gene Associated With Vascular Smooth Muscle Calcification Circ. Res., September 16, 2005; 97(6): 550 - 557. [Abstract] [Full Text] [PDF]
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 G. J. Kahaly and W. H. Dillmann Thyroid Hormone Action in the Heart Endocr. Rev., August 1, 2005; 26(5): 704 - 728. [Abstract] [Full Text] [PDF]
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