Biomedical subjects

Céline Marionneau

Publications and source records attributed to Céline Marionneau.

10 recordsLinked to original sources

PubMed2006

Calmodulin kinase II inhibition shortens action potential duration by upregulation of K+ currents.

The multifunctional Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) is activated by elevated intracellular Ca(2+) (Ca(2+)(i)), and mice with chronic myocardial CaMKII inhibition (Inh) resulting from transgenic expression of a CaMKII inhibitory peptide (AC3-I) unexpectedly showed action potential duration (APD) shortening. Inh mice exhibit increased L-type Ca(2+) current (I(Ca)), because of upregulation of protein kinase A (PKA) activity, and decreased CaMKII-dependent phosphorylation of phospholamban (PLN). We hypothesized that CaMKII is a molecular signal linking Ca(2+)(i) to repolarization. Whole cell voltage-clamp recordings revealed that the fast transient outward current (I(to,f)) and the inward rectifier current (I(K1)) were selectively upregulated in Inh, compared with wild-type (WT) and transgenic control, mice. Breeding Inh mice with mice lacking PLN returned I(to,f) and I(K1) to control levels and equalized the APD and QT intervals in Inh mice to control and WT levels. Dialysis of AC3-I into WT cells did not result in increased I(to,f) or I(K1), suggesting that enhanced cardiac repolarization in Inh mice is an adaptive response to chronic CaMKII inhibition rather than an acute effect of reduced CaMKII activity. Increasing PKA activity, by cell dialysis with cAMP, or inhibition of PKA did not affect I(K1) in WT cells. Dialysis of WT cells with cAMP also reduced I(to,f), suggesting that PKA upregulation does not increase repolarizing K(+) currents in Inh mice. These findings provide novel in vivo and cellular evidence that CaMKII links Ca(2+)(i) to cardiac repolarization and suggest that PLN may be a critical CaMKII target for feedback regulation of APD in ventricular myocytes.

Action Potentials↗

PubMed2005

Targeted deletion of Kv4.2 eliminates I(to,f) and results in electrical and molecular remodeling, with no evidence of ventricular hypertrophy or myocardial dysfunction.

Previous studies have demonstrated a role for voltage-gated K+ (Kv) channel alpha subunits of the Kv4 subfamily in the generation of rapidly inactivating/recovering cardiac transient outward K+ current, I(to,f), channels. Biochemical studies suggest that mouse ventricular I(to,f) channels reflect the heteromeric assembly of Kv4.2 and Kv4.3 with the accessory subunits, KChIP2 and Kvbeta1, and that Kv4.2 is the primary determinant of regional differences in (mouse ventricular) I(to,f) densities. Interestingly, the phenotypic consequences of manipulating I(to,f) expression in different mouse models are distinct. In the experiments here, the effects of the targeted deletion of Kv4.2 (Kv4.2(-/-)) were examined. Unexpectedly, voltage-clamp recordings from Kv4.2(-/-) ventricular myocytes revealed that I(to,f) is eliminated. In addition, the slow transient outward K+ current, I(to,s), and the Kv1.4 protein (which encodes I(to,s)) are upregulated in Kv4.2(-/-) ventricles. Although Kv4.3 mRNA/protein expression is not measurably affected, KChIP2 expression is markedly reduced in Kv4.2(-/-) ventricles. Similar to Kv4.3, expression of Kvbeta1, as well as Kv1.5 and Kv2.1, is similar in wild-type and Kv4.2(-/-) ventricles. In addition, and in marked contrast to previous findings in mice expressing a truncated Kv4.2 transgene, the elimination I(to,f) in Kv4.2(-/-) mice does not result in ventricular hypertrophy. Taken together, these findings demonstrate not only an essential role for Kv4.2 in the generation of mouse ventricular I(to,f) channels but also that the loss of I(to,f) per se does not have overt pathophysiological consequences.

Animals↗

PubMed2005

Chronic heart rate reduction remodels ion channel transcripts in the mouse sinoatrial node but not in the ventricle.

We investigated the effects of chronic and moderate heart rate (HR) reduction on ion channel expression in the mouse sinoatrial node (SAN) and ventricle. Ten-week-old male C57BL/6 mice were treated twice daily with either vehicle or ivabradine at 5 mg/kg given orally during 3 wk. The effects of HR reduction on cardiac electrical activity were investigated in anesthetized mice with serial ECGs and in freely moving mice with telemetric recordings. With the use of high-throughput real-time RT-PCR, the expression of 68 ion channel subunits was evaluated in the SAN and ventricle at the end of the treatment period. In conscious mice, ivabradine induced a mean 16% HR reduction over a 24-h period that was sustained over the 3-wk administration. Other ECG parameters were not modified. Two-way hierarchical clustering analysis of gene expression revealed a separation of ventricles from SANs but no discrimination between treated and untreated ventricles, indicating that HR reduction per se induced limited remodeling in this tissue. In contrast, SAN samples clustered in two groups depending on the treatment. In the SAN from ivabradine-treated mice, the expression of nine ion channel subunits, including Navbeta1 (-25%), Cav3.1 (-29%), Kir6.1 (-28%), Kvbeta2 (-41%), and Kvbeta3 (-30%), was significantly decreased. Eight genes were significantly upregulated, including K+ channel alpha-subunits (Kv1.1, +30%; Kir2.1, +29%; Kir3.1, +41%), hyperpolarization-activated cation channels (HCN2, +24%; HCN4, +52%), and connexin 43 (+26%). We conclude that reducing HR induces a complex remodeling of ion channel expression in the SAN but has little impact on ion channel transcripts in the ventricle.

Animals↗

PubMed2005

Functional genomics of cardiac ion channel genes.

Ion channels are an ensemble of specialized membrane proteins that act in concert to create and modulate the electrical activity of many excitable cells, including cardiac myocytes. Following completion of the sequencing of various genomes, including that of the human, the complete repertoire of ion channel genes has been elucidated for different species. How transcripts issued from this gene collection are expressed and modulated in relation to variable physiological and pathological situations is the subject of functional or physiological genomics. Specialized microarrays (IonChips) comprising probes for the ensemble of ion channel and regulatory genes were developed as an alternative to whole-genome DNA chips. Physiological genomics of cardiac ion channel genes is a growing field that, in combination with genetics, should markedly increase our comprehension of the molecular mechanisms leading to arrhythmias.

Amiodarone↗

PubMed2005

Mouse model of SCN5A-linked hereditary Lenègre's disease: age-related conduction slowing and myocardial fibrosis.

BACKGROUND: We have previously linked hereditary progressive cardiac conduction defect (hereditary Lenègre's disease) to a loss-of-function mutation in the gene encoding the main cardiac Na+ channel, SCN5A. In the present study, we investigated heterozygous Scn5a-knockout mice (Scn5a+/- mice) as a model for hereditary Lenègre's disease. METHODS AND RESULTS: In Scn5a+/- mice, surface ECG recordings showed age-related lengthening of the P-wave and PR- and QRS-interval duration, coinciding with previous observations in patients with Lenègre's disease. Old but not young Scn5a+/- mice showed extensive fibrosis of their ventricular myocardium, a feature not seen in wild-type animals. In old Scn5a+/- mice, fibrosis was accompanied by heterogeneous expression of connexin 43 and upregulation of hypertrophic markers, including beta-MHC and skeletal alpha-actin. Global connexin 43 expression as assessed with Western blots was similar to wild-type mice. Decreased connexin 40 expression was seen in the atria. Using pangenomic microarrays and real-time PCR, we identified in Scn5a+/- mice an age-related upregulation of genes encoding Atf3 and Egr1 transcription factors. Echocardiography and hemodynamic investigations demonstrated conserved cardiac function with aging and lack of ventricular hypertrophy. CONCLUSIONS: We conclude that Scn5a+/- mice convincingly recapitulate the Lenègre's disease phenotype, including progressive impairment with aging of atrial and ventricular conduction associated with myocardial rearrangements and fibrosis. Our work provides the first demonstration that a monogenic ion channel defect can progressively lead to myocardial structural anomalies.

Age Factors↗

PubMed2005

In vitro molecular interactions and distribution of KCNE family with KCNQ1 in the human heart.

OBJECTIVE: The voltage-gated K+ channel KCNQ1 associates with the small KCNE1 beta subunit to underlie the IKs repolarizing current in the heart. Based on sequence homology, the KCNE family is recognized to comprise five members. Controversial data have indicated their participation in several K+ channel protein complexes, including KCNQ1. The expression level and the putative functions of the different KCNE subunits in the human heart still require further investigation. METHODS: We have carried out a comparative study of all KCNE subunits with KCNQ1 using the patch-clamp technique in mammalian cells. Real-time RT-PCR absolute quantification was performed on human atrial and ventricular tissue. RESULTS: While KCNQ1/KCNE1 heteromultimer reached high current density with slow gating kinetics and pronounced voltage dependence, KCNQ1/KCNE2 and KCNQ1/KCNE3 complexes produced instantaneous voltage-independent currents with low and high current density, respectively. Co-expression of KCNE4 or KCNE5 with KCNQ1 induced small currents in the physiological range of voltages, with kinetics similar to those of the KCNQ1/KCNE1 complex. However, co-expression of these inhibitory subunits with a disease-associated mutation (S140G-KCNQ1) led to currents that were almost undistinguishable from the KCNQ1/KCNE1 canonical complex. Absolute cDNA quantification revealed a relatively homogeneous distribution of each transcript, except for KCNE4, inside left atria and endo- and epicardia of left ventricular wall with the following abundance: KCNQ1 >> KCNE4 > or = KCNE1 > KCNE3 > KCNE2 > KCNE5. KCNE4 expression was twice as high in atrium compared to ventricle. CONCLUSIONS: Our data show that KCNQ1 forms a channel complex with 5 KCNE subunits in a specific manner but only interactions with KCNE1, KCNE2, and KCNE3 may have physiological relevance in the human heart.

Animals↗

PubMed2004

Long-term amiodarone administration remodels expression of ion channel transcripts in the mouse heart.

BACKGROUND: The basis for the unique effectiveness of long-term amiodarone treatment on cardiac arrhythmias is incompletely understood. The present study investigated the pharmacogenomic profile of amiodarone on genes encoding ion-channel subunits. METHODS AND RESULTS: Adult male mice were treated for 6 weeks with vehicle or oral amiodarone at 30, 90, or 180 mg x kg(-1) x d(-1). Plasma and myocardial levels of amiodarone and N-desethylamiodarone increased dose-dependently, reaching therapeutic ranges observed in human. Plasma triiodothyronine levels decreased, whereas reverse triiodothyronine levels increased in amiodarone-treated animals. In ECG recordings, amiodarone dose-dependently prolonged the RR, PR, QRS, and corrected QT intervals. Specific microarrays containing probes for the complete ion-channel repertoire (IonChips) and real-time reverse transcription-polymerase chain reaction experiments demonstrated that amiodarone induced a dose-dependent remodeling in multiple ion-channel subunits. Genes encoding Na+ (SCN4A, SCN5A, SCN1B), connexin (GJA1), Ca2+ (CaCNA1C), and K+ channels (KCNA5, KCNB1, KCND2) were downregulated. In patch-clamp experiments, lower expression of K+ and Na+ channel genes was associated with decreased I(to,f), I(K,slow), and I(Na) currents. Inversely, other K+ channel alpha- and beta-subunits, such as KCNA4, KCNK1, KCNAB1, and KCNE3, were upregulated. CONCLUSIONS: Long-term amiodarone treatment induces a dose-dependent remodeling of ion-channel expression that is correlated with the cardiac electrophysiologic effects of the drug. This profile cannot be attributed solely to the amiodarone-induced cardiac hypothyroidism syndrome. Thus, in addition to the direct effect of the drug on membrane proteins, part of the therapeutic action of long-term amiodarone treatment is likely related to its effect on ion-channel transcripts.

Amiodarone↗

PubMed2004

Specific pattern of ionic channel gene expression associated with pacemaker activity in the mouse heart.

Even though sequencing of the mammalian genome has led to the discovery of a large number of ionic channel genes, identification of the molecular determinants of cellular electrical properties in different regions of the heart has been rarely obtained. We developed a high-throughput approach capable of simultaneously assessing the expression pattern of ionic channel repertoires from different regions of the mouse heart. By using large-scale real-time RT-PCR, we have profiled 71 channels and related genes in the sinoatrial node (SAN), atrioventricular node (AVN), the atria (A) and ventricles (V). Hearts from 30 adult male C57BL/6 mice were microdissected and RNA was isolated from six pools of five mice each. TaqMan data were analysed using the threshold cycle (C(t)) relative quantification method. Cross-contamination of each region was checked with expression of the atrial and ventricular myosin light chains. Two-way hierarchical clustering analysis of the 71 genes successfully classified the six pools from the four distinct regions. In comparison with the A, the SAN and AVN were characterized by higher expression of Nav beta 1, Nav beta 3, Cav1.3, Cav3.1 and Cav alpha 2 delta 2, and lower expression of Kv4.2, Cx40, Cx43 and Kir3.1. In addition, the SAN was characterized by higher expression of HCN1 and HCN4, and lower expression of RYR2, Kir6.2, Cav beta 2 and Cav gamma 4. The AVN was characterized by higher expression of Nav1.1, Nav1.7, Kv1.6, Kvbeta1, MinK and Cav gamma 7. Other gene expression profiles discriminate between the ventricular and the atrial myocardium. The present study provides the first genome-scale regional ionic channel expression profile in the mouse heart.

Animals↗

PubMed2003

RhoA expression is controlled by nitric oxide through cGMP-dependent protein kinase activation.

The small G protein RhoA is a convergence point for multiple signals that regulate smooth muscle cell functions. NO plays a major role in the structure and function of the normal adult vessel wall, mainly through modulation of gene transcription. This study was thus performed to analyze in vitro and in vivo the effect of NO signaling on RhoA expression in arterial smooth muscle cells. In rat or human artery smooth muscle cells, sodium nitroprusside or 8-(2-chlorophenylthio)-cGMP induced a rise in RhoA mRNA and protein expression, which was inhibited by the cGMP-dependent protein kinase (PKG) inhibitor (R(p))-8-bromo-beta-phenyl-1,N(2)-ethenoguanosine 3':5'-phosphorothioate. The NO/PKG stimulation of RhoA expression involved both an increase in RhoA protein stability and stimulation of rhoA gene transcription. Cloning and functional analysis of the human rhoA promoter revealed that the effect of NO/PKG involved phosphorylation of ATF-1 and subsequent binding to the cAMP-response element. Chronic inhibition of NO synthesis in N(omega)-nitro-l-arginine-treated rats induced a strong decrease in RhoA mRNA and protein expression in aorta and pulmonary artery associated with inhibition of RhoA-mediated Ca(2+) sensitization. These effects were prevented by oral administration of the cGMP phosphodiesterase inhibitor sildenafil. These results show that NO/PKG signaling positively controls RhoA expression and suggest that the basal release of NO is necessary to maintain RhoA expression and RhoA-dependent functions in vascular smooth muscle cells.

3T3 Cells↗

PubMed2003

Structure-activity relationships of human urotensin II and related analogues on rat aortic ring contraction.

The sequence of human urotensin II (UII) has been recently established as H-Glu-Thr-Pro-Asp-Cys-Phe-Trp-Lys-Tyr-Cys-Val-OH, and it has been reported that UII is the most potent mammalian vasoconstrictor peptide identified so far. A series of UII analogues was synthesized, and the contractile activity of each compound was studied in vitro using de-endothelialised rat aortic rings. Replacement of each amino acid by an L-alanine or by a D-isomer showed that the N- and C-terminal residues flanking the cyclic region of the amidated peptide were relatively tolerant to substitution. Conversely, replacement of any residue of the cyclic region significantly reduced the contractile activity of the molecule. The octapeptide UII(4-11) was 4 times more potent than UII, indicating that the C-terminal region of the molecule possesses full biological activity. Alanine or D-isomer substitutions in UII(4-11) or in UII(4-11)-NH2, respectively, showed a good correlation with the results obtained for UII-NH2. Disulfide bridge disruption or replacement of the cysteine residues by their D-enantiomers markedly reduced the vasoconstrictor effect of UII and its analogues. In contrast, acetylation of the N-terminal residue of UII and UII-NH2 enhanced the potency of the peptide. Finally, monoiodination of the Tyr6 residue in UII(4-11) increased by 5 fold the potency of the peptide in the aortic ring bioassay. This structure-activity relationship study should provide useful information for the rational design of selective and potent UII receptor agonists and antagonists.

Amino Acid Sequence↗