Phosphoproteomics reveals a novel mechanism underlying the proarrhythmic effects of nilotinib, vandetanib, and mobocertinib.

The use of tyrosine kinase inhibitors (TKIs) has resulted in significant occurrence of arrhythmias. However, the precise mechanism of the proarrhythmic effect is not fully understood. In this study, we found that nilotinib (NIL), vandetanib (VAN), and mobocertinib (MOB) induced the development of "cellrhythmia" (arrhythmia-like events) in a concentration-dependent manner in human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs). Continuous administration of NIL, VAN, or MOB in animals significantly prolonged the action potential durations (APD) and increased susceptibility to arrhythmias. Using phosphoproteomic analysis, we identified proteins with altered phosphorylation levels after treatment with 3 µM NIL, VAN, and MOB for 1.5 h. Using these identified proteins as substrates, we performed kinase-substrate enrichment analysis to identify the kinases driving the changes in phosphorylation levels of these proteins. MAPK and WNK were both inhibited by NIL, VAN, and MOB. A selective inhibitor of WNK1, WNK-IN-11, induced concentration- and time-dependent cellrhythmias and prolonged field potential duration (FPD) in hiPSC-CMs in vitro; furthermore, administration in guinea pigs confirmed that WNK-IN-11 prolonged ventricular repolarization and increased susceptibility to arrhythmias. Fingding indicated that WNK1 inhibition had an in vivo and in vitro arrhythmogenic phenotype similar to TKIs. Additionally,three of TKIs reduced hERG and KCNQ1 expression at protein level, not at transcription level. Similarly, the knockdown of WNK1 decreased hERG and KCNQ1 protein expression in hiPSC-CMs. Collectively, our data suggest that the proarrhythmic effects of NIL, VAN, and MOB occur through a kinase inhibition mechanism. NIL, VAN, and MOB inhibit WNK1 kinase, leading to a decrease in hERG and KCNQ1 protein expression, thereby prolonging action potential repolarization and consequently cause arrhythmias.

Transcriptomics Coupled with Proteomics Reveals Osimertinib-induced Myocardial Mitochondrial Dysfunction.

Osimertinib, an irreversible epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI) used for cancer treatment, can cause significant cardiac toxicity. However, the specific mechanism of osimertinib-induced cardiotoxicity is not fully understood. In this study, we administered osimertinib to mice and neonatal rat ventricular myocytes (NRVMs). We observed significant structural and functional damage to the hearts of these mice, along with a marked increase in cardiac injury biomarkers and accompanying ultrastructural damage to mitochondria. We integrated 4D label-free protein quantification and RNA-Seq methods to analyze the sequencing data of NRVMs under osimertinib treatment (0 and 2.5µM). Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis evidenced that differentially expressed genes (DEGs)and differentially expressed proteins (DEPs) were distinctly enriched for oxidative phosphorylation (OXPHOs). Simultaneously, osimertinib primarily affected the contents of adenosine triphosphate (ATP). Further investigations revealed that osimertinib disrupts the functions of the ATP synthase (complex V), leading to a reduction in ATP production. Taken together, our data demonstrated that osimertinib causes mitochondrial dysfunction, which in turn leads to the onset of cardiac toxicity.

Healing of the Torn Anterior Horn of Rabbit Medial Meniscus to Bone after Transtibial Pull-Out Repair and Autologous Platelet-Rich Plasma Gel Injection.

OBJECTIVES: The transtibial pull-out repair (TP) is a relatively new method for treating meniscal root tear; however, the clinical evaluation of its healing effect remains controversial. Due to ethical constraints and limitations of imaging techniques in humans, here we dynamically observe the healing effects of TP and TP with platelet-rich plasma gel (PRG) at the histological level using an animal model. METHODS: Platelet-rich plasma (PRP) and PRG of rabbits were prepared. Platelet-derived growth factor (PDGF) and transforming growth factor-ß1 (TGF-ß1) levels in PRP and PRG were determined using an enzyme-linked immunosorbent assay. A rabbit model of anterior horn tear of the medial meniscus and TP surgery were created. PRG was injected between the anterior horn of the medial meniscus and the tibial tunnel. Rabbits were divided into three groups: the anterior horn tear group (Tear group), the anterior horn tear + TP group (TP group), and the anterior horn tear + TP + PRG group (TP + PRG group). The healing effect was observed dynamically using histopathological studies and biomechanical experiments. RESULTS: The platelet content in PRP significantly increased to approximately 4.57 times that of whole blood. PDGF and TGF-ß1 concentrations in PRG increased to 2.46 and 4.15 times those in PRP, respectively. Hematoxylin and eosin (H&E) and Masson staining showed that the number of inflammatory cells in healing tissue decreased and the collagen fibers significantly increased in TP and TP + PRG groups at 4, 8, and 12 weeks postoperatively compared to those in Tear group. Neatly arranged, interlaced, and dense collagen fibers were found between the anterior horn and bone at 12 weeks. H&E and toluidine blue staining showed that the injury to the femoral condyle cartilage was alleviated. The healing performance in TP + PRG group was better and faster than that in TP group. The maximum tensile fracture strength of the meniscus progressively increased at 8 and 12 weeks postoperatively. CONCLUSIONS: Anterior horn injury of the medial meniscus in rabbits can be repaired using the TP technique, and the addition of autologous PRG to the bone tunnel promotes early healing of the meniscus and bone postoperatively. Meanwhile, both treatments can reduce the secondary damage to the cartilage due to osteoarthritis.

Pharmacological suppression of Nedd4-2 rescues the reduction of Kv11.1 channels in pathological cardiac hypertrophy.

The human ether-á-go-go-related gene (hERG) encodes the pore-forming subunit (Kv11.1), conducting a rapidly delayed rectifier K+ current (I Kr). Reduction of I Kr in pathological cardiac hypertrophy (pCH) contributes to increased susceptibility to arrhythmias. However, practical approaches to prevent I Kr deficiency are lacking. Our study investigated the involvement of ubiquitin ligase Nedd4-2-dependent ubiquitination in I Kr reduction and sought an intervening approach in pCH. Angiotensin II (Ang II) induced a pCH phenotype in guinea pig, accompanied by increased incidences of sudden death and higher susceptibility to arrhythmias. Patch-clamp recordings revealed a significant I Kr reduction in pCH cardiomyocytes. Kv11.1 protein expression was decreased whereas its mRNA level did not change. In addition, Nedd4-2 protein expression was increased in pCH, accompanied by an enhanced Nedd4-2 and Kv11.1 binding detected by immunoprecipitation analysis. Cardiac-specific overexpression of inactive form of Nedd4-2 shortened the prolonged QT interval, reversed I Kr reduction, and decreased susceptibility to arrhythmias. A synthesized peptide containing the PY motif in Kv11.1 C-terminus binding to Nedd4-2 and a cell-penetrating sequence antagonized Nedd4-2-dependent degradation of the channel and increased the surface abundance and function of hERG channel in HEK cells. In addition, in vivo administration of the PY peptide shortened QT interval and action potential duration, and enhanced I Kr in pCH. We conclude that Nedd4-2-dependent ubiquitination is critically involved in I Kr deficiency in pCH. Pharmacological suppression of Nedd4-2 represents a novel approach for antiarrhythmic therapy in pCH.

Downregulation of hERG channel expression by tyrosine kinase inhibitors nilotinib and vandetanib predominantly contributes to arrhythmogenesis.

Cardiotoxicity by tyrosine kinase inhibitors remains an important concern. Nilotinib and vandetanib clinically carry high proarrhythmic risk and the exact mechanism underlying arrhythmogenesis is not fully understood. In this study, we investigated the effects of nilotinib and vandetanib on the abundance of human ether-á-go-go-related gene (hERG) K+ channel and assessed the potential role of acute hERG blockage versus chronic effects in arrhythmogenesis. We found that both nilotinib and vandetanib prolonged the field potential duration reflecting the repolarisation process and induced cellrythmias of human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) in a time-and concentration-dependent manner after, after chronic exposure. Patch-clamp recordings revealed significant reductions of hERG current densities by nilotinib or vandetanib after chronic incubation with hERG-HEK293 cells in addition to the acute inhibition. Western blot analysis showed that nilotinib and vandetanib decreased mature hERG protein (155-kDa) expression, in a greater extent than that of the immature form (135-kDa). A serum and glucocorticoid kinase 1 (SGK1) activator, C4-ceramide, prevented the nilotinib-and vandetanib-induced hERG protein downregulation and thus the incidence of cellrrhythmias. Taken together, our data demonstrated that the downregulation of hERG channel abundance on the cellular membrane predominantly contributed to the proarrhythmic effect of nilotinib and vandetanib.

Inhibitory G-protein-mediated modulation of slow delayed rectifier potassium channels contributes to increased susceptibility to arrhythmogenesis in aging heart.

BACKGROUND: Slow delayed rectifier potassium current (IKs) is an important component of repolarization reserve during sympathetic nerve excitement. However, little is known about age-related functional changes of IKs and its involvement in age-dependent arrhythmogenesis. OBJECTIVE: The purpose of this study was to investigate age-related alteration of the IKs response to ß-adrenergic receptor (ßAR) activation. METHODS: Dunkin-Hartley guinea pigs were used. Whole-cell patch-clamp recording was used to record K+ currents. Optical mapping of membrane potential was performed in ex vivo heart. RESULTS: There was no difference in IKs density in ventricular cardiomyocytes between young and old guinea pigs. However, in contrast to IKs potentiation in young hearts, isoproterenol (ISO) evoked an acute inhibition on IKs in a concentration-dependent manner in old guinea pig hearts. The ß2AR antagonist, but not ß1AR antagonist, reversed the inhibitory response. Preincubation of cardiomyocytes with the inhibitory G protein (Gi) inhibitor pertussis toxin (PTX) also reversed the inhibitory response. In HEK293 cells cotransfected with cloned IKs channel and ß2AR, ISO enhanced the current but reduced it when cells were cotransfected with Gi2, and PTX restored the ISO-induced excitatory response. Moreover, in aging cardiomyocytes, Gßγ inhibitor gallein, PLC inhibitor U73122, or protein kinase C inhibitor Bis-1 prevented the reduction of IKs by ISO. Furthermore, cardiac-specific Gi2 overexpression in young guinea pigs predisposed the heart to ventricular tachyarrhythmias. PTX pretreatment protected the hearts from ventricular arrhythmias. CONCLUSION: ßAR activation acutely induces an inhibitory IKs response in aging guinea pig hearts through ß2AR-Gi signaling, which contributes to increased susceptibility to arrhythmogenesis in aging hearts.

Auxiliary subunits control biophysical properties and response to compound NS5806 of the Kv4 potassium channel complex.

Kv4 pore-forming subunits co-assemble with ß-subunits including KChIP2 and DPP6 and the resultant complexes conduct cardiac transient outward K+ current (Ito). Compound NS5806 has been shown to potentate Ito in canine cardiomyocytes; however, its effects on Ito in other species yet to be determined. We found that NS5806 inhibited native Ito in a concentration-dependent manner (0.1~30 µM) in both mouse ventricular cardiomyocytes and human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs), but potentiated Ito in the canine cardiomyocytes. In HEK293 cells co-transfected with cloned Kv4.3 (or Kv4.2) and ß-subunit KChIP2, NS5806 significantly increased the peak current amplitude and slowed the inactivation. In contrast, NS5806 suppressed the current and accelerated inactivation of the channels when cells were co-transfected with Kv4.3 (or Kv4.2), KChIP2 and another ß-subunit, DPP6-L (long isoform). Western blot analysis showed that DPP6-L was dominantly expressed in both mouse ventricular myocardium and hiPSC-CMs, while it was almost undetectable in canine ventricular myocardium. In addition, low level of DPP6-S expression was found in canine heart, whereas levels of KChIP2 expression were comparable among all three species. siRNA knockdown of DPP6 antagonized the Ito inhibition by NS5806 in hiPSC-CMs. Molecular docking simulation suggested that DPP6-L may associate with KChIP2 subunits. Mutations of putative KChIP2-interacting residues of DPP6-L reversed the inhibitory effect of NS5806 into potentiation of the current. We conclude that a pharmacological modulator can elicit opposite regulatory effects on Kv4 channel complex among different species, depending on the presence of distinct ß-subunits. These findings provide novel insight into the molecular design and regulation of cardiac Ito. Since Ito is a potential therapeutic target for treatment of multiple cardiovascular diseases, our data will facilitate the development of new therapeutic Ito modulators.

Transcriptome analysis reveals new microRNAs-mediated pathway involved in anther development in male sterile wheat.

BACKGROUND: 337S is a novel bi-pole-photo-thermo-sensitive genic male sterile line in wheat, and sensitive to both long day length/high temperature and short day length/low temperature condition. Although the regulatory function of MicroRNAs (miRNAs) in reproductive development has been increasingly studied, their roles in pre-meiotic and meiotic cells formation of plants have not been clearly explored. Here, we explored the roles of miRNAs in regulating male sterility of 337S at short day length/low temperature condition. RESULTS: Small RNA sequencing and degradome analyses were employed to identify miRNAs and their targets in the 337S whose meiotic cells collapsed rapidly during male meiotic prophase, resulting in failure of meiosis at SL condition. A total of 102 unique miRNAs were detected. Noticeably, the largest miRNA family was MiR1122. The target CCR4-associated factor 1 (CAF1) of miR2275, a subunit of the Carbon Catabolite Repressed 4-Negative on TATA-less (CCR4-NOT) complex, contributes to the process of early meiosis, and was first identified here. Further studies showed that the expression of several pivotal anther-related miRNAs was altered in 337S at SL condition, especially tae-miR1127a, which may be related to male sterility of 337S. Here, we also identified a new member of SWI/SNF factors SWI/SNF-related matrix-associated actin-dependent regulator of chromatin subfamily A, member 3-like 3 (SMARCA3L3) targeted by tae-miR1127a, whose function might be involved in faithful progression of meiosis in male reproductive cells. CONCLUSION: The miRNA-target interactions of tae-miR2275-CAF1 and tae-miR1127a-SMARCA3L3 might be involved in regulating male fertility in 337S. Our results also implied that multiple roles for SMARCA3L3 and CAF1 in DNA repair and transcriptional regulation jointly orchestrated a tight and orderly system for maintaining chromatin and genome integrity during meiosis.

Efficacy and Safety of Pulmonary Arterial Hypertension-specific Therapy in Pulmonary Arterial Hypertension: A Meta-analysis of Randomized Controlled Trials.

BACKGROUND: Previous meta-analyses of pulmonary arterial hypertension (PAH)-specific therapy for PAH pooled PAH-specific combination therapy and monotherapy. This flaw may threaten the authenticity of their findings. METHODS: PubMed, Embase, and the Cochrane Library were searched for randomized controlled trials that evaluated any PAH-specific medications in the treatment of PAH. We calculated ORs with 95% CIs for dichotomous data and standardized mean differences for continuous data. RESULTS: In total, 35 randomized controlled trials involving 6,702 patients were included. In monotherapy vs placebo/conventional therapy, significance was obtained in mortality reduction (OR, 0.50 [95% CI, 0.33 to 0.76]; P = .001), 6-min walk test (mean difference, 31.10 m [95% CI, 25.40 to 36.80]; P < .00001), New York Heart Association/World Health Organization functional class (OR, 2.48 [95% CI, 1.51 to 4.07]; P = .0003), and hemodynamic status based on mean pulmonary artery pressure, pulmonary vascular resistance, cardiac index, and incidence of withdrawal due to adverse effects. In combination therapy vs monotherapy, significance was reached for the 6-min walk test (mean difference, 19.96 m [95% CI, 15.35 to 24.57]; P < .00001), functional class (OR, 1.65 [95% CI, 1.20 to 2.28]; P = .002), hemodynamic status, and incidence of withdrawal due to adverse effects (OR, 2.01 [95% CI, 1.54 to 2.61]; P < .00001) but not for mortality reduction (OR, 0.98 [95% CI, 0.57 to 1.68]; P = .94). CONCLUSIONS: Our meta-analysis revealed that PAH-specific monotherapy could improve mortality, exercise capacity, functional class, and hemodynamic status compared with placebo or conventional therapy. However, combination therapy could further improve exercise capacity, functional class, and hemodynamic status compared with monotherapy, but it had no proven effect on mortality. Combination therapy had a much higher incidence of withdrawal due to adverse effects than monotherapy.

Combinations of Histone Modifications for Pattern Genes.

Histone post-translational modifications play important roles in transcriptional regulation. It is known that multiple histone modifications can act in a combinatorial manner. In this study, we investigated the effects of multiple histone modifications on expression levels of five gene categories (four kinds of pattern genes and non-pattern genes) in coding regions. The combinatorial patterns of modifications for the five gene categories were also studied in the regions. Our results indicated that the differences in the expression levels between any two gene categories were significant. There were some corresponding differences in multiple histone modification levels among the five gene categories. Multiple histone modifications jointly impacted expression levels of every gene category. Four mutual combinations of histone modifications were found and analyzed.

Influence factors on the correlations between expression levels of neighboring pattern genes.

Some genes tend to cluster and be co-expressed. Multiple factors affect gene co-expression. In this study, we investigated the relationships between multiple factors and the correlations of expression levels of neighboring genes, which were divided into four kinds of pattern genes and one type of non-pattern gene. Our results indicate that the correlation between expression levels of neighboring non-pattern genes is related to multiple factors with the exception of transcriptional orientations of neighboring genes. The correlation between expression levels of neighboring specific genes or neighboring repressed genes is likely to be dependent on the co-functions of neighboring genes. The correlation between expression levels of neighboring housekeeping genes is associated with histone modifications in intergenic regions.

[Role of calcineurin in down-regulation of left ventricular transmural voltage- dependent K(+) currents in mice with heart failure].

The aim of the present study was to investigate the role of calcineurin in the down-regulation of left ventricular transmural voltage-dependent K(+) currents in heart failure. Transverse aorta was banded by using microsurgical techniques to create mouse heart failure model. Sham-operated (Sham) or aorta banded (Band) mice were randomized to receive calcineurin inhibitor cyclosporine A (CsA) or vehicle. The densities and kinetic properties of voltage-dependent K(+) currents, as well as action potential (AP), of left ventricular subendocardial (Endo) and subepicardial (Epi) myocytes were determined by using whole-cell patch-clamp technique. The results showed that calcineurin activity was significant higher in Endo myocytes than that in Epi ones in all the groups. Compared with Sham group, Band mice showed significantly increased calcineurin activity both in Endo and Epi myocytes. CsA significantly reduced calcineurin activity in Band mice. CsA treatment in Band mice partially reversed the down-regulation of Ito density, completely reversed the down-regulation of IK,slow density both in Endo and Epi myocytes, and Iss density in Endo myocytes. In addition, CsA treatment in Band mice partially antagonized the prolongation of action potential duration (APD), and APD at 50% (APD50) and 90% repolarization (APD90) were significantly reduced. Because of non-parallel shortening of APD in Endo and Epi myocytes, the ratio of Endo/Epi APD90 was reduced from 4.8:1 in Band mice to 2.6:1 in CsA-treated mice, which was close to that in Sham mice. The results suggest that non-parallel activation of calcineurin in Endo and Epi myocytes contributes to the down-regulation of transmural voltage-dependent K(+) currents and the amplification of transmural dispersion of repolarization (TDR) in left ventricular failure hearts. Inhibition of calcineurin may be a potential new therapeutic strategy to prevent and cure arrhythmias and sudden death in heart failure.

Compound ICA-105574 prevents arrhythmias induced by cardiac delayed repolarization.

Impaired ventricular repolarization can lead to long QT syndrome (LQT), a proarrhythmic disease with high risk of developing lethal ventricular tachyarrhythmias. The compound ICA-105574 is a recently developed hERG activator and it enhances IKr current with very high potency by removing the channel inactivation. The present study was designed to investigate antiarrhythmic properties of ICA-105574. For comparison, the effects of another compound NS1643 was in-parallel assessed, which also acts primarily to attenuate channel inactivation with moderate potency. We found that both ICA-105574 and NS1643 concentration-dependently shortened action potential duration (APD) in ventricular myocytes, and QT/QTc intervals in isolated guinea-pig hearts. ICA-105574, but not NS1643, completely prevented ventricular arrhythmias in intact guinea-pig hearts caused by IKr and IKs inhibitors, although both ICA-105574 and NS1643 could reverse the drug-induced prolongation of APD in ventricular myocytes. Reversing prolongation of QT/QTc intervals and antagonizing the increases in transmural dispersion of repolarization and instability of the QT interval induced by IKr and IKs inhibitors contributed to antiarrhythmic effect of ICA-105574. Meanwhile, ICA-105574 at higher concentrations showed a potential proarrhythmic risk in normal hearts. Our results suggest that ICA-105574 has more efficient antiarrhythmic activity than NS1643. However, its potential proarrhythmic risk implies that benefits and risks should be seriously taken into consideration for further developing this type of hERG activators.

[Effect of increased posterior tibial slope or partial posterior cruciate ligament release on knee kinematics of total knee arthroplasty].

OBJECTIVE: To compare the effects of increased posterior tibial slope or partial posterior cruciate ligament (PCL) release on knee kinematics of total knee arthroplasty (TKA). METHODS: Anteroposterior laxity, rotational laxity, varus and valgus laxity and maximum flexion angle were evaluated in 6 normal cadaver knees and the knees after TKA at flexion 0 degrees , 30 degrees , 60 degrees , 90 degrees and 120 degrees . Then the femoral prosthesis was shifted 5 mm posteriorly to simulate the tightly implanted knee. The same tests were performed on the tightly implanted knees. After that, the posterior tibial slope was increased 4 degrees or the PCL was partially released, and the same tests were made as in the normal knees respectively. Statistical analysis of the results was made using student's t test. RESULTS: Anteroposterior laxity, rotational laxity and varus and valgus laxity of the tightly implanted knees at flexion 30 degrees , 60 degrees , 90 degrees and 120 degrees were significantly less than those of the normal TKA knees (P < 0.05). Compared with the tightly implanted knees, anteroposterior laxity, rotational laxity and varus and valgus laxity at flexion 30 degrees , 60 degrees , 90 degrees and 120 degrees significantly improved after increased 4 degrees posterior tibial slope (P < 0.05); in the partial PCL released group, anteroposterior laxity at flexion 30 degrees , 60 degrees , 90 degrees and 120 degrees was significantly improved (P < 0.05), varus and valgus laxity was significantly improved only at flexion 90 degrees (P < 0.05), and rotational laxity was significantly improved at flexion 30 degrees , 60 degrees and 90 degrees (P < 0.05). Compared with PCL released group, varus and valgus laxity at flexion 30 degrees , 60 degrees and 90 degrees and rotational laxity at flexion 0 degrees , 30 degrees , 60 degrees and 90 degrees were significantly improved in the group of increased 4 degrees posterior tibial slope (P < 0.05). Maximum flexion angle of the tightly implanted knee (120.4 degrees ) was less than that of the normal TKA knees (130.3 degrees , P < 0.05) and that of increased 4 degrees posterior tibial slope group (131.1 degrees , P < 0.05). There was no significant difference at the maximum flexion angle between the increased 4 degrees posterior tibial slope group and the PCL released group (131.1 degrees vs 124.0 degrees , P = 0.0816). CONCLUSIONS: Anteroposterior laxity, varus and valgus laxity, rotational laxity and maximum flexion angle of the tightly implanted knees are less than those of the normal TKA knees. After increased 4 degrees posterior tibial slope, these indexes are improved significantly. Partial PCL released can significantly improve the anteroposterior laxity and had less effect on the varus and valgus laxity, rotational laxity and maximum flexion angle. So, a knee that is tight in flexion can be more likely to be corrected by increasing posterior tibial slope than by partially releasing PCL.

[Transmural L-type calcium current in a pressure-overloaded mouse model with heart failure].

Transmural electrical heterogeneity plays an important role in the normal dispersion of repolarizaion and propagation of excitation in the heart. The amplification of transmural electrical heterogeneity contributes to the genesis of arrhythmias in cardiac hypertrophy and failure. We established a mouse model with cardiac failure by aortic banding and investigated the possible contribution of L-type calcium current (I(Ca-L)) to transmural electrical heterogeneity in both normal and failing hearts. Single myocytes were enzymatically isolated from subendocardial and subepicardial myocardium of the free left ventricle wall. The recordings of action potential and I(Ca-L) were performed using the conventional whole-cell patch-clamp technique. The results showed that: (1) The action potential duration at 90% repolarization (APD(90)) of the subendocardial myocytes in normal control mice was (38.2 +/- 6.44) ms, which was significantly longer than that of the subepicardial myocytes [(15.672 +/- 5.31) ms]. The ratio of APD(90) for subendocardial/subepicardial myocytes was about 2.5:1. The peak I(Ca-L) density in subendocardial myocytes was (-2.7 +/- 0.49) pA/pF, which was not different from that in subepicardial myocytes [(-2.54 +/- 0.53) pA/pF]. (2) In failing hearts, both action potential duration at 50% repolarization (APD(50)) and APD(90) were remarkably prolonged either in subendocardial or subepicardial myocytes compared to that in sham hearts. The subendocardial myocytes had much longer APD. The ratio of APD(90) for subendocardial/subepicardial myocytes changed to about 4.2:1. (3) I(Ca-L) density in subendocardial myocytes was significantly decreased in failing hearts compared with that in sham hearts. At four test potentials from +10 mV to +40 mV, the density of I(Ca-L) from subendocardial myocytes in failing hearts was decreased by 20.2%, 21.4%, 21.6% and 25.7%, respectively (P<0.01). However, no significant difference was observed in I(Ca-L) density from subepicardial myocytes in failing hearts. There was no significant difference in the kinetic properties of I(Ca-L) in subendocardial and subepicardial myocytes between the band and sham groups. We conclude that I(Ca-L) may not contribute to the physiological transmural electrical heterogeneity in mouse hearts. The electrical heterogeneity is exaggerated and the density of I(Ca-L) is decreased in the subendocardial myocytes, but not in the subepicardial myocytes in failing hearts. The results obtained suggest that the decreased density of I(Ca-L) in subendocardial myocytes is possibly an adaptive response to the prolongation of action potential due to delayed depolarization and may reduce the transmural dispersion of repolarization in heart failure.

Electrophysiologic effects of adenosine triphosphate on rabbit sinoatrial node pacemaker cells via P1 receptors.

AIM: To study the electrophysiologic effects of adenosine triphosphate (ATP) on rabbit sinoatrial node pacemaker cells and the receptors related with the action of ATP. METHODS: Intracellular microelectrode method was used to record the parameters of action potential (AP) in the rabbit sinoatrial nodes. RESULTS: ATP (0.1-3 mmol/L) decreased the rate of pacemaker firing (RPF) by 16 %-43 % and velocity of diastolic depolarization (VDD) by 33 %-67 %, increased the amplitude of AP (APA) by 6 %-9 % and maximal rate of depolarization (V(max)) by 30 %-76 %, shortened APD50 by 7 %-12 % and APD(90) by 6.3 %-9 %, concentration-dependently. The effects of ATP, adenosine (Ado), and adenosine diphosphate at the same concentration on AP were not different from each other significantly. Neither uridine triphosphate nor alpha,beta-methylene ATP had significant electrophysiologic effects on the sinoatrial node of rabbits. Both the electrophysiologic effects of ATP and Ado on pacemaker cells were inhibited by P1 receptor antagonist aminophylline 0.1 mmol/L (P<0.05) in a closely similar manner, and the effects of ATP were not affected by P2 receptor antagonist reactive blue 2 at 0.05 mmol/L (P>0.05). CONCLUSION: There are no functional P2X(1) and P2Y(2) receptors on pacemaker cells of the rabbit sinoatrial nodes, and the electrophysiologic effects of ATP in the rabbit sinoatrial node pacemaker cells are mediated via P1 receptors by Ado degraded from ATP.