Diabetes increases the risk of developing atrial fibrillation and especially young diabetes patients have a high relative risk. Increased focus on detecting atrial fibrillation in young diabetes patients might prove beneficial, and both anticoagulation treatment and anti-arrhythmic treatment strategies should be considered as soon as possible.

Background Phosphorylation is a key regulatory event in controlling the function of the cardiac gap junction protein connexin43 (Cx43). Recently, three new phosphorylation sites (S296, S297 and S306) were identified on Cx43; two of which (S297 and S306) are dephosphorylated during ischemia. The functional significance of these new sites is currently unknown. Objective To examine the role of S296, S297 and S306 in the regulation of electrical intercellular communication. Methods To mimic constitutive dephosphorylation serine was mutated to alanine at the three sites and expressed in HeLa cells. Electrical coupling and single-channel measurements were performed by double patch clamp; protein expression levels were assayed by western blotting, localization of Cx43 and phosphorylation of S306 by immunolabeling. Free hemichannels were assessed by biotinylation. Results Macroscopic conductance in cells expressing S306A was reduced to 57 % compared to WT while coupling was not significantly changed in either S296A or S297A expressing cells. S306A expressing cells displayed similar protein and free hemichannel abundance compared to WT-Cx43, whereas the fractional area of plaques in cell to cell interfaces was increased. However, single-channel measurements showed a WT-Cx43 main state conductance of 119 pS, whereas the main state conductance of S306A channels was reduced to 95 pS. Furthermore, channel gating was affected in S306A channels. Conclusions Lack of phosphorylation at serine 306 results in reduced coupling, which can be explained by reduced single channel conductance. We suggest that dephosphorylation of S306 partly explains the electrical uncoupling seen in myocardial ischemia.

Osmotic water permeability (P(f)) was studied in spheroid-shaped human airway epithelia explants derived from nasal polyps by the use of a new improved tissue collection and isolation procedure. The fluid-filled spheroids were lined with a single cell layer with the ciliated apical cell membrane facing the outside. They were capable of surviving hours of experiment involving continuous superfusion of the bathing medium and changes of osmolarity. A new image analysis technique was developed for measuring the spheroid diameters, giving high time and measurement resolutions. The transepithelial P(f), determined by the changes of the apical solution osmolarity, was not influenced by the presence of glucose, Na(+), or Na(+)/glucose-cotransport inhibitors in the bath, but was sensitive to the aquaporin (AQP) inhibitor HgCl(2). The measured P(f) levels and the values of activation energy were in the range of those seen in AQP-associated water transport. Together, these results indicate the presence of an AQP in the apical membrane of the spheroids. Notably, identical values for P(f) were found in CF and non-CF airway preparations, as was the case also for the calculated spontaneous fluid absorption rates.

Abstract-Gap junctions provide a low-resistance pathway for cardiac electric propagation. The role of GJ regulation in arrhythmia is unclear, partly because of limited availability of pharmacological tools. Recently, we showed that a peptide called "RXP-E" binds to the carboxyl terminal of connexin43 and prevents chemically induced uncoupling in connexin43-expressing N2a cells. Here, pull-down experiments show RXP-E binding to adult cardiac connexin43. Patch-clamp studies revealed that RXP-E prevented heptanol-induced and acidification-induced uncoupling in pairs of neonatal rat ventricular myocytes. Separately, RXP-E was concatenated to a cytoplasmic transduction peptide (CTP) for cytoplasmic translocation (CTP-RXP-E). The effect of RXP-E on action potential propagation was assessed by high-resolution optical mapping in monolayers of neonatal rat ventricular myocytes, containing Ϸ20% of randomly distributed myofibroblasts. In contrast to control experiments, when heptanol (2 mmol/L) was added to the superfusate of monolayers loaded with CTP-RXP-E, action potential propagation was maintained, albeit at a slower velocity. Similarly, intracellular acidification (pH i 6.2) caused a loss of action potential propagation in control monolayers; however, propagation was maintained in CTP-RXP-E-treated cells, although at a slower rate. Patch-clamp experiments revealed that RXP-E did not prevent heptanol-induced block of sodium currents, nor did it alter voltage dependence or amplitude of Kir2.1/Kir2.3 currents. RXP-E is the first synthetic molecule known to: (1) bind cardiac connexin43; (2) prevent heptanol and acidification-induced uncoupling of cardiac gap junctions; and (3) preserve action potential propagation among cardiac myocytes. RXP-E can be used to characterize the role of gap junctions in the function of multicellular systems, including the heart. (Circ Res. 2008;103:519-526.)Key Words: Cx43CT Ⅲ particle-receptor interaction Ⅲ gap junctions Ⅲ connexin43 Ⅲ rotigaptide C onnexins (Cxs) are integral membrane proteins that oligomerize to form intercellular channels called gap junctions (GJs). The most abundant GJ protein in a number of mammalian systems is Cx43. GJs allow passage of ions and small molecules between cells and are regulated by a variety of chemical interactions between the Cx molecule and the microenvironment. As such, GJs act as active filters to control passage of intercellular messages and modulate function.Our previous work has suggested that regulation of Cx43 results from the association of the carboxyl-terminal (CT) domain, acting as a gating particle, and a separate region of the Cx molecule acting as a receptor for the gating particle. 1,2 Additional studies have shown that this intramolecular particle-receptor interaction can be modulated by other intermolecular interactions in the microenvironment of the GJ plaque. 3-5 Based on the particle-receptor model, we reasoned that regulation of Cx43 could be disrupted by the binding of exogenous molecules to regions of the gating particle requi...

Gap junctions (GJ’s) provide a low resistance pathway for cardiac electrical propagation. The role of GJ regulation in arrhythmias is unclear, partly due to the limited availability of pharmacological tools. Recently, we showed that a peptide called “RXP-E” binds to the carboxyl terminal of connexin43 (Cx43) and prevents chemically-induced uncoupling in Cx43-expressing N2a cells. Here, we used pull-down experiments to show that RXP-E binds to adult cardiac Cx43. Patch-clamp studies revealed that RXP-E prevented heptanol-induced and acidification-induced uncoupling in pairs of neonatal rat ventricular myocytes (NRVM’s). Separately, RXP-E was concatenated to a cytoplasmic transduction peptide for translocation into the cytoplasm (construct CTP-RXP-E). The effect of RXP-E on action potential (AP) propagation was assessed by high resolution optical mapping in monolayers of NRVMs, containing approximately 20% of randomly distributed myofibroblasts. Conduction velocity (CV) was 164 ± 8mm/sec (avg±SEM; n=12; pacing frequency 2Hz) in untreated cells, and 158±10mm/sec, (n=6) and 180±7mm/sec (n=10) in monolayers treated with CTP-RXPE or a scrambled version of the peptide (CTP-Scr), respectively (pNS). Exposure of either untreated, or CTP-Scr-treated monolayers to heptanol caused propagation block. However, when heptanol (2 mmol/L) was added to the superfusate of monolayers loaded with CTP-RXP-E, AP propagation was maintained, albeit at a slower velocity (87±5mm/sec;n=4; P<0.001). Similarly, intracellular acidification (pHi=6.2) caused a loss of AP propagation in control or CTP-Scr monolayers; however, propagation was maintained in CTP-RXP-E treated cells, though at a slower rate (CV=93 ± 28mm/sec; n=4). Consistent with these results, patch clamp experiments revealed that RXP-E did not prevent heptanol-induced block of sodium or calcium currents, nor did it alter the voltage dependence or amplitude of Kir2.1/Kir2.3 currents. RXP-E is the first synthetic molecule known to: bind cardiac Cx43; prevent heptanol and acidification-induced uncoupling of cardiac GJ’s and preserve AP propagation among cardiac myocytes. RXP-E can be used to characterize the role of GJ’s in the function of multicellular systems, including the heart. This research has received full or partial funding support from the American Heart Association, AHA Founders Affiliate (Connecticut, Maine, Massachusetts, New Hampshire, New Jersey, New York, Rhode Island, Vermont).

Thapsigargin is a highly potent and selective inhibitor of sarco-endoplasmic reticulum (SERCA) family of Ca P+ -ATPases and a useful tool in research concerning the function of intracellular Ca P+ stores. We describe here a novel fluorescent derivative (8-O-(4-aminocinnamoyl)-8-O-debutanoylthapsigargin, termed ACTA) of this compound, acting as a Ca P+ -ATPase inhibitor with a potency approaching that of thapsigargin. Binding of ACTA to the skeletal muscle sarcoplasmic reticulum vesicles results in a strong fluorescence enhancement, approximately 66% of which depends on ACTA association with Ca P+ -ATPase. This specific component of ACTA fluorescence is sensitive to the E I -E P conformational equilibrium of the pump. The combined properties of high potency and binding-dependent fluorescence suggest ACTA to be a useful probe for a range of studies involving the SERCA class of ATPases.z 1998 Federation of European Biochemical Societies.

No abstract

Background In patients suspected of acute coronary syndrome, but where the coronary angiography (CAG) has shown unobstructed coronary arteries differential diagnoses include spontaneous coronary artery dissection and takotsubo cardiomyopathy. This case report presents a patient with spontaneous coronary artery dissection but diagnostic signs suspicious of takotsubo cardiomyopathy. Which leads to a consideration of the co-existence of the diseases. Case summary A 57-year-old woman was acutely admitted to the emergency ward with sudden development of chest discomfort, palpitations, and dyspnoea. At hospitalization, the electrocardiography showed T-wave inversions in I, aVL, and V2, and Troponin I was elevated. Initial echocardiography revealed apical akinesia consistent with takotsubo cardiomyopathy. Initially, a diagnosis of acute coronary syndrome or takotsubo cardiomyopathy was suspected. The patient was further diagnostically assessed with CAG including optical coherence tomography which showed spontaneous coronary artery dissection in the left anterior descending artery. At follow-up 3 months later, CAG showed a fully healed coronary artery, and repeated echocardiography showed normalization of the left ventricular function. Discussion In this case report, initially, acute coronary syndrome was suspected due to electrocardiography with T-wave inversions and elevated cardiac biomarkers. Takotsubo cardiomyopathy was suspected when echocardiography showed apical ballooning, but CAG with optical coherence tomography revealed a spontaneous coronary artery dissection. Interestingly no severe obstructions of coronary arteries were seen, and follow-up echocardiography showed fully regained myocardial function. This leads to the debate as to whether this might be a case of co-existing spontaneous coronary artery dissection and takotsubo cardiomyopathy.