E. coli infection disrupts the epithelial barrier and activates intrinsic neurosecretory reflexes in the pig colon

This study aims to assess the barrier integrity and possible activation of enteric neural pathways associated with secretion and motility in the pig colon induced by an enterotoxigenic Escherichia coli (ETEC) challenge. 50 Danbred male piglets were used for this study. 16 were challenged with an oral dose of the ETEC strain F4+ 1.5 × 109 colony-forming unit. Colonic samples were studied 4- and 9-days post-challenge using both a muscle bath and Ussing chamber. Colonic mast cells were stained with methylene blue. In control animals, electrical field stimulation induced neurosecretory responses that were abolished by tetrodotoxin (10−6M) and reduced by the combination of atropine (10−4M) and α-chymotrypsin (10U/mL). Exogenous addition of carbachol, vasoactive intestinal peptide, forskolin, 5-HT, nicotine, and histamine produced epithelial Cl− secretion. At day 4 post-challenge, ETEC increased the colonic permeability. The basal electrogenic ion transport remained increased until day 9 post-challenge and was decreased by tetrodotoxin (10−6M), atropine (10−4M), hexamethonium (10−5M), and ondansetron (10−5M). In the muscle, electrical field stimulation produced frequency-dependent contractile responses that were abolished with tetrodotoxin (10−6M) and atropine (10−6M). Electrical field stimulation and carbachol responses were not altered in ETEC animals in comparison with control animals at day 9 post-challenge. An increase in mast cells, stained with methylene blue, was observed in the mucosa and submucosa but not in the muscle layer of ETEC-infected animals on day 9 post-challenge. ETEC increased the response of intrinsic secretory reflexes and produced an impairment of the colonic barrier that was restored on day 9 post-challenge but did not modify neuromuscular function

Electrophysiological and histological characterization of atrial scarring in a model of isolated atrial myocardial infarction

Background: Characterization of atrial myocardial infarction is hampered by the frequent concurrence of ventricular infarction. Theoretically, atrial infarct scarring could be recognized by multifrequency tissue impedance, like in ventricular infarction, but this remains to be proven. Objective: This study aimed at developing a model of atrial infarction to assess the potential of multifrequency impedance to recognize areas of atrial infarct scar. Methods: Seven anesthetized pigs were submitted to transcatheter occlusion of atrial coronary branches arising from the left coronary circumflex artery. Six weeks later the animals were anesthetized and underwent atrial voltage mapping and multifrequency impedance recordings. The hearts were thereafter extracted for anatomopathological study. Two additional pigs not submitted to atrial branch occlusion were used as controls. Results: Selective occlusion of the atrial branches induced areas of healed infarction in the left atrium in 6 of the 7 cases. Endocardial mapping of the left atrium showed reduced multi-frequency impedance (Phase angle at 307 kHz: from -17.1° ± 5.0° to -8.9° ± 2.6°, p < .01) and low-voltage of bipolar electrograms (.2 ± 0.1 mV vs. 1.9 ± 1.5 mV vs., p < .01) in areas affected by the infarction. Data variability of the impedance phase angle was lower than that of bipolar voltage (coefficient of variability of phase angle at307 kHz vs. bipolar voltage: .30 vs. .77). Histological analysis excluded the presence of ventricular infarction. Conclusion: Selective occlusion of atrial coronary branches permits to set up a model of selective atrial infarction. Atrial multifrequency impedance mapping allowed recognition of atrial infarct scarring with lesser data variability than local bipolar voltage mapping. Our model may have potential applicability on the study of atrial arrhythmia mechanisms.Peer ReviewedPostprint (published version

Beta-blocker treatment of patients with atrial fibrillation attenuates spontaneous calcium release-induced electrical activity

Aims Atrial fibrillation (AF) has been associated with excessive spontaneous calcium release, linked to cyclic AMP (cAMP)-dependent phosphorylation of calcium regulatory proteins. Because ß-blockers are expected to attenuate cAMP-dependent signaling, we aimed to examine whether the treatment of patients with ß-blockers affected the incidence of spontaneous calcium release events or transient inward currents (ITI). Methods The impact of treatment with commonly used ß-blockers was analyzed in human atrial myocytes from 371 patients using patch-clamp technique, confocal calcium imaging or immunofluorescent labeling. Data were analyzed using multivariate regression analysis taking into account potentially confounding effects of relevant clinical factors Results The L-type calcium current (ICa) density was diminished significantly in patients with chronic but not paroxysmal AF and the treatment of patients with ß-blockers did not affect ICa density in any group. By contrast, the ITI frequency was elevated in patients with either paroxysmal or chronic AF that did not receive treatment, and ß-blocker treatment reduced the frequency to levels observed in patients without AF. Confocal calcium imaging showed that ß-blocker treatment also reduced the calcium spark frequency in patients with AF to levels observed in those without AF. Furthermore, phosphorylation of the ryanodine receptor (RyR2) at Ser-2808 and phospholamban at Ser-16 was significantly lower in patients with AF that received ß-blockers. Conclusion Together, our findings demonstrate that ß-blocker treatment may be of therapeutic utility to prevent spontaneous calcium release-induced atrial electrical activity; especially in patients with a history of paroxysmal AF displaying preserved ICa density.Peer ReviewedPostprint (published version

Influence of sex on intracellular calcium homoeostasis in patients with atrial fibrillation

Altres ajuts: Fundació Marato TV3 [20152030/31].Atrial fibrillation (AF) has been associated with intracellular calcium disturbances in human atrial myocytes, but little is known about the potential influence of sex and we here aimed to address this issue. Alterations in calcium regulatory mechanisms were assessed in human atrial myocytes from patients without AF or with long-standing persistent or permanent AF. Patch-clamp measurements revealed that L-type calcium current (I ) density was significantly smaller in males with than without AF (−1.15 ± 0.37 vs. −2.06 ± 0.29 pA/pF) but not in females with AF (−1.88 ± 0.40 vs. −2.21 ± 0.0.30 pA/pF). In contrast, transient inward currents (I ) were more frequent in females with than without AF (1.92 ± 0.36 vs. 1.10 ± 0.19 events/min) but not in males with AF. Moreover, confocal calcium imaging showed that females with AF had more calcium spark sites than those without AF (9.8 ± 1.8 vs. 2.2 ± 1.9 sites/µm 2) and sparks were wider (3.0 ± 0.3 vs. 2.2 ± 0.3 µm) and lasted longer (79 ± 6 vs. 55 ± 8 ms), favouring their fusion into calcium waves that triggers I s and afterdepolarizations. This was linked to higher ryanodine receptor phosphorylation at s2808 in women with AF, and inhibition of adenosine A or beta-adrenergic receptors that modulate s2808 phosphorylation was able to reduce the higher incidence of I in women with AF. Perturbations of the calcium homoeostasis in AF is sex-dependent, concurring with increased spontaneous SR calcium release-induced electrical activity in women but not in men, and with diminished I density in men only. This work was supported by grants from The Spanish Ministry of Science Innovation and Universities

Supporting Information ß2-adrenergic stimulation potentiates spontaneous calcium release by increasing signal mass and co-activation of ryanodine receptor clusters

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Alteraciones en la homeostasis del calcio y la actividad eléctrica que se asocian con la fibrilación auricular en miocitos auriculares humanos y un modelo porcino de infarto auricular

La fibril·lació auricular (FA) s'ha associat a anomalies estructurals, moleculars i electrofisiològiques a l'aurícula. El sistema nerviós autònom té un paper molt rellevant en l'inici i el manteniment d'aquesta malaltia, ja que pot induir canvis significatius i heterogenis en l'electrofisiologia auricular. S'ha demostrat que els mètodes que redueixen la innervació autònoma a l'aurícula redueixen la incidència d'arrítmies, per la qual cosa la neuromodulació pot ser útil per controlar la FA. A més, s'ha observat un augment en la densitat de la innervació simpàtica en mostres auriculars de pacients amb FA. Aquest augment del to simpàtic és el resultat de l'activació dels receptors [Beta]-adrenèrgics acoblats a una proteïna Gs, que estimula adenilil ciclasa augmentant els nivells d'AMP cíclic amb la posterior activació de la proteïna quinasa A. Aquesta última activa proteïnes que s'encarreguen de regular l'homeòstasi del calci com el receptor de rianodina de tipus 2 (RyR2). La hiperfosforilació d'aquest receptor és de gran importància, ja que s'ha relacionat amb un increment en l'alliberament espontani de calci des del reticle sarcoplasmàtic. A la clínica, s'usen antagonistes dels receptors [Beta]-adrenèrgics com a tractament per a pacients amb arrítmies auriculars, sobretot per controlar la freqüència cardíaca dels pacients. Tot i això, aquests fàrmacs també modifiquen l'activació de RyR2 prevenint l'activitat espontània i les arrítmies ventriculars, suggerint que també podrien prevenir l'alliberament espontani de calci i reduir la incidència de postdespolaritzacions en pacients amb FA. Un altre factor que pot contribuir a la generació d'arrítmies auriculars és l'infart auricular (IA). Aquest passa quan s'oclueixen branques coronàries auriculars que normalment provenen dels segments proximals de les artèries coronàries dreta o circumflexa esquerra. A més, el seu diagnòstic, a partir de l'electrocardiograma, és difícil i podria ser degut als sistemes de registre convencionals o al baix impacte de l'oclusió de la branca auricular a l'estructura miocàrdica local i l'electrofisiologia. A la clínica, els canvis a l'ona P i la desviació del segment PR són criteris que permeten sospites d'IA, però són inconstants i poden requerir un ECG de referència. A més, se sap que les alteracions electrofisiològiques intrínseques induïdes per l'infart de miocardi auricular aïllat provoquen alentiment de la conducció auricular local, prolongació del període refractari i canvis en el segment ST i el voltatge dels electrogrames auriculars locals, substrat que podria propiciar les arrítmies. Actualment, la caracterització electrofisiològica de la cicatrització de l'IA i la seva capacitat per induir arrítmies auriculars es veu obstaculitzada per l'escassetat de models animals que imiten aquesta entitat clínica.La fibrilación auricular (FA) se ha asociado a anomalías estructurales, moleculares y electrofisiológicas en la aurícula. El sistema nervioso autónomo desempaña un papel muy relevante en el inicio y mantenimiento de esta enfermedad ya que puede inducir cambios significativos y heterogéneos en la electrofisiología auricular. Se ha demostrado que los métodos que reducen la inervación autónoma en la aurícula reducen la incidencia de arritmias por lo que la neuromodulación puede ser útil para controlar la FA. Además, se ha observado un aumento en la densidad en la inervación simpática en muestras auriculares de pacientes con FA. Este aumento del tono simpática es el resultado de la activación de los receptores [Beta]-adrenérgicos acoplados a una proteína Gs, que estimula adenilil ciclasa aumentando los niveles de AMP cíclico con la posterior activación de la proteína quinasa A. Ésta última, activa proteínas que se encargan de regular la homeostasis del calcio como el receptor de rianodina de tipo 2 (RyR2). La hiperfosforilación de este receptor es de gran importancia ya que ésta se ha relacionado con un incremento en la liberación espontánea de calcio desde el retículo sarcoplasmático. En la clínica, se usan antagonistas de los receptores [Beta]-adrenérgicos como tratamiento para pacientes con arritmias auriculares, sobre todo para controlar la frecuencia cardíaca de los pacientes. Sin embargo, estos fármacos también modifican la activación de RyR2 previniendo la actividad espontánea y las arritmias ventriculares, sugiriendo que también podrían prevenir la liberación espontánea de calcio y reducir la incidencia de postdespolarizaciones en pacientes con FA. Otro factor que puede contribuir en la generación de arritmias auriculares es el infarto auricular (IA). Éste ocurre cuando se ocluyen ramas coronarias auriculares que normalmente provienen de los segmentos proximales de las arterias coronarias derecha o circunfleja izquierda. Además, su diagnóstico, a partir del electrocardiograma (ECG), es difícil y podría deberse a los sistemas de registro convencionales o al bajo impacto de la oclusión de la rama auricular en la estructura miocárdica local y la electrofisiología. En la clínica, los cambios en la onda P y la desviación del segmento PR son criterios que permiten sospechas de IA, pero son inconstantes y pueden requerir de un ECG de referencia. Además, se sabe que las alteraciones electrofisiológicas intrínsecas inducidas por el infarto de miocardio auricular aislado provocan ralentización de la conducción auricular local, prolongación del período refractario y cambios en el segmento ST y el voltaje de los electrogramas auriculares locales, sustrato que podría propiciar las arritmias. Actualmente, la caracterización electrofisiológica de la cicatrización del IA, y su capacidad para inducir arritmias auriculares se ve obstaculizada por la escasez de modelos animales que imiten esta entidad clínica.Atrial fibrillation (AF) has been associated with structural, molecular, and electrophysiological abnormalities in the atrium. The autonomic nervous system plays a crucial role in the onset and maintenance of this disease, as it can induce significant and heterogeneous changes in atrial electrophysiology. Methods that decrease autonomic innervation in the atrium have been shown to reduce the incidence of arrhythmias, making neuromodulation a potential approach to control AF. Furthermore, there has been a noted increase in sympathetic nerve density in atrial samples taken from patients with AF. This heightened sympathetic tone results from the activation of [Beta]-adrenergic receptors coupled to a Gs protein, which stimulates adenylate cyclase and increases cyclic AMP levels, leading to the subsequent activation of protein kinase A (PKA). The PKA activates proteins responsible for regulating calcium homeostasis, such as the type 2 ryanodine receptor (RyR2). Hyperphosphorylation of this receptor is of significant importance, as it has been linked to increased spontaneous calcium release from the sarcoplasmic reticulum (SR). In clinical practice, [Beta]-adrenergic receptor antagonists are used to treat patients with atrial arrhythmias, mainly to control their heart rate. However, these drugs also modify RyR2 activation, which prevents spontaneous activity and ventricular arrhythmias. This suggests that such drugs may also prevent spontaneous calcium release and reduce the incidence of afterdepolarizations in patients with AF. Another factor that may contribute to the generation of atrial arrhythmias is atrial infarction (AI). AIs occur when atrial coronary branches that normally arise from the proximal segments of the left circumflex or right arteries are occluded. Moreover, diagnosing AIs based on electrocardiograms (ECG) is challenging. This might be due to the use of conventional recording systems or to the limited impact that atrial branch occlusion has on the local myocardial structure and electrophysiology. In clinical practice, changes in the P-wave and PR segment deviation are indicators of a potential AI, but they are inconsistent and may require a reference ECG. Additionally, intrinsic electrophysiological alterations induced by isolated atrial myocardial infarction are known to lead to local atrial conduction slowing, prolonged refractory periods, and changes in the ST segment and in the voltage of local atrial electrograms, which could create a substrate for arrhythmias. Currently, the electrophysiological characterization of AI scarring and its potential to induce atrial arrhythmias is hindered by the scarcity of animal models that mimic this clinical entity

Alteraciones en la homeostasis del calcio y la actividad eléctrica que se asocian con la fibrilación auricular en miocitos auriculares humanos y un modelo porcino de infarto auricular

La fibril·lació auricular (FA) s'ha associat a anomalies estructurals, moleculars i electrofisiològiques a l'aurícula. El sistema nerviós autònom té un paper molt rellevant en l'inici i el manteniment d'aquesta malaltia, ja que pot induir canvis significatius i heterogenis en l'electrofisiologia auricular. S'ha demostrat que els mètodes que redueixen la innervació autònoma a l'aurícula redueixen la incidència d'arrítmies, per la qual cosa la neuromodulació pot ser útil per controlar la FA. A més, s'ha observat un augment en la densitat de la innervació simpàtica en mostres auriculars de pacients amb FA. Aquest augment del to simpàtic és el resultat de l'activació dels receptors [Beta]-adrenèrgics acoblats a una proteïna Gs, que estimula adenilil ciclasa augmentant els nivells d'AMP cíclic amb la posterior activació de la proteïna quinasa A. Aquesta última activa proteïnes que s'encarreguen de regular l'homeòstasi del calci com el receptor de rianodina de tipus 2 (RyR2). La hiperfosforilació d'aquest receptor és de gran importància, ja que s'ha relacionat amb un increment en l'alliberament espontani de calci des del reticle sarcoplasmàtic. A la clínica, s'usen antagonistes dels receptors [Beta]-adrenèrgics com a tractament per a pacients amb arrítmies auriculars, sobretot per controlar la freqüència cardíaca dels pacients. Tot i això, aquests fàrmacs també modifiquen l'activació de RyR2 prevenint l'activitat espontània i les arrítmies ventriculars, suggerint que també podrien prevenir l'alliberament espontani de calci i reduir la incidència de postdespolaritzacions en pacients amb FA. Un altre factor que pot contribuir a la generació d'arrítmies auriculars és l'infart auricular (IA). Aquest passa quan s'oclueixen branques coronàries auriculars que normalment provenen dels segments proximals de les artèries coronàries dreta o circumflexa esquerra. A més, el seu diagnòstic, a partir de l'electrocardiograma, és difícil i podria ser degut als sistemes de registre convencionals o al baix impacte de l'oclusió de la branca auricular a l'estructura miocàrdica local i l'electrofisiologia. A la clínica, els canvis a l'ona P i la desviació del segment PR són criteris que permeten sospites d'IA, però són inconstants i poden requerir un ECG de referència. A més, se sap que les alteracions electrofisiològiques intrínseques induïdes per l'infart de miocardi auricular aïllat provoquen alentiment de la conducció auricular local, prolongació del període refractari i canvis en el segment ST i el voltatge dels electrogrames auriculars locals, substrat que podria propiciar les arrítmies. Actualment, la caracterització electrofisiològica de la cicatrització de l'IA i la seva capacitat per induir arrítmies auriculars es veu obstaculitzada per l'escassetat de models animals que imiten aquesta entitat clínica.La fibrilación auricular (FA) se ha asociado a anomalías estructurales, moleculares y electrofisiológicas en la aurícula. El sistema nervioso autónomo desempaña un papel muy relevante en el inicio y mantenimiento de esta enfermedad ya que puede inducir cambios significativos y heterogéneos en la electrofisiología auricular. Se ha demostrado que los métodos que reducen la inervación autónoma en la aurícula reducen la incidencia de arritmias por lo que la neuromodulación puede ser útil para controlar la FA. Además, se ha observado un aumento en la densidad en la inervación simpática en muestras auriculares de pacientes con FA. Este aumento del tono simpática es el resultado de la activación de los receptores [Beta]-adrenérgicos acoplados a una proteína Gs, que estimula adenilil ciclasa aumentando los niveles de AMP cíclico con la posterior activación de la proteína quinasa A. Ésta última, activa proteínas que se encargan de regular la homeostasis del calcio como el receptor de rianodina de tipo 2 (RyR2). La hiperfosforilación de este receptor es de gran importancia ya que ésta se ha relacionado con un incremento en la liberación espontánea de calcio desde el retículo sarcoplasmático. En la clínica, se usan antagonistas de los receptores [Beta]-adrenérgicos como tratamiento para pacientes con arritmias auriculares, sobre todo para controlar la frecuencia cardíaca de los pacientes. Sin embargo, estos fármacos también modifican la activación de RyR2 previniendo la actividad espontánea y las arritmias ventriculares, sugiriendo que también podrían prevenir la liberación espontánea de calcio y reducir la incidencia de postdespolarizaciones en pacientes con FA. Otro factor que puede contribuir en la generación de arritmias auriculares es el infarto auricular (IA). Éste ocurre cuando se ocluyen ramas coronarias auriculares que normalmente provienen de los segmentos proximales de las arterias coronarias derecha o circunfleja izquierda. Además, su diagnóstico, a partir del electrocardiograma (ECG), es difícil y podría deberse a los sistemas de registro convencionales o al bajo impacto de la oclusión de la rama auricular en la estructura miocárdica local y la electrofisiología. En la clínica, los cambios en la onda P y la desviación del segmento PR son criterios que permiten sospechas de IA, pero son inconstantes y pueden requerir de un ECG de referencia. Además, se sabe que las alteraciones electrofisiológicas intrínsecas inducidas por el infarto de miocardio auricular aislado provocan ralentización de la conducción auricular local, prolongación del período refractario y cambios en el segmento ST y el voltaje de los electrogramas auriculares locales, sustrato que podría propiciar las arritmias. Actualmente, la caracterización electrofisiológica de la cicatrización del IA, y su capacidad para inducir arritmias auriculares se ve obstaculizada por la escasez de modelos animales que imiten esta entidad clínica.Atrial fibrillation (AF) has been associated with structural, molecular, and electrophysiological abnormalities in the atrium. The autonomic nervous system plays a crucial role in the onset and maintenance of this disease, as it can induce significant and heterogeneous changes in atrial electrophysiology. Methods that decrease autonomic innervation in the atrium have been shown to reduce the incidence of arrhythmias, making neuromodulation a potential approach to control AF. Furthermore, there has been a noted increase in sympathetic nerve density in atrial samples taken from patients with AF. This heightened sympathetic tone results from the activation of [Beta]-adrenergic receptors coupled to a Gs protein, which stimulates adenylate cyclase and increases cyclic AMP levels, leading to the subsequent activation of protein kinase A (PKA). The PKA activates proteins responsible for regulating calcium homeostasis, such as the type 2 ryanodine receptor (RyR2). Hyperphosphorylation of this receptor is of significant importance, as it has been linked to increased spontaneous calcium release from the sarcoplasmic reticulum (SR). In clinical practice, [Beta]-adrenergic receptor antagonists are used to treat patients with atrial arrhythmias, mainly to control their heart rate. However, these drugs also modify RyR2 activation, which prevents spontaneous activity and ventricular arrhythmias. This suggests that such drugs may also prevent spontaneous calcium release and reduce the incidence of afterdepolarizations in patients with AF. Another factor that may contribute to the generation of atrial arrhythmias is atrial infarction (AI). AIs occur when atrial coronary branches that normally arise from the proximal segments of the left circumflex or right arteries are occluded. Moreover, diagnosing AIs based on electrocardiograms (ECG) is challenging. This might be due to the use of conventional recording systems or to the limited impact that atrial branch occlusion has on the local myocardial structure and electrophysiology. In clinical practice, changes in the P-wave and PR segment deviation are indicators of a potential AI, but they are inconsistent and may require a reference ECG. Additionally, intrinsic electrophysiological alterations induced by isolated atrial myocardial infarction are known to lead to local atrial conduction slowing, prolonged refractory periods, and changes in the ST segment and in the voltage of local atrial electrograms, which could create a substrate for arrhythmias. Currently, the electrophysiological characterization of AI scarring and its potential to induce atrial arrhythmias is hindered by the scarcity of animal models that mimic this clinical entity.Universitat Autònoma de Barcelona. Programa de Doctorat en Neurocièncie

ß2-adrenergic stimulation potentiates spontaneous calcium release by increasing signal mass and co-activation of ryanodine receptor clusters

Aims It is unknown how ß-adrenergic stimulation affects calcium dynamics in individual RyR2 clusters and leads to the induction of spontaneous calcium waves. To address this, we analysed spontaneous calcium release events in green fluorescent protein (GFP)-tagged RyR2 clusters. Methods Cardiomyocytes from mice with GFP-tagged RyR2 or human right atrial tissue were subjected to immunofluorescent labelling or confocal calcium imaging. Results Spontaneous calcium release from single RyR2 clusters induced 91.4% ± 2.0% of all calcium sparks while 8.0% ± 1.6% were caused by release from two neighbouring clusters. Sparks with two RyR2 clusters had 40% bigger amplitude, were 26% wider, and lasted 35% longer at half maximum. Consequently, the spark mass was larger in two- than one-cluster sparks with a median and interquartile range for the cumulative distribution of 15.7 ± 20.1 vs 7.6 ± 5.7 a.u. (P < .01). ß2-adrenergic stimulation increased RyR2 phosphorylation at s2809 and s2815, tripled the fraction of two- and three-cluster sparks, and significantly increased the spark mass. Interestingly, the amplitude and mass of the calcium released from a RyR2 cluster were proportional to the SR calcium load, but the firing rate was not. The spark mass was also higher in 33 patients with atrial fibrillation than in 36 without (22.9 ± 23.4 a.u. vs 10.7 ± 10.9; P = .015). Conclusions Most sparks are caused by activation of a single RyR2 cluster at baseline while ß-adrenergic stimulation doubles the mass and the number of clusters per spark. This mimics the shift in the cumulative spark mass distribution observed in myocytes from patients with atrial fibrillation.Postprint (published version

Changes in Local Atrial Electrograms and Surface ECG Induced by Acute Atrial Myocardial Infarction

Atrial coronary branch occlusion is a hardly recognizable clinical entity that can promote atrial fibrillation. The low diagnostic accuracy of the ECG could deal with the characteristics of the ischemia-induced changes in local atrial electrograms, but these have not been described. We analyzed the effects of selective acute atrial branch occlusion on local myocardial structure, atrial electrograms, and surface ECG in an experimental model close to human cardiac anatomy and electrophysiology. Six anesthetized open-chest anesthetized pigs underwent surgical occlusion of an atrial coronary branch arising from the right coronary artery during 4 h. Atrial electrograms and ECG were simultaneously recorded. One additional pig acted as sham control. In all cases, the hearts were processed for anatomopathological analysis. Atrial branch occlusion induced patchy atrial necrosis with sharp border zone. During the first 30 min of occlusion, atrial electrograms showed progressive R wave enlargement (1.8 ± 0.6 mV vs. 2.5 ± 1.1 mV, p < 0.01), delayed local activation times (28.5 ± 8.9 ms vs. 36.1 ± 16.4 ms, p < 0.01), ST segment elevation (−0.3 ± 0.3 mV vs. 1.0 ± 1.0 mV, p < 0.01), and presence of monophasic potentials. Atrial ST segment elevation decreased after 2 h of occlusion. The electrical border zone was ∼1 mm and expanded over time. After 2 h of occlusion, the ECG showed a decrease in P wave amplitude (from 0.09 ± 0.04 mV to 0.05 ± 0.04 mV after 165 min occlusion, p < 0.05) and duration (64.4 ± 8.0 ms vs. 80.9 ± 12.6 ms, p < 0.01). Selective atrial branch occlusion induces patchy atrial infarction and characteristic changes in atrial activation, R/S wave, and ST segment that are not discernible at the ECG. Only indirect changes in P wave amplitude and duration were appreciated in advanced stages of acute coronary occlusion

Impact of R-Carvedilol on β2-Adrenergic Receptor-Mediated Spontaneous Calcium Release in Human Atrial Myocytes

A hallmark of atrial fibrillation is an excess of spontaneous calcium release events, which can be mimicked by β1- or β2-adrenergic stimulation. Because β1-adrenergic receptor blockers (β1-blockers) are primarily used in clinical practice, we here examined the impact of β2-adrenergic stimulation on spontaneous calcium release and assessed whether the R- and S-enantiomers of the non-selective β- blocker carvedilol could reverse these effects. For this purpose, human atrial myocytes were isolated from patients undergoing cardiovascular surgery and subjected to confocal calcium imaging or immunofluorescent labeling of the ryanodine receptor (RyR2). Interestingly, the β2-adrenergic agonist fenoterol increased the incidence of calcium sparks and waves to levels observed with the non-specific β-adrenergic agonist isoproterenol. Moreover, fenoterol increased both the amplitude and duration of the sparks, facilitating their fusion into calcium waves. Subsequent application of the non β-blocking R-Carvedilol enantiomer reversed these effects of fenoterol in a dose-dependent manner. R-Carvedilol also reversed the fenoterol-induced phosphorylation of the RyR2 at Ser-2808 dose-dependently, and 1 µM of either R- or S-Carvedilol fully reversed the effect of fenoterol. Together, these findings demonstrate that β2-adrenergic stimulation alone stimulates RyR2 phosphorylation at Ser-2808 and spontaneous calcium release maximally, and points to carvedilol as a tool to attenuate the pathological activation of β2-receptors.Medicine, Faculty ofNon UBCReviewedFacultyResearche