Atrial fibrillation as a risk factor for exercise-induced pulmonary haemorrhage following a standardised exercise test.

BACKGROUND: Atrial fibrillation (AF) has been proposed as a risk factor for exercise-induced pulmonary haemorrhage (EIPH) due to increased pressure in the left atrium. OBJECTIVE: To evaluate if AF was associated with EIPH following a standardised exercise test (SET) to fatigue. STUDY DESIGN: Two-arm controlled experiment. METHODS: Ten untrained Standardbred mares mean (standard deviation [SD]) age 6 (2) years performed a SET on the treadmill in sinus rhythm (SR) (SET1) and 25-44 days after induction of self-sustained AF (SET2). AF was induced by tachypacing using a pacing device. Endoscopy, including tracheal wash and bronchoalveolar lavage (BAL), was performed 48-72 h before and 24 h after the two SETs. In addition, endoscopic grading of tracheal blood was performed 2 h after each SET. RESULTS: After SET1, none of the horses showed blood in the trachea, and two horses showed erythrophagocytosis. Following SET2, two horses had grade 1 blood in the trachea and free erythrocytes and erythrophagocytosis in the BAL, while another two horses had erythrophagocytosis in the BAL. In SET2, the overall performance on the treadmill was decreased with a lower maximum velocity (SET1 10.3 ± 0.8 m/s vs. SET2 8.9 ± 0.9 m/s, p = 0.004), a higher heart rate (284 ± 21 vs. 221 ± 18 bpm, p = 0.003) and more abnormal QRS complexes (p < 0.001) compared with SET1. CONCLUSIONS: Two horses showed signs of EIPH, resulting in visible blood in the trachea, when exercising in AF compared with SR. However, a possible link between EIPH, pulmonary pressure and AF needs to be further elucidated.

Increased fibroblast accumulation in the equine heart following persistent atrial fibrillation.

BACKGROUND: Fibroblasts maintain the extracellular matrix homeostasis and may couple to cardiomyocytes through gap junctions and thereby increase the susceptibility to slow conduction and cardiac arrhythmias, such as atrial fibrillation (AF). In this study, we used an equine model of persistent AF to characterize structural changes and the role of fibroblasts in the development of an arrhythmogenic substrate for AF. MATERIAL AND METHODS: Eleven horses were subjected to atrial tachypacing until self-sustained AF developed and were kept in AF for six weeks. Horses in sinus rhythm (SR) served as control. In terminal open-chest experiments conduction velocity (CV) was measured. Tissue was harvested and stained from selected sites. Automated image analysis was performed to assess fibrosis, fibroblasts, capillaries and various cardiomyocyte characteristics. RESULTS: Horses in SR showed a rate-dependent slowing of CV, while in horses with persistent AF this rate-dependency was completely abolished (CV•basic cycle length relation p = 0.0295). Overall and interstitial amounts of fibrosis were unchanged, but an increased fibroblast count was found in left atrial appendage, Bachmann's bundle, intraatrial septum and pulmonary veins (p < 0.05 for all) in horses with persistent AF. The percentage of α-SMA expressing fibroblasts remained the same between the groups. CONCLUSION: Persistent AF resulted in fibroblast accumulation in several regions, particularly in the left atrial appendage. The increased number of fibroblasts could be a mediator of altered electrophysiology during AF. Targeting the fibroblast proliferation and differentiation could potentially serve as a novel therapeutic target slowing down the structural remodeling associated with AF.

Vitrification yields higher cryo-survival rate than slow freezing in biopsied bovine in vitro produced blastocysts.

Vitrification and slow freezing are the two commonly used embryo cryopreservation methods. In most studies, vitrification of intact embryos has proven superior in several respects, including cell and embryo survival and pregnancy rate. However, there is a lack of data for comparing these two methods in in vitro produced (IVP) bovine blastocysts, which have been subjected to the retrieval of trophectoderm (TE) biopsy. Day 7 IVP blastocysts were pooled and randomized into four groups: 1) non-biopsy (NB), 2) biopsy (B), 3) biopsy-vitrification (BV), 4) biopsy-slow freeze (BSF). The blastocysts in the B, BV, and BSF groups were subjected to TE biopsy. For the B group, this was followed by 5 hours (h) incubation and subsequent scoring of the biopsy-survival (re-expansion) rate before processing for further analyses. For the BV and BSF groups, the biopsy procedure was followed by 2 h incubation, allowing for a quick re-expansion, after which the blastocysts were subjected to vitrification and slow freezing, respectively. After warming and thawing, respectively, they were then incubated for 5 h followed by scoring the cryo-survival (re-expansion) rates before processing for further analyses. These included quantification of ICM and TE cells, cleaved caspase-3- and TUNEL-positive cells, quantitative PCR on cellular stress markers (SOD1 and PRDX1), and ultrastructural analysis. The biopsy-survival rate in the B group was 94% (307/326). The cryo-survival rate in BV (86%, 138/161) was higher than that in BSF (57%, 81/142; P < 0.001). No differences were noted between the average ICM, TE, and total cell numbers of the groups. The percentages of cleaved caspase-3-positive cells were higher in BV vs. NB (P < 0.05), in BSF vs. NB (P < 0.001), and in BSF vs. B (P < 0.001). The percentages of TUNEL-positive cells were higher in BV vs. NB (P < 0.05) and in BSF vs. NB (P < 0.001). The levels of mRNA abundance for SOD1 and PRDX1 in B, BV, and BSF were not different from that in NB. The ultrastructural analysis of blastocysts in the BV and BSF groups showed distension of extracellular spaces and appearance of intracellular vacuoles in the ICM, distension of mitochondria, and disorganization of mitochondrial cristae in both ICM and TE, and weakened tight junctions between adjacent TE cells. In summary, our findings demonstrate that vitrification yields a higher cryo-survival rate than slow freezing in biopsied bovine IVP blastocysts. However, biopsy-vitrification and biopsy-slow-freeze values are comparable in terms of ICM, TE, and total blastocyst cell numbers, as well as cleaved caspase-3- and TUNEL-positive cell rates. Moreover, biopsy and cryopreservation performed alone had no effect on ICM, TE, total blastocyst cell numbers, or TUNEL-positive cell rates. Biopsy and vitrification performed alone had no effect on the cleaved caspase-3 positive cell rates, whereas slow freezing resulted in an increased rate. Furthermore, double traumatization with a combination of biopsy and cryopreservation, either vitrification or slow freezing, resulted in increased rates of cleaved caspase-3- and TUNEL-positive cells.

Inhibition of Small-Conductance Calcium-Activated Potassium Current (I K,Ca) Leads to Differential Atrial Electrophysiological Effects in a Horse Model of Persistent Atrial Fibrillation.

BACKGROUND: Small-conductance Ca2+-activated K+ (KCa2) channels have been proposed as a possible atrial-selective target to pharmacologically terminate atrial fibrillation (AF) and to maintain sinus rhythm. However, it has been hypothesized that the importance of the KCa2 current-and thereby the efficacy of small-conductance Ca2+-activated K+ current (I K,Ca) inhibition-might be negatively related to AF duration and the extent of AF-induced remodeling. EXPERIMENTAL APPROACH AND METHODS: To address the hypothesis of the efficacy of I K,Ca inhibition being dependent on AF duration, the anti-arrhythmic properties of the I K,Ca inhibitor NS8593 (5 mg/kg) and its influence on atrial conduction were studied using epicardial high-density contact mapping in horses with persistent AF. Eleven Standardbred mares with tachypacing-induced persistent AF (42 ± 5 days of AF) were studied in an open-chest experiment. Unipolar AF electrograms were recorded and isochronal high-density maps analyzed to allow for the reconstruction of wave patterns and changes in electrophysiological parameters, such as atrial conduction velocity and AF cycle length. Atrial anti-arrhythmic properties and adverse effects of NS8593 on ventricular electrophysiology were evaluated by continuous surface ECG monitoring. RESULTS: I K,Ca inhibition by NS8593 administered intravenously had divergent effects on right and left AF complexity and propagation properties in this equine model of persistent AF. Despite global prolongation of AF cycle length, a slowing of conduction in the right atrium led to increased anisotropy and electrical dissociation, thus increasing AF complexity. In contrast, there was no significant change in AF complexity in the LA, and cardioversion of AF was not achieved. CONCLUSIONS: Intra-atrial heterogeneity in response to I K,Ca inhibition by NS8593 was observed. The investigated dose of NS8593 increased the AF cycle length but was not sufficient to induce cardioversion. In terms of propagation properties during AF, I K,Ca inhibition by NS8593 led to divergent effects in the right and left atrium. This divergent behavior may have impeded the cardioversion success.

Thoracotomy and Pericardiotomy for Access to the Heart in Horses: Surgical Procedure and Effects on Anesthetic Variables.

Thoracotomy is an uncommon procedure in horses but remains essential in a variety of cases of pleuropneumonia, pericarditis, thoracic trauma or diaphragmatic herniation, and for experimental thoracic and cardiac procedures. This study aimed at developing an experimental surgical procedure allowing access to the entire circumference of the heart and describing the effect of thoracotomy on pulmonary gas exchange in these horses. The study consisted of two arms, arm one (undergoing thoracotomy), was a terminal experimental study that included 11 Standardbred horses with experimentally induced (by tachypacing) atrial fibrillation. Arm two consisted of 6 Standardbred horses undergoing anesthesia for reasons unrelated to the present study. These horses functioned as controls. Anesthesia was induced using zolazepam and tiletamine. Anesthesia was maintained with isoflurane in 100% oxygen and ventilation with intermittent positive pressure (IPPV); no positive end-expiratory pressure (PEEP) was performed. Rib resection and pericardiotomy were performed for complete exposure of the entire circumference of the heart. Arterial blood samples were collected prior to, 5 and 30 minutes after puncture of pleura parietalis. In 10 horses, resection of the fifth rib was adequate for exposure of the heart. In one horse, removal of the sixth rib was also necessary. The duration of the surgical procedure (thoracotomy, pericardiotomy) was < 45 minutes. During a thoracotomy, PaO2 decreased significantly (P < .05) from 291.8 ± 82.8 mmHg to 165.2 ± 73.5 mmHg but was not different from normal anesthetized controls. The PaCO2 remained within normal limits. This surgical approach provided access to the entire circumference of the heart.

Detection of atrial fibrillation with implantable loop recorders in horses.

BACKGROUND: Cardiac arrhythmias in horses are diagnosed by auscultation or electrocardiogram (ECG), which results in a low sensitivity for detecting arrhythmias that occur sporadically. Implantable loop recorders (ILRs) are small ECG devices placed subcutaneously, to automatically detect arrhythmias in human patients. OBJECTIVES: To test ILRs ability to detect atrial fibrillation (AF) in horses. Furthermore, we hypothesised that anatomical location of the implant site might influence signal quality. Signal quality was evaluated both during exercise and over time. STUDY DESIGN: Experimental study. METHODS: In five Standardbred mares, eleven ILRs were implanted subcutaneously in up to three different positions (Front: pectoral region, Left-6: sixth left intercostal space and Ventral: xiphoid region) and AF induced. The R- and T-wave amplitudes were measured in all positions over time during AF. AF burden automatically registered by the ILRs over a 2-month period was compared with selected Holter ECG recordings. RESULTS: All three positions had stable R- and T-wave amplitudes during the study period and were of sufficient quality to allow AF detection at rest. The position Left-6 showed significantly higher R- and T-wave amplitudes compared with the other positions. During submaximal exercise only the Left-6 position was able to record ECG signals of diagnostic quality. No position yielded diagnostic signals at maximum exercise due to artefacts. MAIN LIMITATIONS: Few horses and ILRs included and no spontaneous AF episodes were studied. CONCLUSIONS: This preliminary study indicates that ILRs can be used for AF detection in horses, but the anatomical location is important for optimal ECG quality. Despite insufficient quality during exercise, ILRs were suitable for AF detection at rest. Therefore, the ILR may be a valuable diagnostic tool for detecting paroxysmal AF in horses.

Effect of selective IK,ACh inhibition by XAF-1407 in an equine model of tachypacing-induced persistent atrial fibrillation.

BACKGROUND AND PURPOSE: Inhibition of the G-protein gated ACh-activated inward rectifier potassium current, IK,ACh may be an effective atrial selective treatment strategy for atrial fibrillation (AF). Therefore, the anti-arrhythmic and electrophysiological properties of a novel putatively potent and highly specific IK,ACh inhibitor, XAF-1407 (3-methyl-1-[5-phenyl-4-[4-(2-pyrrolidin-1-ylethoxymethyl)-1-piperidyl]thieno[2,3-d]pyrimidin-6-yl]azetidin-3-ol), were characterised for the first time in vitro and investigated in horses with persistent AF. EXPERIMENTAL APPROACH: The pharmacological ion channel profile of XAF-1407 was investigated using cell lines expressing relevant ion channels. In addition, eleven horses were implanted with implantable cardioverter defibrillators enabling atrial tachypacing into self-sustained AF. The electrophysiological effects of XAF-1407 were investigated after serial cardioversions over a period of 1 month. Cardioversion success, drug-induced changes of atrial tissue refractoriness, and ventricular electrophysiology were assessed at baseline (day 0) and days 3, 5, 11, 17, and 29 after AF induction. KEY RESULTS: XAF-1407 potently and selectively inhibited Kir 3.1/3.4 and Kir 3.4/3.4, underlying the IK,ACh current. XAF-1407 treatment in horses prolonged atrial effective refractory period as well as decreased atrial fibrillatory rate significantly (~20%) and successfully cardioverted AF, although with a decreasing efficacy over time. XAF-1407 shortened atrioventricular-nodal refractoriness, without effect on QRS duration. QTc prolongation (4%) within 15 min of drug infusion was observed, however, without any evidence of ventricular arrhythmia. CONCLUSION AND IMPLICATIONS: XAF-1407 efficiently cardioverted sustained tachypacing-induced AF of short duration in horses without notable side effects. This supports IK,ACh inhibition as a potentially safe treatment of paroxysmal AF in horses, suggesting potential clinical value for other species including humans.

Longitudinal study of electrical, functional and structural remodelling in an equine model of atrial fibrillation.

BACKGROUND: Large animal models are important in atrial fibrillation (AF) research, as they can be used to study the pathophysiology of AF and new therapeutic approaches. Unlike other animal models, horses spontaneously develop AF and could therefore serve as a bona fide model in AF research. We therefore aimed to study the electrical, functional and structural remodelling caused by chronic AF in a horse model. METHOD: Nine female horses were included in the study, with six horses tachypaced into self-sustained AF and three that served as a time-matched sham-operated control group. Acceleration in atrial fibrillatory rate (AFR), changes in electrocardiographic and echocardiographic variables and response to medical treatment (flecainide 2 mg/kg) were recorded over a period of 2 months. At the end of the study, changes in ion channel expression and fibrosis were measured and compared between the two groups. RESULTS: AFR increased from 299 ± 33 fibrillations per minute (fpm) to 376 ± 12 fpm (p < 0.05) and atrial function (active left atrial fractional area change) decreased significantly during the study (p < 0.05). No changes were observed in heart rate or ventricular function. The AF group had more atrial fibrosis compared to the control group (p < 0.05). No differences in ion channel expression were observed. CONCLUSION: Horses with induced AF show signs of atrial remodelling that are similar to humans and other animal models.

Time-dependent antiarrhythmic effects of flecainide on induced atrial fibrillation in horses.

BACKGROUND: Pharmacological treatment of atrial fibrillation (AF) in horses can be challenging because of low efficacy and adverse effects. Flecainide has been tested with variable efficacy. OBJECTIVE: To test whether the efficacy of flecainide is dependent on AF duration. ANIMALS: Nine Standardbred mares. METHODS: Factorial study design. All horses were instrumented with a pacemaker and assigned to a control or an AF group. On day 0, all horses were in sinus rhythm and received 2 mg/kg flecainide IV. Atrial fibrillation subsequently was induced in the AF group by pacemaker stimulation. On days 3, 9, 27, and 55, flecainide was administered to all horses, regardless of heart rhythm. RESULTS: All horses in AF cardioverted to sinus rhythm on days 3 and 9. On day 27, 5/6 horses cardioverted, whereas only 2/6 cardioverted on day 55. The time from the start of flecainide infusion to cardioversion (range, 3-185 min, log transformed) showed linear correlation with the cumulative duration of AF (r2 = .80, P < .0001). Flecainide induced abnormal QRS complexes in 4/6 AF horses and 1/3 controls. A positive correlation was found between heart rate before flecainide infusion and number of abnormal QRS complexes (0.14, P < .05). One horse suffered from cardiac arrest and died after flecainide infusion. CONCLUSIONS AND CLINICAL IMPORTANCE: Flecainide is effective for cardioversion of short-term induced AF, but the effect decreases with AF duration. Controlling heart rate may minimize adverse effects caused by flecainide, but the drug should be used with great caution.

Effect of induced chronic atrial fibrillation on exercise performance in Standardbred trotters.

BACKGROUND: Atrial fibrillation (AF) is the most common arrhythmia affecting performance in horses. However, no previous studies have quantified the performance reduction in horses suffering from AF. OBJECTIVES: To quantify the effect of AF on maximum velocity (Vmax ), maximum heart rate (HRmax ), heart rate recovery (T100 ), hematologic parameters and development of abnormal QRS complexes. ANIMALS: Nine Standardbred trotters. METHODS: Two-arm controlled trial. Six horses had AF induced by means of a pacemaker and 3 served as sham-operated controls. All horses were subjected to an exercise test to fatigue before (SET1) and after (SET2) 2 months of AF or sham. The Vmax and HRmax were assessed using a linear mixed normal model. Abnormal QRS complexes were counted manually on surface ECGs. RESULTS: Atrial fibrillation resulted in a 1.56 m/sec decrease in Vmax (P < .0001). In the AF group, HRmax ± SD increased from 226 ± 11 bpm at SET1 to 311 ± 27 bpm at SET 2. The AF group had higher HRmax at SET2 compared with controls (P < .0001), whereas no difference between the control and AF groups was observed at SET1 (P = .96). Several episodes of wide complex tachycardia were observed during exercise in 3 of the AF horses during SET2. CONCLUSIONS AND CLINICAL IMPORTANCE: Atrial fibrillation resulted in a significant reduction in performance, an increase in HR and development of abnormal QRS complexes during exercise, which may be a risk factor for collapse or sudden cardiac death.