INTRODUCTION:The electrophysiologic basis for characteristic rate-dependent, constant-late-coupled (390 + 54 milliseconds) premature ventricular beats (PVBs) present 4-5 days following coronary artery occlusion were examined in 108 anesthetized dogs. METHODS AND RESULTS:Fractionated/double potentials were observed in injured zone bipolar and composite electrograms at prolonged sinus cycle lengths (1,296 ± 396 milliseconds). At shorter cycle lengths, conduction of the delayed potential decremented, separating from the initial electrogram by a progressively prolonged isoelectric interval. With sufficient delay of the second potential following an isoelectric interval, a PVB was initiated. Both metastable and stable constant-coupled PVBs were associated with Wenckebach-like patterns of delayed activation following an isoelectric interval. Signal-averaging from the infarct border confirmed the presence of an isoelectric interval preceding the PVBs (N = 15). Pacing from the site of double potential formation accurately reproduced the surface ECG morphology (N = 15) of spontaneous PVBs. Closely-spaced epicardial mapping demonstrated delayed activation across an isoelectric interval representing "an arc of conduction block." Rate-dependent very slow antegrade conduction through a zone of apparent conduction block (N = 8) produced decremental activation delays until the delay was sufficient to excite epicardium distal to the original "arc of conduction block," resulting in PVB formation. CONCLUSION:The present experiments demonstrate double potential formation and rate-dependent constant-coupled late PVB formation in infarcted dog hearts. Electrode recordings demonstrate a prolonged isoelectric period preceding PVB formation consistent with very slow conduction (<70 mm/s) across a line of apparent conduction block and may represent a new mechanism of PVB formation following myocardial infarction.

The electrophysiologic mechanism for rate‐dependent PVBs associated with double potentials (DPs) was investigated in infarcted canine hearts using bipolar and intracellular microelectrode recordings.

OBJECTIVES We investigated the hypothesis that re-entrant pulmonary vein (PV) tachycardias may serve as a mechanism for initiating and sustaining paroxysmal atrial fibrillation (PAF). BACKGROUND The mechanisms of rapid repetitive discharges from the PV initiating PAF remain incompletely understood. Pulmonary vein myocardial sleeves appear to provide a favorable substrate for re-entry formation. METHODS The electrophysiologic properties of canine PV sleeves were investigated using a combination of high-resolution optical mapping (n 5) and extracellular bipolar and intracellular microelectrode recordings (n 56) in a superfused PV preparation. RESULTS From the left atrium to distal PV, there was progressive shortening of the action potential (AP) duration, reduction in AP and bipolar electrogram amplitude, and depolarization of resting membrane potentials. Sustained PV tachycardias were induced exclusively in the presence of acetylcholine (10 7 to 10 6 mol/l, n 12). Sustained PV tachycardias were rapid (mean cycle length 93 15 ms), regular, and capable of induction, termination, and resetting by single extrastimuli. Re-entry as the mechanism underlying PV tachycardias was confirmed by optical mapping (n 5). Acetylcholine also reduced the slope of the AP restitution curve and suppressed AP alternans (n 6). Importantly, PV tachycardias exhibited 1:1 conduction into the atrium at short cycle lengths ( 100 ms), emphasizing the potential role of re-entrant PV tachycardia in atrial fibrillation. CONCLUSIONS Pulmonary veins provide a favorable substrate for re-entry formation. Heterogeneity of the electrophysiologic properties and marked abbreviation of action potential duration and refractoriness by acetylcholine combine to produce rapid and stable re-entrant PV tachycardias. Elevated parasympathetic tone and re-entrant PV tachycardia may serve as a mechanism underlying the perpetuation of PAF. (J Am Coll Cardiol 2005;45:1871–7) © 2005 by the ublished by Elsevier Inc. doi:10.1016/j.jacc.2005.02.070

BACKGROUND AND PURPOSE:It is known that encapsulation can alter the delivery of plasminogen activators by flow to accelerate fibrinolysis while other experimental studies suggest encapsulation may reduce the risk of hemorrhage with administration of the agent. The aim of this research is to resolve the effect of encapsulation on fibrinolysis and bleeding in the microcirculation.METHODS:An established rabbit model of fibrinolytic hemorrhage was utilized to explore the potential of encapsulation to limit bleeding. Equal dosages of free or microencapsulated streptokinase (MESK) were infused to initiate thrombolysis of small vessel clots while tracking blood loss.RESULTS:Compared to free streptokinase, significant improvements in bleeding were observed with MESK as demonstrated by (1) delayed onset of bleeding, (2) shortened duration, and (3) reduction in the volume of lost blood, consistent with less systemic fibrinogen degradation.CONCLUSIONS:Findings demonstrate that encapsulation of streptokinase can inhibit clot lysis in small vessels. Combined with prior work on accelerated thrombolysis, results suggest a time-based regimen for avoiding bleeding complications during thrombolytic therapy with encapsulated agent.

INTRODUCTION:There has been a long-standing controversy regarding the mechanism(s) to explain the irregular ventricular response during atrial tachycardia (AT) or atrial fibrillation (AF) and where the site of block, if any, resides in the atrioventricular (AV) junction.METHODS:We studied 12 Langendorff preparations perfused with modified Tyrode's solution containing 5-10 mM diacetyl monoxime which suppressed contractility but allowed the use of intracellular action potential (AP) recordings. Octapolar catheters (2-mm rings, 2-mm spacing) were secured along the tricuspid annulus from the apex to the base of the triangle of Koch and along the anterior limbus of the fossa ovalis to record extracellular, slow pathway, fast pathway, His bundle (Hb) and AV nodal (AVN) extracellular potentials as well as intracellular action potentials.RESULTS:AT or AF induced by rapid atrial pacing showed a variety of irregular responses due to: (1) Wenckebach conduction showing decrement within the AVN and progressive diminution of extracellular AVN potentials (n = 5); (2) repetitive concealed conduction proximal to the AVN (n = 3); (3) ectopic beats arising within the AVN (n = 2); (4) ectopic beats arising at the Hb (n = 2).CONCLUSIONS:In this experimental preparation, extracellular and intracellular recordings provided presumptive evidence for the mechanisms causing the irregularities of the ventricular response such as repetitive concealed conduction, enhanced automaticity or electrotonically triggered activity. Also more definitive determinations of the site of block in the AV junction were also obtained.

BackgroundActivating autoantibodies to β-adrenergic receptors (AAβ1/2AR) and M2 muscarinic receptors (AAM2R) have been reported in several cardiac diseases and may have pathophysiologic relevance. However, the interactions and relative effects of AAβ1AR, AAβ2AR and AAM2R on contractile function have not been characterized.MethodsThe inotropic effects of IgG from 18 selected patients with cardiomyopathy and/or atrial tachyarrhythmias positive by ELISA for antibodies to β1/2AR were studied using an isolated canine Purkinje fiber contractility assay. M2R-blockade was tested using atropine while selective β1AR and β2AR blockade used CGP-20712A and ICI-118551 respectively.ResultsFifteen of the 18 anti-β1/2AR ELISA-positive samples demonstrated evidence for negative inotropic muscarinic effects which were blocked using atropine. Atropine failed to uncover a positive inotropic response in 2 of the 18 IgG samples (false positive ELISA for AAβAR). In the remaining 16 AAβAR true-positive subjects, the β1AR-induced increase in contractility (concurrent M2/β2 blockade) was augmented to 140.5±12.2% of baseline compared to 127.4±7.2% of baseline with M2 blockade (atropine) only (p<0.001, n=16). The β2AR-induced increase in contractility (concurrent M2/β1 blockade) was only 114.5±4.3% of baseline (p<0.001, n=16). Combined M2 and β1/β2 blockade eliminated any increase in contractility.ConclusionsThe inherently positive inotropic effect of AAβ1AR was negatively modulated by AAM2R and AAβ2AR. These opposing effects of receptor-activating autoantibodies may alter cardiac performance and influence clinical outcome depending on their receptor type and relative contractile activity.

Background: Increased titers of antibodies activating beta-adrenergic and muscarinic-2 (M-2) receptors (ELISA assays) are present in the serum and purified immunoglobulins (IgG) of patients with Gr...

The ubiquitin-proteasome system (UPS) plays a central role in intracellular protein degradation and regulates many cellular processes, including cell proliferation, inflammation, adaptation to stress, cell death, and the removal of damaged or misfolded proteins. Numerous studies have demonstrated that altered UPS function is involved in the pathogenesis of a wide range of cardiac diseases including hypertrophy and failure, myocardial ischemia, atherosclerosis, and diabetic cardiovascular disease. Impairment of proteasome function is a common feature of cardiac disease; however several studies have also demonstrated increased proteasome activity in models similar but not identical with those having decreased function. Recent studies have shown that use of proteasome inhibitors before or following production of the model of cardiac disease may confer cardioprotection under certain conditions.

The intrinsic cardiac autonomic nervous system (ganglionated plexuses [GP]) plays a significant role in the initiation and maintenance of atrial fibrillation (AF) in both experimental models and AF patients. Left atrial GP, located in epicardial fat pads and the ligament of Marshall, contain afferent neurons from the atrial myocardium and the central autonomic nervous system, efferent neurons (cholinergic and adrenergic neurons), and interconnecting neurons, which allow communication between GP. Stimulation of the GP produces both parasympathetic stimulation ( markedly shortens action potential duration) and sympathetic stimulation (increases calcium transient) in the pulmonary vein (PV) myocardium and atrial myocardium. In a canine model, GP stimulation resulted in early afterdepolarizations, and calcium transient triggered firing in the adjacent PV and initiated AF. Fractionated atrial potentials (FAP) were consistently located in the left atrium close to the stimulated GP. Ablation of the stimulated GP eliminated the FAP surrounding the GP. In patients with paroxysmal AF, epicardial and endocardial high-frequency stimulation produced a positive vagal response ( transient AV block during AF and hypotension), allowing the identification and localization of five major left atrial GP (superior left GP, inferior left GP, Marshall tract GP, anterior right GP, inferior right GP). High-density electroanatomic maps of the left atrium and PVs obtained during AF showed the FAP are located in four main left atrial areas (left atrial appendage ridge FAP area, superior-left FAP area, inferoposterior FAP area, anterior-right FAP area). All five GP are located within one of the four FAP areas. In 63 patients with paroxysmal AF, GP ablation alone (before PV antrum isolation) significantly decreased the occurrence of PV firing (47/63 patients before ablation vs 9/63 patients after ablation, P < .01). GP ablation also decreased the inducibility of sustained AF (43/63 patients vs 23/63 patients, P < .01) and markedly reduced or eliminated the left atrial FAP areas.

Objectives We studied activating autoantibodies to beta-1 adrenergic receptors (AA beta 1AR) and activating autoantibodies to M2 muscarinic receptors (AAM2R) in the genesis of atrial fibrillation (AF) in Graves' hyperthyroidism.Background Atrial fibrillation frequently complicates hyperthyroidism. Both AA beta 1AR and AAM2R have been described in some patients with dilated cardiomyopathy and AF. We hypothesized that their copresence would facilitate AF in autoimmune Graves' hyperthyroidism.Methods Immunoglobulin G purified from 38 patients with Graves' hyperthyroidism with AF (n = 17) or sinus rhythm (n = 21) and 10 healthy control subjects was tested for its effects on isolated canine Purkinje fiber contractility with and without atropine and nadolol. Immunoglobulin G electrophysiologic effects were studied using intracellular recordings from isolated canine pulmonary veins. Potential cross-reactivity of AA beta 1AR and AAM2R with stimulating thyrotropin receptor (TSHR) antibodies was evaluated before and after adsorption to Chinese hamster ovary cells expressing human TSHRs using flow cytometry and enzyme-linked immunosorbent assays.Results The frequency of AA beta 1AR and/or AAM2R differed significantly between patients with AF and sinus rhythm (AA beta 1AR = 94% vs. 38%, p < 0.001; AAM2R = 88% vs. 19%, p < 0.001; and AA beta 1AR+AAM2R = 82% vs. 10%, p < 0.001). The copresence of AA beta 1AR and AAM2R was the strongest predictor of AF (odds ratio: 33.61, 95% confidence interval: 1.17 to 964.11, p = 0.04). Immunoglobulin G from autoantibody-positive patients induced hyperpolarization, decreased action potential duration, enhanced early afterdepolarization formation, and facilitated triggered firing in pulmonary veins by local autonomic nerve stimulation. Immunoadsorption studies showed that AA beta 1AR and AAM2R were immunologically distinct from TSHR antibodies.Conclusions When present in patients with Graves' hyperthyroidism, AA beta 1AR and AAM2R facilitate development of AF. (J Am Coll Cardiol 2009; 54: 1309-16) (C) 2009 by the American College of Cardiology Foundation

After the sequential successes of catheter ablation for the treatment of pre-excitation syndromes (WPW), junctional reentry (AVNRT) atrial flutter (AFL) and ventricular arrhythmias, clinical electrophysiologists have focused on the myocardial basis of atrial fibrillation (AF). Thus, the strategy for ablation of drug and cardioversion refractory AF was to isolate the myocardial connections from the focal firing pulmonary veins (PVs) in addition to altering the atrial substrate maintaining AF. However, the overall success rates have not achieved those of the other types of ablation procedures. In this review we have summarized the favorable aspects and drawbacks of pulmonary vein isolation (PVI). As for the role of the Intrinsic Cardiac Autonomic Nervous System (ICANS), both basic and clinical evidence has shown that ganglionated plexi (GP) stimulation promotes initiation and maintenance of AF, and that GP ablation reduces recurrence of AF following catheter or surgical ablation of these structures. Based on these findings, the GP Hyperactivity Hypothesis has been proposed to explain, at least in part, the mechanistic basis for the focal form of AF. For example, PV isolation may not always be necessary for elimination of AF, as in the early stages of paroxysmal AF. GP ablation alone, in these cases, may suffice for focal AF termination. In the persistent and long standing persistent forms the substrate for AF may be more extensive and therefore require GP ablation plus PV isolation and/or CFAE ablations. Clinical reports, both catheter based as well as minimally invasive surgical procedures, which include PVI plus GP ablation have shown relatively long-term success rates much closer to or equal to those achieved by myocardial ablation procedures in patients with WPW, AVNRT and AFL.

Background—The objective of this study was to develop an acute experimental model showing both focal and macroreentrant sustained atrial fibrillation (AF). Methods and Results—In 31 anesthetized dogs, bilateral thoracotomies allowed the attachment of electrode catheters at the right and left superior pulmonary veins, atrial free walls, and atrial appendages. Acetylcholine, 100 mmol/L, was applied topically to either appendage. Sequential radiofrequency ablation was achieved for the ganglionated plexi (GP), found adjacent to the 4 pulmonary veins. In 12 separate studies, a propafenone bolus, 2 mg/kg, was given before and after GP ablations at the start of acetylcholine-induced AF. Acetylcholine caused abrupt onset of AF (n=22) or induced AF by burst pacing (n=9) that lasted ≥10 minutes. Rapid, regular, or fractionated atrial electrograms were consistently seen (average cycle length, 37±7 ms) at the appendages versus cycle lengths of 114±23 ms at other atrial sites. After ablations of GP, AF abruptly terminated (n=25). In 6 dogs, sustained atrial tachyarrhythmias continued. Pacing at specific atrial sites organized electrograms of one atrium or also captured the other atrium. The latter resulted in termination when pacing was stopped in 4 of these 6 experiments. Propafenone did not change the duration of focal AF before GP ablation (17±9 versus 14±8 minutes; control, P=0.6) but terminated reentrant atrial tachyarrhythmias (12±3 versus 2±1 minutes, P=0.0009). Conclusions—Before GP ablation, acetylcholine (100 mmol/L) induced sustained AF characterized by rapid, focal firing. GP ablations were associated with loss of focal firing and regularization of electrograms in both atria before termination. Propafenone failed to terminate focal AF but rapidly terminated entrainable macroreentrant atrial tachyarrhythmias.

A 71-year-old male with well-controlled hypertension developed atrial tachyarrhythmias in 2002 and a restrictive cardiomyopathy in 2006 to 2007. Sera from 1992, 2001, and 2006 to 2008 demonstrated activating autoantibodies against β-adrenergic (AAβAR) and M2 muscarinic receptors (AAM2R). These sera have been characterized for bioactivity using in vitro assays of cardiac contractility and automaticity using a canine cardiac Purkinje fiber assay as well as protein kinase assay activation in H9c2 cells. These assays demonstrated concurrent positive βAR and inhibitory M2R effects that were blocked by nadolol and atropine, respectively. In a canine pulmonary vein atrial sleeve preparation, sera diluted 1:100 produced atrial hyperpolarization that was blocked by atropine. Atrial tachyarrhythmias developed in 2002 in the presence of a persistent bradycardia. Serial echocardiograms demonstrated progressive diastolic dysfunction in the absence of cardiac hypertrophy between 2006 and 2007. A dual-chamber pacemaker was installed with combined βAR (nadolol) and M2<3R (oxybutynin) blockade, resulting in marked suppression of atrial ectopy and improved diastolic function. The estimated pulmonary artery pressure decreased and exercise tolerance returned. Blood pressure has remained normal with β-blockade. AAβAR and AAM2R prospectively influenced atrial and ventricular function in this patient, and specific receptor blockade was associated with improved cardiac function.

OBJECTIVES:We sought to systematically investigate the role of the ligament of Marshall (LOM) and inferior left ganglionated plexi (ILGP) in modulating electrophysiological functions. METHODS:The following structures were exposed in 36 dogs: (1) LOM, (2) superior left GP (SLGP) near the junction of left superior pulmonary vein (LSPV) and left atrium, (3) ILGP near the left inferior pulmonary vein-atrial junction, (4) anterior right GP (ARGP) near the sino-atrial node, and (5) inferior right GP (IRGP) at the junction of inferior vena cava and atria. High frequency stimulation (HFS; 0.6-8.0 V, 20 Hz, 0.1 msec in duration) was applied to the LOM, SLGP, ILGP, ARGP, IRGP, or vagosympathetic trunk. Ventricular rate (VR) during atrial fibrillation (AF) was compared before and after ablation of GP in different sequences. RESULTS:ARGP + ILGP ablation but not ARGP ablation alone eliminated the VR slowing response induced by LOM stimulation, suggesting that all the autonomic innervation from the LOM to AV node passes the ILGP. LOM ablation attenuated the VR slowing response caused by SLGP or left vagosympathetic stimulation, suggesting that LOM modulates the autonomic innervation between the AV node and the left vagosympathetic trunk or SLGP. ARGP attenuated while ARGP + ILGP ablation eliminated the VR slowing response induced by left vagosympathetic stimulation, suggesting that both ARGP and ILGP modulate the AV nodal innervation of the extrinsic and intrinsic cardiac autonomic nervous system (ANS). CONCLUSION:The LOM and ILGP function as the "integration centers" that modulate the autonomic interactions between extrinsic and intrinsic cardiac ANS on AV nodal function.

Background: Orthostatic hypotension (OH) is characterized by an abnormal autonomic response to upright posture. Activating autoantibodies to beta 1/2-adrenergic (AA beta 1/2AR) and M2/3 muscarinic receptors (AAM2/3R) produce vasodilative changes in the vasculature that may contribute to OH.Methods: Immunoglobulin (Ig)G from 6 patients with idiopathic OH harboring autoantibodies and from 10 healthy control subjects were examined for: 1) beta 1AR and M2R activity with a perfused Purkinje fiber assay and PKA assay in H9c2 cells and 2) vasodilator beta 2AR and M3R activity using a pressurized cremaster resistance arteriole assay. Changes in IgG activity with and without propranolol, atropine, and L-NAME were used to estimate AA beta AR, AAM2R, and AAM3R activation of their respective functions.Results: All six patients had elevated enzyme-linked immunosorbent assay titers to at least one of the receptors compared with controls. beta AR-mediated contractility activity and M2R activity were increased in five of the six patients. IgG from all six patients produced a direct vasodilator effect on cremaster arterioles. beta AR and nitric oxide synthase blockade led to near normalization of IgG-induced vasodilation.Conclusion: AA beta 1/2AR and AAM2/3R are present in some patients with idiopathic OH compatible with an in vivo effect. These autoantibodies and their cardiovascular effects provide new mechanistic insights into the pathophysiology of OH. J Am Soc Hypertens 2012;6(1):40-47. Published by Elsevier Inc on behalf of American Society of Hypertension.

Background: We previously demonstrated that acetylcholine (Ach) injected into cardiac ganglionated plexi (GP) causes pulmonary vein (PV) ectopy initiating atrial fibrillation (AF). Objective: To determine the effects of Ach applied at non‐PV sites. Methods: Overall, 54 dogs were anesthetized with Na‐pentobarbital. A right and left thoracotomy allowed the placement of multielectrode catheters to record from the superior PVs, mid portion of the atrium and the atrial appendages (AA). A monophasic action potential (MAP) was recorded from the AA. Ach (1, 10, 100 mM) was applied sequentially to the AA. Results: In 19 of 26 animals, Ach 100 mM on the right (n = 15) or left (n = 4) AA induced focal, sustained AF (≥10 minutes) with rapid regular firing (cycle length = 37 ± 7 ms) at the AA. A clamp with teeth placed across the AA caused arrest in the AA. However, AF was sustained only when PV sites adjacent to the GP manifested complex fractionated atrial electrograms (CFAE). Clamping the AA prior to Ach (100 mM) application resulted in focal AF arising at the PVs but not at the AA. When a clamp without teeth was applied prior to Ach application, no AF at either AA or PV site could be induced. Conclusion: Isolation of the focal AF at the AA (primary trigger) by clamping caused cessation of activity in the AA, but AF continued due to secondary triggers arising from PVs. The possible mechanism(s) responsible for these findings are discussed, and various ancillary experiments (n = 28) were added to help elucidate mechanisms.