Tetsuji Shinohara

Continuous Low-Level Vagus Nerve Stimulation Reduces Stellate Ganglion Nerve Activity and Paroxysmal Atrial Tachyarrhythmias in Ambulatory Canines

Background— We hypothesize that left-sided low-level vagus nerve stimulation (LL-VNS) can suppress sympathetic outflow and reduce atrial tachyarrhythmias in ambulatory dogs. Methods and Results— We implanted a neurostimulator in 12 dogs to stimulate the left cervical vagus nerve and a radiotransmitter for continuous recording of left stellate ganglion nerve activity, vagal nerve activities, and ECGs. Group 1 dogs (N=6) underwent 1 week of continuous LL-VNS. Group 2 dogs (N=6) underwent intermittent rapid atrial pacing followed by active or sham LL-VNS on alternate weeks. Integrated stellate ganglion nerve activity was significantly reduced during LL-VNS (7.8 mV/s; 95% confidence interval [CI] 6.94 to 8.66 versus 9.4 mV/s [95% CI, 8.5 to 10.3] at baseline; P=0.033) in group 1. The reduction was most apparent at 8 AM, along with a significantly reduced heart rate (P=0.008). Left-sided low-level vagus nerve stimulation did not change vagal nerve activity. The density of tyrosine hydroxylase–positive nerves in the left stellate ganglion 1 week after cessation of LL-VNS were 99 684 &mgr;m2/mm2 (95% CI, 28 850 to 170 517) in LL-VNS dogs and 186 561 &mgr;m2/mm2 (95% CI, 154 956 to 218 166; P=0.008) in normal dogs. In group 2, the frequencies of paroxysmal atrial fibrillation and tachycardia during active LL-VNS were 1.4/d (95% CI, 0.5 to 5.1) and 8.0/d (95% CI, 5.3 to 12.0), respectively, significantly lower than during sham stimulation (9.2/d [95% CI, 5.3 to 13.1]; P=0.001 and 22.0/d [95% CI, 19.1 to 25.5], P<0.001, respectively). Conclusions— Left-sided low-level vagus nerve stimulation suppresses stellate ganglion nerve activities and reduces the incidences of paroxysmal atrial tachyarrhythmias in ambulatory dogs. Significant neural remodeling of the left stellate ganglion is evident 1 week after cessation of continuous LL-VNS.

Small-Conductance Calcium-Activated Potassium Channel and Recurrent Ventricular Fibrillation in Failing Rabbit Ventricles

Rationale: Fibrillation/defibrillation episodes in failing ventricles may be followed by action potential duration (APD) shortening and recurrent spontaneous ventricular fibrillation (SVF). Objective: We hypothesized that activation of apamin-sensitive small-conductance Ca2+-activated K+ (SK) channels is responsible for the postshock APD shortening in failing ventricles. Methods and Results: A rabbit model of tachycardia-induced heart failure was used. Simultaneous optical mapping of intracellular Ca2+ and membrane potential (Vm) was performed in failing and nonfailing ventricles. Three failing ventricles developed SVF (SVF group); 9 did not (no-SVF group). None of the 10 nonfailing ventricles developed SVF. Increased pacing rate and duration augmented the magnitude of APD shortening. Apamin (1 &mgr;mol/L) eliminated recurrent SVF and increased postshock APD80 in the SVF group from 126±5 to 153±4 ms (P<0.05) and from 147±2 to 162±3 ms (P<0.05) in the no-SVF group but did not change APD80 in nonfailing group. Whole cell patch-clamp studies at 36°C showed that the apamin-sensitive K+ current (IKAS) density was significantly larger in the failing than in the normal ventricular epicardial myocytes, and epicardial IKAS density was significantly higher than midmyocardial and endocardial myocytes. Steady-state Ca2+ response of IKAS was leftward-shifted in the failing cells compared with the normal control cells, indicating increased Ca2+ sensitivity of IKAS in failing ventricles. The Kd was 232±5 nmol/L for failing myocytes and 553±78 nmol/L for normal myocytes (P=0.002). Conclusions: Heart failure heterogeneously increases the sensitivity of IKAS to intracellular Ca2+, leading to upregulation of IKAS, postshock APD shortening, and recurrent SVF.

Diastolic Intracellular Calcium-Membrane Voltage Coupling Gain and Postshock Arrhythmias Role of Purkinje Fibers and Triggered Activity

Rationale: Recurrent ventricular arrhythmias after initial successful defibrillation are associated with poor clinical outcome. Objective: We tested the hypothesis that postshock arrhythmias occur because of spontaneous sarcoplasmic reticulum Ca release, delayed afterdepolarization (DAD), and triggered activity (TA) from tissues with high sensitivity of resting membrane voltage (Vm) to elevated intracellular calcium (Cai) (high diastolic Cai–voltage coupling gains). Methods and Results: We simultaneously mapped Cai and Vm on epicardial (n=14) or endocardial (n=14) surfaces of Langendorff-perfused rabbit ventricles. Spontaneous Cai elevation (SCaE) was noted after defibrillation in 32% of ventricular tachycardia/ventricular fibrillation at baseline and in 81% during isoproterenol infusion (0.01 to 1 &mgr;mol/L). SCaE was reproducibly induced by rapid ventricular pacing and inhibited by 3 &mgr;mol/L of ryanodine. The SCaE amplitude and slope increased with increasing pacing rate, duration, and dose of isoproterenol. We found TAs originating from 6 of 14 endocardial surfaces but none from epicardial surfaces, despite similar amplitudes and slopes of SCaEs between epicardial and endocardial surfaces. This was because DADs were larger on endocardial surfaces as a result of higher diastolic Cai–voltage coupling gain, compared to those of epicardial surfaces. Purkinje-like potentials preceded TAs in all hearts studied (n=7). IK1 suppression with CsCl (5 mmol/L, n=3), BaCl2 (3 &mgr;mol/L, n=3), and low extracellular potassium (1 mmol/L, n=2) enhanced diastolic Cai–voltage coupling gain and enabled epicardium to also generate TAs. Conclusions: Higher diastolic Cai–voltage coupling gain is essential for genesis of TAs and may underlie postshock arrhythmias arising from Purkinje fibers. IK1 is a major factor that determines the diastolic Cai–voltage coupling gain.

Genesis of phase 3 early afterdepolarizations and triggered activity in acquired long-QT syndrome.

Background—Both phase 2 and phase 3 early afterdepolarizations (EADs) occur in long-QT syndromes, but their respective roles in generating arrhythmias in intact cardiac tissue are incompletely understood. Methods and Results—Intracellular Ca (Cai) and membrane voltage (Vm) were optically mapped in a quasi 2-dimensional model of cryoablated Langendorff-perfused rabbit ventricles (n=16). E-4031 (an IKr blocker) combined with reduced extracellular K ([K+]o) and Mg ([Mg2+]o) prolonged action potential duration heterogeneously and induced phase 2 and phase 3 EADs. Whereas phase 2 EADs were Cai-dependent, phase 3 EADs were not. The origins of 47 triggered activity episodes were attributed to phase 2 EADs in 12 episodes (26%) and phase 3 EADs in 35 episodes (74%). When phase 2 EADs accompanied phase 3 EADs, they accentuated action potential duration heterogeneity, creating a large Vm gradient across the boundary between long and short action potential duration regions from which triggered activity emerged. The amplitude of phase 3 EADs correlated with the Vm gradient (r=0.898, P<0.001). Computer simulation studies showed that coupling of cells with heterogeneous repolarization could extrinsically generate phase 3 EADs via electrotonic current flow. Alternatively, reduced IK1 caused by low [K+]o could generate intrinsic phase 3 EADs capable of inducing triggered activity at the boundary zone. Conclusions—Phase 3 EADs can be extrinsic as the result of electrotonic current across steep repolarization gradients or intrinsic as the result of low IK1 and do not require spontaneous sarcoplasmic reticulum Ca release. Reduction of IK1 by low [K+]o strongly promotes ventricular arrhythmias mediated by phase 3 EADs in acquired long-QT syndrome caused by IKr blockade.

Patterns of baseline autonomic nerve activity and the development of pacing-induced sustained atrial fibrillation

BACKGROUND Whether autonomic nerve activity is important in the development of pacing-induced sustained atrial fibrillation (AF) is unclear. OBJECTIVE The purpose of this study was to test the hypothesis that patterns of baseline autonomic nerve activity are important in the development of pacing-induced sustained AF. METHODS Radiotransmitters were implanted in 12 ambulatory dogs to record left stellate ganglion nerve activity (SGNA) and vagal nerve activity (VNA). Sustained (>48 hours) AF was induced with intermittent rapid atrial pacing. RESULTS At baseline (before pacing), 1-minute integrated nerve activity between SGNA and VNA demonstrated either a single linear relationship with excellent correlation (group 1, N = 3, r = 0.816 ± 0.105) or nonlinear relationships with poor correlation (group 2, N = 9, r = 0.316 ± 0.162, P <.05 vs group 1). Group 1 dogs had higher VNA (97.0 ± 11.5 mV-s) compared to group 2 (33.4 ± 21.7 mV-s, P <.001). Group 1 dogs had more frequent sympathovagal co-activation episodes than did group 2 (50 ± 19 per day vs 15 ± 6 per day, P <.05) and more paroxysmal atrial tachycardia (PAT; 5 ± 1 per day vs 2 ± 1 per day, P <.05) at baseline. Sustained AF occurred after 16 ± 4 days (range 13-20 days) of pacing in group 1 and after 46 ± 18 days (range 23-72 days) of pacing in group 2 (P <.05). In the week before development of sustained AF, VNA of group 2 dogs was significantly increased compared to baseline (P <.05). CONCLUSION Ambulatory dogs with good linear sympathovagal correlation and higher vagal tone at baseline have more PAT episodes at baseline and faster induction of sustained AF by rapid pacing. Rapid atrial pacing increased the VNA of the remaining dogs before induction of sustained AF.

Phosphatidylinositol 3-kinase-dependent activation of akt, an essential signal for hyperthermia-induced heat-shock protein 72, is attenuated in streptozotocin-induced diabetic heart

We tested the hypothesis that phosphatidylinositol 3-kinase (PI 3-kinase)-dependent activation of Akt is essential for the expression of cardiac heat-shock protein 72 (HSP72) and that this pathway is impaired in the streptozotocin (STZ)-induced diabetic heart. STZ-induced male diabetic rats were treated with insulin (STZ-insulin group, n = 26) or vehicle (STZ-vehicle group, n = 61) for 3 weeks. Whole-body hyperthermia (43°C for 20 min) was applied, and the heart was isolated 24 h later. Compared with control heart, hyperthermia-induced HSP72 expression and phosphorylation of Akt were attenuated in the STZ-vehicle heart. Pretreatment with wortmannin attenuated hyperthermia-induced HSP72 expression and phosphorylation of Akt. In isolated perfused heart experiments, the hyperthermia-treated STZ-vehicle heart showed poor left ventricular functional recovery during reperfusion after no-flow global ischemia compared with hyperthermia-treated control heart. Insulin treatment restored HSP72 expression and reperfusion-induced functional recovery. In cultured neonatal rat cardiomyocytes, hyperthermia-induced HSP72 expression was enhanced by insulin, together with tolerance against hypoxia-reoxygenation injury. Wortmannin and LY294002 inhibited hyperthermia-induced HSP72 expression and phosphorylation of Akt. Our results indicate that activation of Akt, in a PI 3-kinase–dependent manner, is essential for hyperthermia-induced HSP72 expression in association with cardioprotection, suggesting impairment of this signaling pathway in the STZ-induced diabetic heart, probably due to insulin deficiency.

Heart Failure Decreases Nerve Activity in the Right Atrial Ganglionated Plexus

Reduced Vagal Control in Heart Failure.

Role of leptin signaling in the pathogenesis of angiotensin II-mediated atrial fibrosis and fibrillation.

Background—We examined the hypothesis that leptin signaling contributes to the atrial fibrosis and atrial fibrillation (AF) evoked by angiotensin II (AngII). Methods and Results—Eight-week-old male CL57/B6 (CNT) and leptin-deficient ob/ob mice (Ob) were subcutaneously infused with AngII (2.0 mg/kg per day). Two weeks later, transesophageal burst pacing and an electrophysiological study using isolated perfused hearts were performed. Left-atrial tissues were collected to determine interstitial fibrosis by Masson trichrome staining, and the expressions of mRNAs related to inflammatory profibrotic signals were assessed. Left-atrial fibroblasts were isolated from adult Sprague-Dawley and Zucker rats. The effects of leptin (100 ng/mL) or AngII (100 nmol/L) treatment were evaluated. In CNT-AngII mice, leptin expression in the left atrium was upregulated (P<0.01). Transesophageal burst pacing induced atrial fibrillation in 88% (7/8) of CNT-AngII mice, but not in Ob-AngII mice (0/8; P<0.01). In isolated perfused hearts, atrial fibrillation was induced only in CNT-AngII mice (4/6; 67%). Interatrial conduction time was prolonged in CNT-AngII mice (P<0.01), but not in Ob-AngII mice. The upregulation of collagen 1, collagen 3, transforming growth factor-&bgr;1, &agr;-smooth muscle actin, MCP-1, F4/80, and RANTES mRNA, which was seen in CNT-AngII mice, was attenuated in Ob-AngII mice. In cultured Sprague-Dawley rat atrial fibroblasts, AngII treatment increased leptin expression (P<0.01). Addition of leptin increased transforming growth factor-&bgr;1, &agr;-smooth muscle actin, MCP-1, and RANTES expressions in Sprague-Dawley rat atrial fibroblasts, but not in Zucker rat atrial fibroblasts. Conclusions—Our results demonstrate for the first time that leptin signaling is essential for the development of atrial fibrosis and atrial fibrillation evoked by AngII.

Establishment of a model of atrial fibrillation associated with chronic kidney disease in rats and the role of oxidative stress

BACKGROUND An animal model of atrial fibrillation (AF) associated with chronic kidney disease (CKD) has not been available. OBJECTIVE The purpose of this study was to test the validity of 5/6 nephrectomy (5.6Nx) as an appropriate model of AF associated with CKD and to investigate the role of oxidative stress. METHODS Male Sprague-Dawley rats were subjected to 5.6Nx. A novel derivative of lipoic acid, sodium zinc dihydrolipoylhistidinate (DHLHZn), was subcutaneously infused. Four weeks later, hearts were isolated. RESULTS We observed 5 main findings. (1) 5.6Nx induced renal dysfunction with elevation of systolic blood pressure and impaired glucose tolerance. (2) In the left atrium (LA), expressions of α-smooth muscle action and collagen type I, the compositional proteins of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase, and malondialdehyde were increased by 5.6Nx, which was reversed by DHLHZn treatment. (3) In the LA, the tissue content of angiotensin II was elevated by 5.6Nx, which was also reversed by DHLHZn. (4) Masson trichrome staining revealed that heterogeneous LA interstitial fibrosis was induced by 5.6Nx, which was attenuated by DHLHZn. (5) In isolated perfused heart experiments, 5.6Nx caused slowing of interatrial conduction. In the hearts of rats of the 5.6Nxgroup, right atrial extrastimuli invariably induced AF (8/8 [100%]), which were suppressed by DHLHZn (3/8 [38%], P <.05). CONCLUSION We successfully established an appropriate model of AF associated with CKD in rats. Because the amount of NADPH oxidase was increased and the potent antioxidant agent DHLHZn was effective, oxidative stress may be involved in the pathogenesis of LA fibrosis and enhanced AF vulnerability in our model.

Effect of cardiac resynchronization therapy on cardiac sympathetic nervous dysfunction and serum C-reactive protein level.

Background:  Cardiac autonomic dysfunction is associated with a poor prognosis in patients with heart failure (HF). Systemic inflammation is elevated in patients with HF. We hypothesized that cardiac resynchronization therapy (CRT) improves cardiac sympathetic nervous dysfunction and systemic inflammation. To test our hypothesis, we evaluated cardiac sympathetic activity and serum levels of high sensitive C‐reactive protein (hs‐CRP) before and after CRT.