Author + information
- Received July 27, 2016
- Revision received January 23, 2018
- Accepted February 8, 2018
- Published online March 28, 2018.
- Richard S. Shelton, MDa,
- Masahiro Ogawa, MD, PhDa,b,
- Hongbo Lin, MSc,
- Changyu Shen, PhDc,
- Johnson Wong, BSa,
- Shien-Fong Lin, PhDd,
- Peng-Sheng Chen, MDa and
- Thomas H. Everett IV, PhDa,∗ ()
- aKrannert Institute of Cardiology and the Division of Cardiology, Department of Medicine, Indiana University School of Medicine, Indianapolis, Indiana
- bDepartment of Cardiology, Fukuoka University School of Medicine, Fukuoka, Japan
- cDivision of Cardiology, Department of Medicine, Indiana University School of Medicine, Indianapolis, Indiana
- dInstitute of Biomedical Engineering, National Chiao Tung University, Hsin-Chu, Taiwan
- ↵∗Address for correspondence:
Dr. Thomas H. Everett IV, Krannert Institute of Cardiology, Indiana University School of Medicine, 1800 North Capitol Avenue, Indianapolis, Indiana 46202.
Objectives This study aimed to test the hypothesis that subcutaneous nerve activity (SCNA) can adequately estimate the cardiac sympathetic tone and the effects of cryoablation of the stellate ganglion in dogs with pacing-induced heart failure (HF).
Background Recording of SCNA is a new method to estimate sympathetic tone in dogs. HF is known to increase sympathetic tone and atrial arrhythmias.
Methods Twelve dogs with pacing-induced HF were studied using implanted radiotransmitters to record the stellate ganglia nerve activity (SGNA), vagal nerve activity, and SCNA. Of these, 6 dogs (ablation group) underwent bilateral stellate ganglia cryoablation before the rapid ventricular pacing; the remaining 6 dogs (control group) had rapid ventricular pacing only. In both groups, SCNA was compared with SGNA and the occurrence of arrhythmias.
Results SCNA invariably increased before the 360 identified atrial tachyarrhythmia episodes in the 6 control dogs before and after HF induction. SCNA and SGNA correlated in all dogs with an average correlation coefficient of 0.64 (95% confidence interval: 0.58 to 0.70). Cryoablation of bilateral stellate ganglia significantly reduced SCNA from 0.34 ± 0.033 μV to 0.25 ± 0.028 μV (p = 0.03) and eliminated all atrial tachyarrhythmias.
Conclusions SCNA can be used to estimate cardiac sympathetic tone in dogs with pacing-induced HF. Cryoablation of the stellate ganglia reduced SCNA and arrhythmia vulnerability.
Sympathetic tone measured by stellate ganglion nerve activity (SGNA) has been shown to influence atrial electrophysiology and the onset of atrial arrhythmias. We (1–3) recently proposed a new method to simultaneously record electrocardiogram (ECG) and subcutaneous nerve activity (SCNA) using bipolar electrodes placed under the skin. The electrical signals were low-pass filtered to optimize ECG and high-pass filtered to reveal SCNA. Because a good correlation has been shown between SCNA and the simultaneously recorded SGNA (1), we proposed that SCNA may be used to estimate cardiac sympathetic tone. The noninvasive estimation of sympathetic tone may provide new insights into the mechanisms of cardiac arrhythmias and may also provide a direct method to measure the success of neuromodulation procedures. Ogawa et al. (4,5) previously performed 2 studies in dogs with pacing-induced heart failure (HF). In the first study, it was shown that HF increases SGNA and occurrence of cardiac arrhythmias (4). In a second study using the same HF model, cryoablation of the caudal half of the left and right stellate ganglia and T2 to T4 thoracic sympathetic ganglia was performed. It was shown that cryoablation significantly reduced both SGNA and paroxysmal atrial tachycardia (PAT) episodes (5). The data from these studies have not been analyzed to determine if SCNA increased after induction of HF and decreased after cryoablation of the stellate ganglion. We retrieved the data from both studies and manually analyzed de novo the SGNA and atrial tachyarrhythmia episodes. We also performed analyses of SCNA using methods previously reported (1). The purpose of the present study was to test the hypothesis that: 1) HF increases both SCNA and SGNA; 2) both SCNA and SGNA preceded the onset of PAT at baseline and after induction of HF; and 3) cryoablation of the stellate ganglion can reduce both SCNA and SGNA in ambulatory dogs.
Original data from 2 previously published reports (4,5) were retrieved and manually analyzed. The research protocols were approved by the Institutional Animal Use and Care Committees of the Cedars-Sinai Medical Center, Los Angeles, California, and the Indiana University School of Medicine at the Methodist Research Institute, Indianapolis, Indiana.
The surgical preparations have been described previously. Briefly, all surgeries were performed under isoflurane anesthesia. Subcutaneous bipolar electrodes were placed in the left thorax of dogs, 6 to 10 cm apart, to record SCNA. The electrodes were stainless steel wires with the terminal 5 mm of the wires stripped of their insulation and used for electrical recording. The leads were attached to a Data Science International D70-EEE radiotransmitter (DSI, St. Paul, Minnesota), which was also implanted in the subcutaneous space. In the same procedure, through a left thoracotomy, a pair of bipolar electrodes from the DSI radiotransmitter was sutured onto the left stellate ganglion (LSG) to record SGNA. Another set of bipolar electrodes was placed on the left thoracic vagal nerve to record vagal nerve activity (VNA).
In all dogs, a pacing lead was implanted in the right ventricular apex and connected to a Medtronic Itrel neurostimulator for high-rate ventricular pacing. After recovery from the initial surgery, the dogs had 2 weeks of continuous baseline ambulatory monitoring. After the baseline recording, the first group of 6 dogs (group 1) underwent right ventricular pacing at 150 beats/min for 3 days, at 200 beats/min for 3 days, and then at 250 beats/min for 3 weeks to induce HF, which was confirmed by ECG (6). The pacemakers were then turned off to allow ambulatory monitoring for an additional 2 weeks. In the second group of 6 dogs (group 2), the caudal half of the left stellate ganglion and T2 to T4 thoracic sympathetic ganglia were cryoablated through the left thoracotomy using a 7-cm SurgiFrost probe (CryoCath Technologies, Inc., Montreal, Canada) during the same surgery in which the DSI radiotransmitter was implanted for monitoring of SCNA, SGNA, and VNA. The dogs then underwent the same surgery and protocol for implantation of the Medtronic Itrel neurostimulator for subsequent high-rate ventricular pacing to induce HF. These dogs were also monitored for 2 weeks before and after HF induction.
Recordings from the bipolar electrodes connected to the radiotransmitter were analyzed to obtain SCNA, SGNA, VNA, and ECG using custom-written software. The impedance of the electrodes was 0.7 to 0.8 Ω and the recordings were amplified 10,000 times and sampled at 1,000 Hz. We obtained an ECG for analyses by applying a bandpass filter (5 to 100 Hz) on the vagus nerve channel or subcutaneous nerve channel of the radiotransmitter. SCNA, SGNA, and VNA were obtained by high-pass filtering at 150 Hz. The SGNA and VNA were rectified and integrated over 1 min. The SCNA was also rectified and integrated over the same minute after adjusting the SCNA with wavelet analysis as described previously (7). In particular, spike-triggered averaging was performed to allow removal of the ventricular electrogram from the SCNA by subtracting a ventricular electrogram template obtained from averaged ventricular electrograms in the observational window. Finally, the ECG was used to calculate the heart rate over the same minute. The sum of all digitized nerve activity was then divided by the total number of samples during the same period to obtain averaged SGNA (aSGNA), averaged SCNA (aSCNA), and averaged VNA (aVNA) per sample. Nerve activity was also quantified by determining the number of spikes and bursts within the nerve activity (8). The filtered and rectified signals from a 24-h recording were integrated over a 100-ms sliding window. Within this integrated signal, nerve spikes and bursts of nerve activity were detected. A spike was defined as the continuous period in which the amplitude is higher than the threshold value. Each spike’s duration and average amplitude were measured within that period. The threshold value was calculated as 3 times the baseline noise level, which was considered to be the lowest 10th percentile of all values during the 24-h recording. Spikes with an inter-spike interval of <5 s were grouped into bursts.
The aSGNA, aSCNA, aVNA, and heart rate were calculated every hour of a 24-h period (2:00 pm to 2:00 pm the next day) before pacing at baseline and of every hour of the 24-h period (2:00 pm to 2:00 pm the next day) immediately after pacing was stopped. To avoid periods of dropped radiotransmission or artifactual noise caused by movement of the dog, which was greater than nerve activity, the first 10 analyzable minutes of each hour were used for the calculation of the averaged nerve activity and heart rate.
Paroxysmal atrial tachyarrhythmias were defined as episodes of tachycardia (≥175 beats/min) with abrupt onset (>50 beats/min/s) and a duration of >5 s. Of these episodes, one-half were randomly selected for P-wave analysis. This analysis involved comparing the P waves before the initiation of the tachycardia to the P waves during the tachycardia episode and analyzing the changes in P-wave morphology and PR interval.
The averaged nerve activities were compared between the control dogs and cryoablated dogs using 1-sided Student t tests both before and after pacing. In addition, the aSCNA for each dog was used to look at the effect of ventricular pacing on both control and cryoablated dogs by 1-sided paired t tests. The data were presented as mean ± SD. To test for circadian variation, generalized additive mixed-effects models were fitted to the longitudinal data. A Pearson correlation coefficient was used to compare the SCNA and SGNA between the control and the ablation groups for 24 h before and after pacing. A p value ≤0.05 was considered significant.
Effect of stellate ganglion cryoablation on nerve activity
The group 1 dogs did not have ablation, whereas the group 2 dogs had stellate ganglion ablation during the first surgery. Figure 1 shows that, at baseline (before rapid ventricular pacing), aSGNA was significantly lower in the group 2 dogs (0.56 ± 0.13 μV) than group 1 dogs (1.14 ± 0.11 μV) (p = 0.003), indicating ablation significantly reduced aSGNA. aSCNA was also significantly lower in group 2 versus group 1 dogs (0.25 ± 0.028 μV vs. 0.34 ± 0.033 μV, p = 0.03). The aVNA was lower in group 2 (0.21 ± 0.0068 μV) than in group 1 (0.48 ± 0.066 μV, p = 0.001) dogs. The heart rate was also lower in group 2 (88 ± 7.9 beats/min) than group 1 (109 ± 4.0 beats/min, p = 0.02). Consistent with a previous report (4), rapid ventricular pacing induced HF increased the nerve activities in both groups of dogs. Figure 2 shows the differences between group 1 and group 2 dogs after HF had been induced. As shown in that figure, the group 2 dogs had lower nerve activities than group 1 dogs, but the differences did not reach statistical significance for aSCNA and aSGNA. The aSCNA was 0.28 ± 0.024 μV in group 2 and 0.43 ± 0.079 μV in group 1 (p = 0.06). The aSGNA was 0.91 ± 0.27 μV for group 2 and 1.23 ± 0.27 μV for group 1 (p = 0.21). The VNA was 0.21 ± 0.011 μV for group 2 and 0.55 ± 0.082 μV for group 1 (p = 0.001). The heart rate was 87 ± 4.9 beats/min for group 2 and 96 ± 3.9 beats/min for group 1 (p = 0.11). The nerve activity was also quantified by detecting the spikes and bursts of activity within the nerve recording. Figure 3 shows that the amplitude of the bursts was significantly decreased after cryoablation for each type of nerve recording (SCNA, SGNA, and VNA). Although there was no significant change in the burst frequency, because of the significant reduction in burst amplitude, total nerve activity is reduced.
SCNA and atrial tachyarrhythmias
Dogs normally have PAT at baseline (9). Consistent with that report, we were able to identify a total of 243 PAT episodes in 6 group 1 dogs during a 24-hour period of monitoring at baseline and 117 PAT episodes after induction of HF by rapid pacing. Our new analyses confirmed the previous report that no PAT episodes were present in group 2 dogs either at baseline or after rapid ventricular pacing (6).
In all PAT episodes, an increase in the SCNA and SGNA correlated with the onset of the tachycardia (Figure 4). At the arrows labeled A and B, the SGNA and SCNA respectively begin to increase dramatically. The heart rate also increased suddenly, increasing at a rate of 87 beats/min/s (abrupt onset). The heart rate increased to >175 beats/min for over 7 s, fulfilling the diagnostic criteria for PAT. In all episodes, as in this example, increased SCNA was closely related to increases in SGNA during PAT episodes.
P-wave changes were found in most randomly selected episodes of arrhythmias, including P-wave morphology changes and PR interval shortening, as shown in Figure 5. In the atrial tachyarrhythmia in Figure 4, the P-wave before the tachycardia started with a biphasic P-wave with an initial upward phase followed by a negative phase (Figure 5A), but after the tachycardia, the P-wave was initially negative with a small positive phase, and then was even more negative (Figure 5B). Furthermore, the PR interval before the tachycardia was 125 ms and was shortened to 105 ms during the tachycardia. These findings indicate that the PAT episodes are unlikely to be sinus tachycardia.
Correlation between SGNA and SCNA
The nerve activity of stellate ganglia and subcutaneous nerves for both groups 1 and 2 at baseline and after pacing were compared by plotting SGNA by SCNA. Linear regression was then used to fit the data and calculate the correlation coefficient. Figure 6 shows the results for dog A, including the correlation coefficient of r = 0.61 and p < 0.0005. The correlation coefficient and p value for each dog both before and after pacing is shown in Table 1. In general, SCNA correlated moderately well with SGNA, with an average of all correlation coefficients of all dogs at 0.64 (95% confidence interval: 0.58 to 0.70). To compare the 2 groups, before pacing, the mean difference between the ablation group and the control group (ablation – control) is 0.2567 (p = 0.01). After pacing, the mean difference between the ablation group and the control group (ablation – control) is –0.035 (p = NS).
Figure 7 summarizes the SCNA data for all dogs. The SCNA had a significant circadian variation (p < 0.001) when fitted to a generalized additive mixed-effects model. Figure 8 shows the generalized additive mixed-effect model fitted to the SCNA data. Previous studies have shown a similar significant circadian variation for the SGNA (5). Compared with the SCNA in Figure 7A, they both show a progressive increase in the morning hours, with a secondary increase in the late afternoon.
Using SCNA to measure sympathetic tone
HF is characterized by increased sympathetic tone and cardiac arrhythmias (4). The sympathetic nerve activity as measured by microneurography is reduced by cardiac resynchronization therapy in the responders, but not in nonresponders (10). Because microneurography techniques are difficult to perform and cannot be used in ambulatory patients however, sympathetic nerve activity measurements have not been clinically useful in assessing the effects of cardiac resynchronization therapy or other neuromodulation methods. The results of this study suggest that the SCNA might be a useful method to directly measure sympathetic tone in HF and determine the effects of HF therapy. A second possible use of SCNA is to determine if cardiac arrhythmias are triggered by sympathetic activation. A previous study showed that SCNA was observed before a majority of episodes of ventricular tachycardia and ventricular fibrillation in an ambulatory canine model of sudden cardiac death (2). In the present study, we have shown that SCNA preceded the onset of all PAT episodes both at baseline and after the induction of HF. These findings suggest that SCNA can be useful in determining whether PAT was caused by sympathetic activation.
Sources of SCNA
Axonal tracer studies have shown that a significant portion of the skin sympathetic nerves of the neck and upper thorax originate from the ipsilateral stellate ganglion (11,12). Several other studies have demonstrated a strong relationship between sympathetic nerve structures in the chest and skin nerves. Donadio et al. have shown in skin biopsies an abundance of sympathetic nerves in arteriovenous anastomosis, arrector pilorum muscles, and arterioles (13). Baron et al. showed in axonal tracer studies that nearly all skin sympathetic nerve somata are located in the middle cervical and stellate ganglia (11). The middle cervical ganglion and the stellate ganglion are highly interconnected to intrinsic cardiac neurons. In addition, it has been shown that SCNA can be used as an estimate of sympathetic tone (1) and is more accurate than heart rate variability (14). The current study confirms that SGNA and SCNA are closely related because they both increase before the onset of paroxysmal atrial tachyarrhythmias and both decrease after cryoablation.
Effects of LSG ablation on SCNA
Previous studies have shown that the right stellate ganglion (RSG) controlled heart rate and only had an effect on blood pressure after the left stellate ganglion was completely resected. This suggests that the RSG innervates the sinus node and plays a role in regulating the heart rate. The LSG controls blood pressure and is thought to be the main influence in cardiac sympathetic control (15). In a separate canine study, it was demonstrated that LSG stimulation significantly increased atrial fibrillation inducibility over RSG stimulation. Removing the LSG significantly reduced the incidence of atrial fibrillation (16); however, in a clinical study, both RSG and LSG block showed similar results in reduced atrial fibrillation inducibility and duration (17). In addition to controlling atrial arrhythmias in canine models (18), LSG ablation is known to be effective in decreasing sympathetic outflow and controlling ventricular arrhythmias (19–22). In animal experiments, it has also been shown that partial LSG ablation can reduce or eliminate PAT in ambulatory canine models (5,18). Left cardiac sympathetic denervation (LCSD) has been used as a therapeutic option for patients with catecholaminergic polymorphic ventricular tachycardia that have recurrent ventricular fibrillation/ventricular tachycardia episodes even with β-blocker therapy. Surgical approaches include thoracoscopic, transaxillary, and supraclavicular to expose the left-sided sympathetic chain from T4 to T1. Resection or ablation of the sympathetic ganglia from T1 to T2 then has shown to have a positive effect in these patients. A more minimally invasive procedure using video-assisted thoracic surgery to perform LCSD has also proven to be effective in reducing cardiac events. Even though we have previously shown that LCSD therapy with cryoablation can be used as a therapy for atrial arrhythmias, it is still not a commonly used approach as a clinical therapeutic option for atrial arrhythmias.
If SGNA is a source of SCNA, then partial LSG ablation should reduce SCNA. The latter findings would further support the validity of SCNA in estimating SGNA. As shown in Figure 1, however, the LSG ablation seems to have more effects on aSGNA than aSCNA. One reason is that the subcutaneous nerves in the thorax may not all come from the LSG; therefore, the baseline correlation between aSGNA and aSCNA was only moderate. A second reason is that LSG is a large nerve structure that generates strong electrical signals. Because of the high signal-to-noise ratio, the amplitude of reduction of SGNA after cryoablation is easier to visualize. In contrast, the subcutaneous nerves are small (average <1 μV at baseline) and have a smaller signal-to-noise ratio. Given that inherent noise of the recording system is a significant portion of the signal, a reduction of SCNA may not be as apparent after LSG ablation. Monitoring SCNA has many advantages over monitoring SGNA as an indicator of sympathetic nerve activity. Monitoring SGNA or other indicators of sympathetic nerve activity have involved an open chest procedure to implant electrodes on the stellate ganglion or other nerve structures. On the other hand, implantation of subcutaneous electrodes in the chest could be accomplished much less invasively. In addition, the study of subcutaneous nerve activity may lead to the use of surface electrodes for the monitoring of sympathetic nerve activity. Preliminary results have shown that it is possible to record the skin sympathetic nerve activity using ordinary ECG patch electrodes (3,23). For these reasons, SCNA may be a useful tool in minimally invasively monitoring sympathetic nerve activity involved in the arrhythmogenesis of atrial tachyarrhythmias and in estimating the efficacy of neuromodulation procedures.
The electrical signals within the recordings from subcutaneous electrodes may come from multiple sources, including low-frequency motion artifacts, ECG, respiratory muscle activity, and nerve activities. Although some noise can be effectively filtered with conventional filters, the ECG can contain high-frequency content that is within the range of the frequency bandwidth of the nerve activity; therefore, we decided to use a wavelet filter to filter out much of the ECG as we have done in previous studies with large ECG signals accompanying the nerve activity of interest (7). Although integrated SCNA showed a significant correlation with integrated SGNA, integrated wavelet SCNA appeared to have a greater correlation with integrated SGNA. The dogs do not have sweat glands in the thorax where the recordings were made. Whether sweating affects the SCNA recording cannot be determined in this study.
In addition, our study used widely spaced electrodes to monitor subcutaneous nerve activity. Other studies in our laboratory have used more narrowly spaced electrodes with varying success (1,3). When using the DSI D70-EEE radiotransmitter with narrowly spaced electrodes, the bandwidth and sampling rate of the device makes the signal-to-noise ratio insufficient to accurately monitor subcutaneous nerve activity; however, when using narrowly spaced electrodes with recording equipment capable of recording with higher bandwidth and higher sampling rate of 10,000 Hz, we were able to high-pass filter at 500 Hz, improving the signal-to-noise ratio and increasing the accuracy of monitoring the subcutaneous nerve activity (1).
Subcutaneous nerve activity correlated with SGNA and the onset of arrhythmias. Cryoablation of the stellate ganglia reduced the SCNA, further validating the use of SCNA to estimate cardiac sympathetic tone. This study suggests that using equipment with an appropriate bandwidth and sampling rate, recording subcutaneous nerve activity with an implantable device may be possible and the SCNA can be used as a surrogate for SGNA.
COMPETENCY IN MEDICAL KNOWLEDGE: Microneurography is used to directly measure sympathetic nerve activity; however, microneurography techniques are difficult to perform and cannot be used in ambulatory patients. The results of this study suggest that the SCNA might be a useful method to directly measure sympathetic tone in HF, to determine the effects of HF therapy, and to determine if cardiac arrhythmias are triggered by sympathetic activation.
TRANSLATIONAL OUTLOOK: Neuromodulation therapy including the ablation of the ganglionated plexi has been used for controlling atrial fibrillation. Techniques that can be used as a surrogate for microneurography can increase the clinical usefulness of sympathetic nerve activity measurements to help in assessing the efficacy of neuromodulation.
This study was supported in part by National Institutes of Health Grants P01HL78931, R01HL71140, R41HL124741, and R42DA043391, U18TR002208, and R01HL139829; a Medtronic-Zipes Endowment; the Charles Fisch Cardiovascular Research Award endowed by Dr. Suzanne B. Knoebel of the Krannert Institute of Cardiology; and the Indiana University Health-Indiana University School of Medicine Strategic Research Initiative. Drs. Lin, Wong, and Everett have equity interest in Arrhythmotech, LLC. Cyberonics, Medtronic, and St. Jude Medical Inc. donated research equipment to Dr. Chen’s research laboratory.
All authors attest they are in compliance with human studies committees and animal welfare regulations of the authors’ institutions and Food and Drug Administration guidelines, including patient consent where appropriate. For more information, visit the JACC: Clinical Electrophysiology author instructions page.
- Abbreviations and Acronyms
- averaged stellate ganglia nerve activity
- averaged subcutaneous nerve activity
- averaged vagus nerve activity
- heart failure
- left cardiac sympathetic denervation
- left stellate ganglion
- paroxysmal atrial tachycardia
- right stellate ganglion
- subcutaneous nerve activity
- stellate ganglia nerve activity
- vagus nerve activity
- Received July 27, 2016.
- Revision received January 23, 2018.
- Accepted February 8, 2018.
- 2018 American College of Cardiology Foundation
- Doytchinova A.,
- Patel J.,
- Zhou S.,
- et al.
- Jiang Z.,
- Zhao Y.,
- Doytchinova A.,
- et al.
- Ogawa M.,
- Zhou S.,
- Tan A.Y.,
- et al.
- Ogawa M.,
- Tan A.Y.,
- Song J.,
- et al.
- Park H.W.,
- Shen M.J.,
- Han S.,
- et al.
- Choi E.K.,
- Shen M.J.,
- Han S.,
- et al.
- Wu G.,
- DeSimone C.V.,
- Suddendorf S.H.,
- et al.
- Zhou Q.,
- Hu J.,
- Guo Y.,
- et al.
- Leftheriotis D.,
- Flevari P.,
- Kossyvakis C.,
- et al.
- Tan A.Y.,
- Zhou S.,
- Ogawa M.,
- et al.
- Schwartz P.J.,
- Priori S.G.,
- Cerrone M.,
- et al.
- Buckley U.,
- Yamakawa K.,
- Takamiya T.,
- Andrew Armour J.,
- Shivkumar K.,
- Ardell J.L.
- Doytchinova A.,
- Hassel J.,
- Yuan Y.,
- et al.