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Summary

Abstract

Introduction

Protocol

Representative Results

Discussion

Acknowledgements

Materials

References

Medicine

Multi-system Monitoring for Identification of Seizures, Arrhythmias and Apnea in Conscious Restrained Rabbits

Published: March 27th, 2021

DOI:

10.3791/62256

1Department of Pharmacology, Upstate Medical University, 2Department of Neurology, Upstate Medical University, 3Strong Epilepsy Center, Departments of Neurology, Anesthesiology / Perioperative Medicine, & Pediatrics, University of Rochester Medical Center

Using simultaneous video-EEG-ECG-oximetry-capnography, we developed a methodology to evaluate the susceptibility of rabbit models to develop provoked arrhythmias and seizures. This novel recording system establishes a platform to test the efficacy and safety of therapeutics and can capture the complex cascade of multi-system events that culminate in sudden death.

Patients with ion channelopathies are at a high risk of developing seizures and fatal cardiac arrhythmias. There is a higher prevalence of heart disease and arrhythmias in people with epilepsy (i.e., epileptic heart.) Additionally, cardiac and autonomic disturbances have been reported surrounding seizures. 1:1,000 epilepsy patients/year die of sudden unexpected death in epilepsy (SUDEP). The mechanisms for SUDEP remain incompletely understood. Electroencephalograms (EEG) and electrocardiograms (ECG) are two techniques routinely used in the clinical setting to detect and study the substrates/triggers for seizures and arrhythmias. While many studies and descriptions of this methodology are in rodents, their cardiac electrical activity differs significantly from humans. This article provides a description of a non-invasive method for recording simultaneous video-EEG-ECG-oximetry-capnography in conscious rabbits. As cardiac electrical function is similar in rabbits and humans, rabbits provide an excellent model of translational diagnostic and therapeutic studies. In addition to outlining the methodology for data acquisition, we discuss the analytical approaches for examining neuro-cardiac electrical function and pathology in rabbits. This includes arrhythmia detection, spectral analysis of EEG and a seizure scale developed for restrained rabbits.

Electrocardiography (ECG) is routinely used in the clinical setting to assess the dynamics of cardiac electrical conduction and the electrical activation-recovery process. ECG is important for detecting, localizing, and assessing the risk of arrhythmias, ischemia, and infarctions. Typically, electrodes are affixed to the patient's chest, arms, and legs in order to provide a three-dimensional view of the heart. A positive deflection is produced when the direction of myocardial depolarization is toward the electrode and a negative deflection is produced when the direction of myocardial depolarization is away from the electrode. Electrographic components of the cardi....

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All experiments were carried out in accordance with the National Institutes of Health (NIH) guidelines and Upstate Medical University Institutional Animal Care and Use Committee (IACUC). In addition, an outline of this protocol is provided in Figure 1.

1. Preparing recording equipment

  1. Connect the computer to an amplifier with a 64-pin headbox.
    NOTE: Each animal has four straight subdermal scalp pin electrodes (7 or 13 mm) for EE.......

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The method described above is capable of detecting abnormalities in the electrical conduction system of the brain and the heart as well as respiratory disturbances. A data acquisition software is used to assess the ECG morphology and detect any abnormal heart rates, conduction disturbances, or ECG rhythms (atrial/ventricular ectopic beats, and brady-/tachy-arrhythmias) (Figure 6). In addition to visualizing the ECG morphology, the traces are analyzed to quant.......

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This experimental setup facilitates detailed simultaneous video-EEG-ECG-oximetry-capnography recordings and analyses in rabbits, particularly in models of cardiac and/or neuronal diseases. The results of this article show that this method is capable of detecting seizures and arrhythmias and differentiating them from electrographic artifacts. Expected results were obtained when giving rabbits a proconvulsant, which induced seizures. The data obtained from the video-EEG recordings were able to be further analyzed to differ.......

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Authors acknowledge this study was supported by grants from the American Heart Association, American Epilepsy Society, and SUNY Upstate Department of Pharmacology.

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Name Company Catalog Number Comments
0.9% Sodium Chloride Irrigation, USP - Flexible Container PFIZER (HOSPIRA) 7983-09 Dilutant
10cc Luer Lock syringe with 20G x 1" Needle Sur-Vet SS-10L2025 Used as a flush after drug injection
4x4 gauze sponges Fisher Scientific 22-415-469 Rolled in a tube to splint ear with angiocatheter
Apple Sauce Kirkland 897971 Vehicle for oral medications
Computer Dell Optiplex 5040 Acquisition computer
E-4031 Tocris 1808 Agent known to prolong the QT interval
ECG Electrode RhythmLink RLSND116-2.5 13mm 35-degree bent (0.4 mm diameter) subdermal pin electrodes
EEG Electrode RhythmLink RLSP513 5-twist 13mm straight (0.4mm diameter) subdermal pin electrodes
EEGLAB (2020) Swartz Center for Computational Neuroscience Open Access Can perform spectral analysis of EEG
Ethernet-to-ethernet adapter Linksys USB3G16 Adapter for connecting the camera to the computer
Euthanasia-III Solution Med-Pharmex ANADA 200-280 Contains pentobarbital sodium and phenytoin sodium, controlled substance
Foam padding Generic N/A Reduces pressure applied to the neck of small rabbits by the restrainer in order to prevent the adverse cardiorespiratory effects of neck compression
Heparin Lock Flush Medline EMZ50051240 To maintain patency of angiocatheter
IR Light Bosch EX12LED-3BD-8W Facilitates recordings in the dark
LabChart Pro (2019, Version 8.1.16) ADInstruments N/A ECG Analysis
JELCO PROTECTIV Safety I.V. Catheters, 25 gauge Smiths Medical 3060 Used to catherize marginal ear vein
MATLAB (R2019b, Update 5) MathWorks N/A Required to run EEGLAB
Microphone Sony Stereo ECM-D570P Recording of audible manifestions of seizures
Micropore Medical Tape, Paper, White 3M 1530-1 Used to secure wires and create ear splint
Natus NeuroWorks Natus LC101-8 Acquisition and review software
Pentylenetetrazol (1 - 10 mg/kg always in 1mL volume) Sigma-Aldrich 88580 Dilutions prepared in saline
Photic Stimulator Grass PS22 Stimulator to control frequency, delay, duration, intensity of the light pulses
Plastic wire organizer / bundler 12Vwire.com LM-12-100-BLK Bundle wires to cut down on noise
PS 22 Photic Stimulator Grass Instruments BZA641035 Strobe light with adjustable flash frequency, delay, and intensity
PVC pipe Generic N/A Prevents small rabbits from kicking their hind legs and causing spinal injury
Quantum Amplifier Natus 13926 Amplifier / digitizer
Quantum HeadBox Amplifier Natus 22134 64-pin breakout box
Rabbit Restrainer Plas-Labs 501-TC Various size rabbit restrainers are available. 6" x 18" x 6" in this study.
Rubber pad (booster) Generic N/A Raises small rabbits up in the restrainer to prevent neck compression
SpO2 ear clip NONIN 61000 PureSAT/SpO2
SpO2 sensor adapter NONIN 13931 XPOD PureSAT/SpO2
SRG-X120 1080p PTZ Camera with HDMI, IP & 3G-SDI Output Sony SRG-X120 Impela Camera
Terumo Sur-Vet Tuberculin Syringe 1cc 25G X 5/8" Regular Luer Sur-Vet 13882 Used to inject intravenous medications
Veterinary Injection Plug Luer Lock Sur-Vet SRIP2V Injection plug for inserting the needle for intravenous medication
Webcol Alcohol Prep, Sterile, Large, 2-ply Covidien 5110 To prepare ear vein before catheterization

  1. Kaese, S., et al. The ECG in cardiovascular-relevant animal models of electrophysiology. Herzschrittmacherther Elektrophysiology. 24 (2), 84-91 (2013).
  2. Pogwizd, S. M., Bers, D. M. Rabbit models of heart disease. Drug Discovery Today: Disease Models. 5 (3), 185-193 (2008).
  3. O'Hara, T., Rudy, Y. Quantitative comparison of cardiac ventricular myocyte electrophysiology and response to drugs in human and nonhuman species. American Journal of Physiology. Heart and Circulatory Physiology. 302 (5), 1023-1030 (2012).
  4. Brunner, M., et al. Mechanisms of cardiac arrhythmias and sudden death in transgenic rabbits with long QT syndrome. Journal of Clinical Investigation. 118 (6), 2246-2259 (2008).
  5. Lengyel, C., et al. Pharmacological block of the slow component of the outward delayed rectifier current (I(Ks)) fails to lengthen rabbit ventricular muscle QT(c) and action potential duration. British Journal of Pharmacology. 132 (1), 101-110 (2001).
  6. Baczko, I., Hornyik, T., Brunner, M., Koren, G., Odening, K. E. Transgenic rabbit models in proarrhythmia research. Frontiers in Pharmacology. 11, 853 (2020).
  7. Rudy, Y., et al. Systems approach to understanding electromechanical activity in the human heart: a national heart, lung, and blood institute workshop summary. Circulation. 118 (11), 1202-1211 (2008).
  8. Zhu, Y., Ai, X., Oster, R. A., Bers, D. M., Pogwizd, S. M. Sex differences in repolarization and slow delayed rectifier potassium current and their regulation by sympathetic stimulation in rabbits. Archives. 465 (6), 805-818 (2013).
  9. Nerbonne, J. M., Nichols, C. G., Schwarz, T. L., Escande, D. Genetic manipulation of cardiac K(+) channel function in mice: what have we learned, and where do we go from here. Circulation Research. 89 (11), 944-956 (2001).
  10. Eckardt, L., et al. Drug-related torsades de pointes in the isolated rabbit heart: comparison of clofilium, d,l-sotalol, and erythromycin. Journal of Cardiovascular Pharmacology. 32 (3), 425-434 (1998).
  11. Baczko, I., Jost, N., Virag, L., Bosze, Z., Varro, A. Rabbit models as tools for preclinical cardiac electrophysiological safety testing: Importance of repolarization reserve. Progress on Biophysics and Molecular Biology. 121 (2), 157-168 (2016).
  12. Richig, J. W., Sleeper, M. M. . Electrocardiography of Laboratory Animals. , (2019).
  13. Edwards, A. G., Louch, W. E. Species-dependent mechanisms of cardiac arrhythmia: A cellular focus. Clinical Medicine Insights. Cardiology. 11, 1179546816686061 (2017).
  14. Salama, G., London, B. Mouse models of long QT syndrome. Journal of Physiology. 578, 43-53 (2007).
  15. Zhang, Y., Wu, J., King, J. H., Huang, C. L., Fraser, J. A. Measurement and interpretation of electrocardiographic QT intervals in murine hearts. American Journal of Physiology. Heart and Circulation Physiology. 306 (11), 1553-1557 (2014).
  16. Auerbach, D. S., et al. Altered cardiac electrophysiology and SUDEP in a model of dravet syndrome. PLoS One. 8 (10), 15 (2013).
  17. Aiba, T., Tomaselli, G. F. Electrical remodeling in the failing heart. Current Opinion in Cardiology. 25 (1), 29-36 (2010).
  18. Auerbach, D. S., et al. Genetic biomarkers for the risk of seizures in long QT syndrome. Neurology. 87 (16), 1660-1668 (2016).
  19. Anderson, L. L., et al. Antiepileptic activity of preferential inhibitors of persistent sodium current. Epilepsia. 55 (8), 1274-1283 (2014).
  20. Johnson, J. N., et al. Identification of a possible pathogenic link between congenital long QT syndrome and epilepsy. Neurology. 72 (3), 224-231 (2009).
  21. Devinsky, O., Hesdorffer, D. C., Thurman, D. J., Lhatoo, S., Richerson, G. Sudden unexpected death in epilepsy: epidemiology, mechanisms, and prevention. Lancet Neurology. 15 (10), 1075-1088 (2016).
  22. Bagnall, R. D., et al. Exome-based analysis of cardiac arrhythmia, respiratory control, and epilepsy genes in sudden unexpected death in epilepsy. Annals in Neurology. 79 (4), 522-534 (2016).
  23. Frasier, C. R., et al. Channelopathy as a SUDEP biomarker in dravet syndrome patient-derived cardiac myocytes. Stem Cell Reports. 11 (3), 626-634 (2018).
  24. Glasscock, E. Genomic biomarkers of SUDEP in brain and heart. Epilepsy and Behavior. 38, 172-179 (2014).
  25. Olejniczak, P. Neurophysiologic basis of EEG. Journal of Clinical Neurophysiology. 23 (3), 186-189 (2006).
  26. Gastaut, H., Hunter, J. An experimental study of the mechanism of photic activation in idiopathic epilepsy. Electroencephalography and Clinical Neurophysiology. 2 (3), 263-287 (1950).
  27. Fisher, R. S., et al. Photic- and pattern-induced seizures: A review for the Epilepsy Foundation of America Working Group. Epilepsia. 46 (9), 1426-1441 (2005).
  28. Specchio, N., et al. Diagnosing photosensitive epilepsy: fancy new versus old fashioned techniques in patients with different epileptic syndromes. Brain Development. 33 (4), 294-300 (2011).
  29. Kasteleijn-Nolst Trenite, D., et al. Methodology of photic stimulation revisited: updated European algorithm for visual stimulation in the EEG laboratory. Epilepsia. 53 (1), 16-24 (2012).
  30. Mishra, V., Gautier, N. M., Glasscock, E. Simultaneous video-EEG-ECG monitoring to identify neurocardiac dysfunction in mouse models of epilepsy. Journal of Visualized Experiments. (131), e57300 (2018).
  31. Green, J. D., Maxwell, D. S., Schindler, W. J., Stumpf, C. Rabbit EEG "theta" rhythm: Its anatomical source and relation to activity in single neurons. Journal of Neurophysiology. 23 (4), 403-420 (1960).
  32. Petersen, J., Diperri, R., Himwich, W. A. The comparative development of the EEG in rabbit, cat and dog. Electroencephalography and Clinical Neurophysiology. 17, 557-563 (1964).
  33. Strain, G. M., Van Meter, W. G., Brockman, W. H. Elevation of seizure thresholds: a comparison of cerebellar stimulation, phenobarbital, and diphenylhydantoin. Epilepsia. 19 (5), 493-504 (1978).
  34. Cheng, Y., et al. Effectiveness of retigabine against levobupivacaine-induced central nervous system toxicity: A prospective, randomized animal study. Journal of Anesthesia. 30 (1), 109-115 (2016).
  35. Nascimento, F. A., et al. Pulmonary and cardiac pathology in sudden unexpected death in epilepsy (SUDEP). Epilepsy and Behavior. 73, 119-125 (2017).
  36. Buchanan, G. F. Impaired CO2-Induced Arousal in SIDS and SUDEP. Trends in Neuroscience. 42 (4), 242-250 (2019).
  37. Van Egmond, P., Binnie, C. D., Veldhuizen, R. The effect of background illumination on sensitivity to intermittent photic stimulation. Electroencephalography and Clinical Neurophysiology. 48 (5), 599-601 (1980).
  38. Harding, G. F., Fylan, F. Two visual mechanisms of photosensitivity. Epilepsia. 40 (10), 1446-1451 (1999).
  39. Kuwada, S., Stanford, T. R., Batra, R. Interaural phase-sensitive units in the inferior colliculus of the unanesthetized rabbit: effects of changing frequency. Journal of Neurophysiology. 57 (5), 1338-1360 (1987).
  40. Kalume, F., et al. Sudden unexpected death in a mouse model of Dravet syndrome. Journal of Clinical Investigation. 123 (4), 1798-1808 (2013).
  41. Xiang, C., et al. Threshold for maximal electroshock seizures (MEST) at three developmental stages in young mice. Zoology Research. 40 (3), 231-235 (2019).
  42. Ross, K. C., Coleman, J. R. Developmental and genetic audiogenic seizure models: behavior and biological substrates. Neuroscience and Biobehavior Reviews. 24 (6), 639-653 (2000).
  43. Faingold, C. L., Randall, M., Tupal, S. DBA/1 mice exhibit chronic susceptibility to audiogenic seizures followed by sudden death associated with respiratory arrest. Epilepsy and Behavior. 17 (4), 436-440 (2010).

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