Name: Advanced Cardiac Rhythm Management by Applying Optogenetic Multi-Site Photostimulation in Murine Hearts (Video) | JoVE Journal
Uploaded: 2021-08-26T13:00:00.000+00:00
Duration: 8 min 43 s
Description: Learn optogenetic photostimulation of Langendorff-perfused murine hearts using a micro-LED array for precise cardiac rhythm control. This method enables targeted arrhythmia termination with real-time optical mapping.

ai_agent_IP

ai_agent

maintenance

instantTrialDurationMinutes

isBiopharma

enableNewMyJoveSectionDesign

educationFeatures

seoMetaApi

ai_copilot_institution_blacklist

ai_chat_institution_blacklist

freeAccessWindow

ssoRoleCapture

scheduledMaintenanceBanner

joveLabsFeatures

disable_autoplay_for_bots

extended_trial_duration

5 minutes

The authors would like to thank Marion Kunze and Tina Althaus for their excellent technical support during experiments. The research leading to the results has received funding from the European Community&#39;s Seventh Framework Programme FP7/2007-2013 under grant agreement number HEALTH-F2-2009-241526. Support was also provided by the German Center for Cardiovascular Research, DZHK e.V. (Project MD28), partner site Goettingen, the German Research Foundation CRC 1002 (project C03), and the Max Planck Society. This work was partly supported by BrainLinks-BrainTools, Cluster of Excellence funded by the German Research Foundation (DFG, grant number EXC 1086).

All experiments strictly followed the animal welfare regulation, in agreement with German legislation, local stipulations, and in accordance with recommendations of the Federation of European Laboratory Animal Science Associations (FELASA). The application for approval of animal experiments has been approved by the responsible animal welfare authority, and all experiments were reported to our animal welfare representatives.
1. Experiment preparation and materials
<ol>
	<li>Optical mapping setup 
	NOTE: The optical setup, as well as the electrical setup, are shown in Figure 1. All components used in the optical and electrical setup are listed in detail in the Table of Materials.
	<ol>
		<li>Use LED 1 and LED 2 for induction of arrhythmia and backup defibrillation. Choose high power LEDs with a wavelength &#955;blue near 475 nm, which is the peak of the excitation wavelength of ChR26. To further narrow the optical spectrum, use a 470 &#177; 20 nm bandpass filter. 
		NOTE: In this work, LED 1 and LED 2 have a typical radiant flux of 3.9 to 5.3 W, according to the datasheet20.</li>
		<li>Illuminate the epicardium for optical mapping with a high-power red LED (LED 3 in Figure 1), which emits light with a center wavelength of &#955;red&#160;=&#160;625&#160;nm and a radiant flux of 700 mW21. The red light is filtered with a 628 &#177; 20 nm bandpass filter and reflected by a long pass dichroic mirror (DM) with a cutoff wavelength of &#955;DM = 685 nm.</li>
		<li>Use an emission filter with &#955;filter-cam&#160;=&#160;775&#160;&#177;&#160;70&#160;nm in front of the camera objective to only record the fluorescence emission of the cardiac activity. Use a fast objective that is well suited for low light applications. 
		NOTE: The frequency of fibrillation of a mouse heart ranges from 20 to 35 Hz; therefore, use a fast enough camera to record with a frequency of 1 to 2 kHz, or even higher.</li>
	</ol>
	</li>
	<li>Micro-LED array 
	NOTE: The micro-LED arrays applied here are realized using microsystems processing as further detailed elsewhere16,17.
	<ol>
		<li>Spin coat a 5 &#181;m-thick polyimide (PI) layer onto 4-inch silicon substrates (single-side polished, 525-&#181;m thick).</li>
		<li>Cure this PI layer at a maximum temperature of 450&#160;&#176;C under a nitrogen atmosphere. Keep the maximum temperature constant for 10 min.</li>
		<li>Deposit and pattern an image reversal photoresist (PR) using ultraviolet (UV) lithography and sputter deposit a 250-nm thin platinum layer (Pt).</li>
		<li>Thicken this Pt-based metallization by electroplating a 1 &#181;m-thick gold (Au) layer&#160;with the patterned PR serving as a masking layer.</li>
		<li>Before spin-coating&#160;a second PI layer, expose the wafer with its first PI layer and the Au-electroplated metallization to an oxygen plasma that chemically activates the surface of the PI layer.</li>
		<li>Cure the second PI layer again at 450 &#176;C, apply UV lithography to pattern a PR layer and open the contact pads of the array for the micro-LED chips and the interfacing printed circuit board (PCB) by reactive ion etching (RIE) using the patterned PR as a masking layer. 
		NOTE: In this RIE process steps, it is recommended to apply 200 W and 100 W for 10 and 30 min, respectively, to define the contact pad openings as well as the outer shape of the two-dimensional (2D) micro-LED array.</li>
		<li>Strip the PR using solvents and plasma etching. Further thicken the contact pads by electroplating an additional 6 &#181;m-thick gold layer.</li>
		<li>Attach the micro-LED chips to the contact pads using a flip-chip bonder.</li>
		<li>Activate the PI surface in an oxygen plasma and underfill the micro-LED chips with a solvent-free adhesive. Cure then the adhesive for 12 h at 120 &#176;C.</li>
		<li>To encapsulate the micro-LED chips, perform another plasma treatment with Argon and apply a thin fluoropolymer layer manually. Pre-cure this layer at 80 &#176;C for 1 h.</li>
		<li>Manually apply silicone as the final encapsulation layer after exposing the micro-LED array to an oxygen plasma, used to improve the silicon adhesion to the underlying fluoropolymer layer. Cure the silicone layer at 80 &#176;C and 180 &#176;C for 1 h each. These final curing steps also cure the fluoropolymer layer completely.</li>
		<li>Solder the contact pads of the PI substrate to a printed circuit board which carries strip connectors for interconnection of the array to an external instrumentation. Cover the solder pads on the PCB using an adhesive.</li>
	</ol>
	</li>
	<li>Electrical setup
	<ol>
		<li>Use electrodes suited for recording an electrocardiogram (ECG), e.g., silver/silver-chloride electrodes or Monophasic Action Potential (MAP) electrodes and an ECG amplifier to monitor the electrical activity of the heart continuously. Furthermore, use an appropriate acquisition device (AD) to record all electrical signals obtained.</li>
		<li>Choose a well-suited driver for the high-power LEDs (LED 1, LED 2, and LED 3), which can manage the maximum current applied to each device. Use an arbitrary function generator (AFG) to control the output of the LED drivers accurately.</li>
		<li>Use a multi-channel LED driver to control the current flowing through the micro-LED array. An AFG with multiple outputs is as well suitable for this task. 
		NOTE: It is advisable to choose LED drivers&#160;limiting the current to the maximum current of the micro-LED, otherwise the diodes might get damaged. One example of a multi-channel micro-LED driver is described in another work18. If necessary, the AFG or any other LED driver might be connected to a computer to remotely control the micro-LED settings. If this is the case connect the LED driver to the computer with the communication protocol of your choice, e.g., General Purpose Interface Bus (GPIB) or a serial connection.</li>
	</ol>
	</li>
</ol>
&#160; &#160;

2. Experimental procedures
<ol>
	<li>Solution preparation
	<ol>
		<li>Prepare Tyrode&#39;s solution: 130&#160;mM&#160;NaCl, 4&#160;mM&#160;KCl, 1&#160;mM&#160;MgCl2, 24&#160;mM&#160;NaHCO3, 1.8&#160;mM&#160;CaCl2, 1.2&#160;mM&#160;KH2PO4, 5.6&#160;mM&#160;Glucose, 0.1%&#160;BSA/Albumin.</li>
		<li>Prepare Low-K+ Tyrode&#39;s solution: Low-K+ Tyrode&#39;s is made in the same way as regular Tyrode&#39;s solution except that only half the amount of KCl is added (2 mM instead of 4 mM KCl). 
		&#8203;NOTE: For an experiment lasting 3 h usually 2-3 L of Low-K+ Tyrode&#39;s (additionally mixed with Blebbistatin (Step 2.1.5) if optical mapping is performed) and 1-2 L of regular Tyrode&#39;s are sufficient.</li>
		<li>Add Pinacidil to the Low-K+ Tyrode&#39;s solution to ease the process of arrhythmia induction, as described in22, to obtain a 100 mM concentration. Wear protective laboratory gloves when handling Pinacidil.</li>
		<li>Prepare 1 mL of 50 &#181;M DI-4-ANBDQPQ with regular Tyrode&#39;s solution. Protect the dye from light to prevent photobleaching.</li>
		<li>Make a 10 mM stock solution of Blebbistatin. For optical mapping, mix Blebbistatin with the 100 mM Pinacidil-Tyrode&#39;s-solution (Step 2.1.3) to obtain a 5&#160;&#181;M&#160;solution. Wear protective laboratory gloves when handling Blebbistatin. 
		NOTE: Keep both the dye and the Blebbistatin solution aside until optical mapping begins.</li>
	</ol>
	</li>
	<li>Langendorff perfusion 
	NOTE: The setup consists of two reservoirs for the two Tyrode&#39;s solutions. They are connected to a bubble trap via tubes with three-way cocks. The heart is later attached to the bubble trap by a Luer lock connector, and it is then suspended in a hexagonal water bath. The water bath is, in turn, connected to a waste container to collect the used Tyrode&#39;s solution.
	<ol>
		<li>Clean all tubes before every experiment with fully demineralized water.</li>
		<li>Aerate both Tyrode&#39;s solutions with Carbogen (5% CO2 and 95% O2) for 30 min at room temperature before beginning of the experiment. Adjust the pH value of the Tyrode&#39;s solutions to 7.4 with NaOH.</li>
		<li>Fill 500 mL of each Tyrode&#39;s solution in the corresponding reservoir and de-aerate the tubes as well as the bubble trap by running Tyrode&#39;s solution through the perfusion system until no more trapped air bubbles are seen in the tubes or in the bubble trap.</li>
		<li>Continue aerating the Tyrode&#39;s solutions during the whole experiment in the reservoirs with Carbogen to ensure that the pH of the perfusate remains stable later during perfusion.</li>
		<li>Heat the perfusion system to 37 &#176;C with a water heat pump. Keep the perfusate temperature constant within the water bath by using an additional heating element such as a waterproof heating cable. 
		NOTE: During the experiment, it is crucial to refill the Tyrode&#39;s reservoirs before they run empty. Otherwise, air bubbles can enter the heart, which can clog the vessels and lead to ischemia.</li>
	</ol>
	</li>
	<li>Mouse Preparation
	<ol>
		<li>Inject subcutaneously 0.1 mL of 500 I.E. Heparin 30&#160;min&#160;before heart isolation procedure.</li>
		<li>Fill a 6 cm Petri dish and a 2 mL syringe with ice-cold Tyrode&#39;s solution. Place under the stereoscopic microscope.</li>
		<li>Perform short time anesthesia of mice by a saturated Isoflurane environment for 2 min and immediate cervical dislocation afterwards. 
		&#8203;NOTE: In order to verify sufficient anesthesia a check for the negative inter-toe reflex is absolutely necessary.</li>
		<li>Open the chest, remove the heart, as described elsewhere23, and place it into the 6 cm Petri dish with ice-cold Tyrode&#39;s solution. Cardiac beating will be diminished due to temperature drop.</li>
		<li>Do the fine preparation under a stereoscopic microscope, as detailed elsewhere23. Attach the aorta onto the blunt needle and fix the vessel with suture material.</li>
		<li>As a control, inject ice-cold Tyrode&#39;s solution through the needle into the heart and check that the heart is tightly mounted. This step also rinses the remaining blood out of the heart.</li>
		<li>Transfer the mounted heart to the perfusion system. Ensure that the perfusate is flowing to prevent air from entering the heart while connecting the needle with the bubble trap. Check that the heart is covered with Tyrode&#39;s solution in the water bath. Steps 2.3.4, 2.3.5, and 2.3.7 are illustrated in Figure 2.</li>
		<li>Ensure that the heart starts beating within a few minutes. Let the heart adapt to the perfusion setup for 15 to 20 min, then switch to low-K+ Tyrode&#39;s solution with Pinacidil (Step 2.1.3) respectively low-K+ Tyrode&#39;s solution with Pinacidil and Blebbistatin (Step 2.1.5) if optical mapping is to be performed.</li>
	</ol>
	</li>
	<li>Arrhythmia induction and optical defibrillation
	<ol>
		<li>Place one of the ECG electrodes as close as possible to the heart surface to ensure good signal quality. Suspend the second ECG electrode in the Tyrode&#39;s solution. Make sure that the ECG acquired is being recorded by the AD of choice.</li>
		<li>Place the micro-LED array on the area of interest of the study, for example, onto the left ventricle.</li>
		<li>Change the perfusion to low-K+ Tyrode&#39;s with Pinacidil and perfuse the heart for 15 to 30 min.</li>
		<li>To induce arrhythmia, illuminate the heart with LED 1 and LED 2 with a train of 20 to 50 light pulses with a frequency find of 25 to 35 Hz, pulse duration Wind of 2 to 15 ms, and light intensity LIopt_ind of 2.8 mW mm-2.</li>
		<li>Repeat the process until arrhythmia is induced. 
		NOTE: Arrhythmias are easy to identify in the ECG signal because the frequency and morphology of the signal differ from normal sinus rhythm. Should the arrhythmia terminate within the next 5 s, classify it as self-terminated, and start a new induction attempt.</li>
		<li>Once a sustained arrhythmia is visually detected, apply a burst of pulses with different widths Wdef and frequencies fdef, using three, six, or nine micro-LEDs of the array at a pulsing current Ipulse of 15&#160;mA yielding to a light intensity LI&#181;LED&#160;=&#160;33.31&#160;&#177;&#160;2.05&#160;mW&#160;mm-2.</li>
		<li>Should the arrhythmia keep ongoing after five micro-LED array-based defibrillation trials, classify the attempt as unsuccessful and start backup defibrillation.</li>
		<li>For backup defibrillation, use LED 1 and LED 2 using the same timing parameters as set for the micro-LED array. 
		&#8203;NOTE: Because the heart is exposed to ischemic and metabolic stress over the whole experimental period, it is possible that termination attempts of arrhythmia are unsuccessful even with backup defibrillation. Whenever this happens, change the perfusion solution to the regular Tyrode&#39;s and let the heart recover for 5 to 10 min. When the ECG returns to sinus rhythm, repeat the protocol from Step 2.4.3 again.</li>
	</ol>
	</li>
	<li>Optical Mapping
	<ol>
		<li>Perfuse the heart with the Blebbistatin solution prepared in Step 2.1.5 and wait until mechanical uncoupling occurs. This is accomplished when the heart stops beating, but an ECG signal is still measurable. 
		NOTE: Mixing the Blebbistatin solution to the mentioned concentration and keeping the heart perfused with this solution maintains the cardiac mechanical activity uncoupled from the electrical activity during the whole experiment.</li>
		<li>Give the 1 mL voltage dye DI-4-ANBDQPQ (prepared in Step 2.1.4) as a bolus in the bubble trap of the Langendorff perfusion. Wait for 5 to 10 min to allow the dye to perfuse the heart uniformly. 
		&#8203;NOTE: Avoid photobleaching of the dye by turning off the red light whenever no recording is being made. If the signal-to-noise ratio of the recording becomes too small (acquired signal is too noisy), repeat steps 2.1.4 and 2.5.2.</li>
		<li>Focus the camera onto the heart surface, turn on LED 3, and apply 1.27 mW mm-2 optical power.</li>
		<li>Turn off the laboratory lights and start recording. Ensure that an optical signal is being acquired by comparing the frequency of the obtained signal to the frequency of the recorded ECG. This ensures that the obtained optical signal is purely related to the electrical activity of the heart. 
		NOTE: Since the fluorescence light emitted by the dye is very week, optical mapping is done in a dark room. This avoids signal interference from any other light sources.</li>
	</ol>
	</li>
</ol>

<ol><li>Davidenko, J. M., Pertsov, A. V., Salamonsz, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Stationary+and+drifting+spiral+waves+of+excitation+in+isolated+cardiac+muscle%5BTitle%5D)+AND+%22Nature%22%5BJournal%5D)">Stationary and drifting spiral waves of excitation in isolated cardiac muscle.</a> Nature. 355, 349-351 (1992).</li>
<li>Fenton, F. H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Termination+of+atrial+fibrillation+using+pulsed+low-energy+far-field+stimulation%5BTitle%5D)+AND+%22Circulation%22%5BJournal%5D)">Termination of atrial fibrillation using pulsed low-energy far-field stimulation.</a> Circulation. 120 (6), 467-476 (2009).</li>
<li>Luther, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Low-energy+control+of+electrical+turbulence+in+the+heart%5BTitle%5D)+AND+%22Nature%22%5BJournal%5D)">Low-energy control of electrical turbulence in the heart.</a> Nature. 475, 235-239 (2011).</li>
<li>Pumir, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Wave+emission+from+heterogeneities+opens+a+way+to+controlling+chaos+in+the+heart%5BTitle%5D)+AND+%22Physical+Review+Letters%22%5BJournal%5D)">Wave emission from heterogeneities opens a way to controlling chaos in the heart.</a> Physical Review Letters. 99, 208101(2007).</li>
<li>Deisseroth, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Optogenetics%5BTitle%5D)+AND+%22Nature+Methods%22%5BJournal%5D)">Optogenetics.</a> Nature Methods. 8, 26-29 (2011).</li>
<li>Boyden, E. S., Zhang, F., Bamberg, E., Nagel, G., Deisseroth, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Millisecond-timescale%2C+genetically+targeted+optical+control+of+neural+activity%5BTitle%5D)+AND+%22Nature+Neuroscience%22%5BJournal%5D)">Millisecond-timescale, genetically targeted optical control of neural activity.</a> Nature Neuroscience. 8, 1263-1268 (2005).</li>
<li>Nagel, G., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Channelrhodopsin-2%2C+a+directly+light-gated+cation-selective+membrane+channel%5BTitle%5D)+AND+%22Proceedings+of+the+National+Academy+of+Sciences%22%5BJournal%5D)">Channelrhodopsin-2, a directly light-gated cation-selective membrane channel.</a> Proceedings of the National Academy of Sciences. 100 (24), 13940-13945 (2003).</li>
<li>Bruegmann, T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Optogenetic+control+of+heart+muscle+in+vitro+and+in+vivo%5BTitle%5D)+AND+%22Nature+Methods%22%5BJournal%5D)">Optogenetic control of heart muscle in vitro and in vivo.</a> Nature Methods. 7, 897-900 (2010).</li>
<li>Natasha, G., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((et+al.Channelrhodopsins%3A+visual+regeneration+and+neural+activation+by+a+light+switch%5BTitle%5D)+AND+%22New+Biotechnology%22%5BJournal%5D)">et al.Channelrhodopsins: visual regeneration and neural activation by a light switch.</a> New Biotechnology. 30 (5), 461-474 (2013).</li>
<li>Zhang, F., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Multimodal+fast+optical+interrogation+of+neural+circuitry%5BTitle%5D)+AND+%22Nature%22%5BJournal%5D)">Multimodal fast optical interrogation of neural circuitry.</a> Nature. 446, 633-639 (2007).</li>
<li>Alilain, W. J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Light-induced+rescue+of+breathing+after+spinal+cord+injury%5BTitle%5D)+AND+%22Journal+of+Neuroscience%22%5BJournal%5D)">Light-induced rescue of breathing after spinal cord injury.</a> Journal of Neuroscience. 28 (46), 11862-11870 (2008).</li>
<li>Ahmad, A., Ashraf, S., Komai, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Optogenetics+applications+for+treating+spinal+cord+injury%5BTitle%5D)+AND+%22Asian+Spine+Journal%22%5BJournal%5D)">Optogenetics applications for treating spinal cord injury.</a> Asian Spine Journal. 9 (2), 299-305 (2015).</li>
<li>Dieter, A., Keppeler, D., Moser, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Towards+the+optical+cochlear+implant%3A+Optogenetic+approaches+for+hearing+restoration%5BTitle%5D)+AND+%22EMBO+Molecular+Medicine%22%5BJournal%5D)">Towards the optical cochlear implant: Optogenetic approaches for hearing restoration.</a> EMBO Molecular Medicine. 12 (4), e11618(2020).</li>
<li>Keppeler, D., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Multichannel+optogenetic+stimulation+of+the+auditory+pathway+using+microfabricated+LED+cochlear+implants+in+rodents%5BTitle%5D)+AND+%22Science+Translational+Medicine%22%5BJournal%5D)">Multichannel optogenetic stimulation of the auditory pathway using microfabricated LED cochlear implants in rodents.</a> Science Translational Medicine. 12 (553), eabb8086(2020).</li>
<li>Verhoefen, M. K., Bamann, C., Blöcher, R., Förster, U., Bamberg, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((The+photocycle+of+channelrhodopsin-2%3A+ultrafast+reaction+dynamics+and+subsequent+reaction+steps%5BTitle%5D)+AND+%22ChemPhysChem%22%5BJournal%5D)">The photocycle of channelrhodopsin-2: ultrafast reaction dynamics and subsequent reaction steps.</a> ChemPhysChem. 11 (14), 3113-3122 (2010).</li>
<li>Schwaerzle, M., Elmlinger, P., Paul, O., Ruther, P. Miniaturized tool for optogenetics based on an LED and an optical fiber interfaced by a silicon housing. 36th Annual Internation Conference IEEE Engineering in Medicine and Biology Society, Chicago, IL, , 5252-5255 (2014).</li>
<li>Schwaerzle, M., Elmlinger, P., Paul, O., Ruther, P. Miniaturized 3 x 3 optical fiber array for optogenetics with integrated 460 nm light sources and flexible electrical interconnection. 28th IEEE Proceedigns. MEMS, Estoril, , 162-165 (2015).</li>
<li>Diaz-Maue, L., Schwaerzle, M., Ruther, P., Luther, S., Richter, C. Follow the light - From low-energy defibrillation to multi-site photostimulation. 40thAnnual International Conference of IEEE Engineering in Medicine and Biology Society, Honolulu, HI, , 4832-4835 (2018).</li>
<li>Zgierski-Johnston, C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Cardiac+pacing+using+transmural+multi-LED+probes+in+channelrhodopsin-expressing+mouse+hearts%5BTitle%5D)+AND+%22Progress+in+Biophysics+and+Molecular+Biology%22%5BJournal%5D)">Cardiac pacing using transmural multi-LED probes in channelrhodopsin-expressing mouse hearts.</a> Progress in Biophysics and Molecular Biology. , 51-61 (2020).</li>
<li>mouser.de, LED Engin, [Online]. , Available: 
 <a target="_blank" href=" https://www.mouser.de/datasheet/2/228/5412893-LED_2520Engin_Datasheet_LuxiGen_LZ4-00B208-
1531969.pdf"> https://www.mouser.de/datasheet/2/228/5412893-LED_2520Engin_Datasheet_LuxiGen_LZ4-00B208-
1531969.pdf</a> (2020).</li>
<li>thorlabs.com, thorlabs, [Online]. , Available: 
 <a target="_blank" href=" https://www.thorlabs.com/_sd.cfm?fileName=25135-S01.pdf&partNumber=M625L3"> https://www.thorlabs.com/_sd.cfm?fileName=25135-S01.pdf&partNumber=M625L3</a> (2020).</li>
<li>Bruegmann, T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Optogenetic+defibrillation+terminates+ventricular+arrhythmia+in+mouse+hearts+and+human+simulations%5BTitle%5D)+AND+%22Journal+of+Clinical+Investigation%22%5BJournal%5D)">Optogenetic defibrillation terminates ventricular arrhythmia in mouse hearts and human simulations.</a> Journal of Clinical Investigation. 126 (10), 3894-3904 (2016).</li>
<li>Richter, C., Christoph, J., Lehnart, S. E., Luther, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Optogenetic+light+crafting+tools+for+the+control+of+cardiac+arrhythmias%5BTitle%5D)+AND+%22Methods+in+Molecular+Biology%22%5BJournal%5D)">Optogenetic light crafting tools for the control of cardiac arrhythmias.</a> Methods in Molecular Biology. 1408, 293-302 (2016).</li>
<li>Quiñonez Uribe, R. A., Luther, S., Diaz-Maue, L., Richter, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Energy-reduced+arrhythmia+termination+using+global+photostimulation+in+optogenetic+murine+hearts%5BTitle%5D)+AND+%22Frontiers+in+Physiology%22%5BJournal%5D)">Energy-reduced arrhythmia termination using global photostimulation in optogenetic murine hearts.</a> Frontiers in Physiology. 9 (1651), (2018).</li>
<li>Moreno, I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((LED+irradiance+pattern+at+short+distances%5BTitle%5D)+AND+%22Applied+Optics%22%5BJournal%5D)">LED irradiance pattern at short distances.</a> Applied Optics. 59 (1), 190-195 (2020).</li>
<li>Predicting unpinning success rates for a pinned spiral in an excitable medium. Behrend, A., Bittihn, P., Luther, S. Computing in Cardiology, Belfast, , 345-348 (2010).</li>
<li>Kappadan, V., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((High-resolution+optical+measurement+of+cardiac+restitution%2C+contraction%2C+and+fibrillation+dynamics+in+beating+vs.+blebbistatin-uncoupled+isolated+rabbit+hearts%5BTitle%5D)+AND+%22Frontiers+in+Physiology%22%5BJournal%5D)">High-resolution optical measurement of cardiac restitution, contraction, and fibrillation dynamics in beating vs. blebbistatin-uncoupled isolated rabbit hearts.</a> Frontiers in Physiology. 11 (464), (2020).</li>
<li>Christoph, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Electromechanical+vortex+filaments+during+cardiac+fibrillation%5BTitle%5D)+AND+%22Nature%22%5BJournal%5D)">Electromechanical vortex filaments during cardiac fibrillation.</a> Nature. 555, 667-672 (2018).</li>
<li>O'Shea, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Cardiac+optogenetics+and+optical+mapping+-+Overcoming+spectral+congestion+in+all-optical+cardiac+electrophysiology%5BTitle%5D)+AND+%22Frontiers+in+Physiology%22%5BJournal%5D)">Cardiac optogenetics and optical mapping - Overcoming spectral congestion in all-optical cardiac electrophysiology.</a> Frontiers in Physiology. 10 (182), (2019).</li>
<li>Aras, K. K., Faye, N. R., Cathey, B., Efimov, I. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Critical+volume+of+human+myocardium+necessary+to+maintain+ventricular+fibrillation%5BTitle%5D)+AND+%22Circulation%3A+Arrhythmia+and+Electrophysiology%22%5BJournal%5D)">Critical volume of human myocardium necessary to maintain ventricular fibrillation.</a> Circulation: Arrhythmia and Electrophysiology. 11 (11), e006692(2018).</li>
<li>Trayanova, N., Doshi, A. N., Prakosa, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((How+personalized+heart+modeling+can+help+treatment+of+lethal+arrhythmias%3A+A+focus+on+ventricular+tachycardia+ablation+strategies+in+post-infarction+patients%5BTitle%5D)+AND+%22Wiley+Interdisciplinary+Reviews+in+System+Biology+and+Medicine%22%5BJournal%5D)">How personalized heart modeling can help treatment of lethal arrhythmias: A focus on ventricular tachycardia ablation strategies in post-infarction patients.</a> Wiley Interdisciplinary Reviews in System Biology and Medicine. 12 (3), 1477(2020).</li>
<li>Bingen, B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Light-induced+termination+of+spiral+wave+arrhythmias+by+optogenetic+engineering+of+atrial+cardiomyocytes%5BTitle%5D)+AND+%22Cardiovascular+Research%22%5BJournal%5D)">Light-induced termination of spiral wave arrhythmias by optogenetic engineering of atrial cardiomyocytes.</a> Cardiovascular Research. 104 (1), 194-205 (2014).</li>
<li>Burton, R. A. B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Optical+control+of+excitation+waves+in+cardiac+tissue%5BTitle%5D)+AND+%22Nature+Photonics%22%5BJournal%5D)">Optical control of excitation waves in cardiac tissue.</a> Nature Photonics. 9 (12), 813-816 (2015).</li>
<li>Dura, M., Schröder-Schetelig, J., Luther, S., Lehnart, S. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Toward+panoramic+in+situ+mapping+of+action+potential+propagation+in+transgenic+hearts+to+investigate+initiation+and+therapeutic+control+of+arrhythmias%5BTitle%5D)+AND+%22Frontiers+in+Physiology%22%5BJournal%5D)">Toward panoramic in situ mapping of action potential propagation in transgenic hearts to investigate initiation and therapeutic control of arrhythmias.</a> Frontiers in Physiology. 5, 337(2014).</li>
<li>Crocini, C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Optogenetics+design+of+mechanistically-based+stimulation+patterns+for+cardiac+defibrillation%5BTitle%5D)+AND+%22Science+Reports%22%5BJournal%5D)">Optogenetics design of mechanistically-based stimulation patterns for cardiac defibrillation.</a> Science Reports. 6 (35628), (2016).</li>
<li>Nyns, E. C. A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Optogenetic+termination+of+ventricular+arrhythmias+in+the+whole+heart%3A+towards+biological+cardiac+rhythm+management%5BTitle%5D)+AND+%22European+Heart+Journal%22%5BJournal%5D)">Optogenetic termination of ventricular arrhythmias in the whole heart: towards biological cardiac rhythm management.</a> European Heart Journal. 38 (27), 2132-2136 (2017).</li>
<li>Wilde, A. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((K%2Batp+channel+opening+and+arrhythmogenesis%5BTitle%5D)+AND+%22Journal+of+Cardiovascular+Pharmacology%22%5BJournal%5D)">K+atp channel opening and arrhythmogenesis.</a> Journal of Cardiovascular Pharmacology. 24 (4), 35-40 (1994).</li>
<li>Christoph, J., Luther, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Marker-free+tracking+for+motion+artifact+compensation+and+deformation+measurements+in+optical+mapping+videos+of+contracting+hearts%5BTitle%5D)+AND+%22Frontiers+in+Physiology%22%5BJournal%5D)">Marker-free tracking for motion artifact compensation and deformation measurements in optical mapping videos of contracting hearts.</a> Frontiers in Physiology. 9 (1483), (2018).</li>
<li>Christoph, J., Schröder-Schetelig, J., Luther, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Electromechanical+optical+mapping%5BTitle%5D)+AND+%22Progress+in+Biophysics+and+Molecular+Biology%22%5BJournal%5D)">Electromechanical optical mapping.</a> Progress in Biophysics and Molecular Biology. 130(B), 150-169 (2017).</li>
</ol>

The authors do not declare any conflict of interests.

A successful treatment of cardiac tachyarrhythmias is key to cardiac therapy. However, the biophysical mechanisms underlying arrhythmia initiation, perpetuation and termination are not fully understood. Therefore, cardiac research aims to optimize electrical shock therapy towards a more gentle termination of arrhythmias, thereby increasing the quality of life of patients28,29,30,31. Low energy ...

Early investigations of the spatial-temporal dynamics during arrhythmia revealed that the complex electrical patterns during cardiac fibrillation are driven by vortex-like rotating excitation waves1. This finding gave new insights into the underlying mechanisms of arrhythmias, which then led to the development of novel electrical termination therapies based on multi-site excitation of the myocardium2,3,4. However, treatments using electric field stimulation are non-local and may innervate all surrounding excitable cells, including muscle tissue, causing cellular and tissue damage, as well as intolerable pain. In contrast to electrical therapies, optogenetic approaches provide a specific and tissue-protective technique for evoking cardiomyocyte action potentials with high spatial and temporal precision. Therefore, optogenetic stimulation has the potential for minimal invasive control of the chaotic activation patterns during cardiac fibrillation.
The introduction of the light-sensitive ion channel channelrhodopsin-2 (ChR2) into excitable cells via genetic manipulation5,6,7, enabled the depolarization of the membrane potential of excitable cells using photostimulation. Several medical applications, including the activation of neuronal networks, the control of cardiac activity, the restoration of vision and hearing, the treatment of spinal cord injuries, and others8,9,10,11,12,13,14 have been developed. The application of ChR2 in cardiology has significant potential due to its millisecond response time15, making it well suited for the targeted control of arrhythmic cardiac dynamics.
In this study, multi-site photostimulation of intact hearts of a transgenic mouse model is shown. In summary, a transgenic alpha-MHC-ChR2 mouse line was established within the scope of the European Community&#39;s Seventh Framework Programme FP7/2007-2013 (HEALTH-F2-2009-241526) and kindly provided by Prof. S. E. Lehnart. In general, transgenic adult male C57/B6/J, expressing Cre-recombinase under control of alpha-MHC were paired to mate with female B6.Cg-Gt(ROSA)26Sortm27.1(CAG-COP4*H134R/tdTomato)Hye/J. Since the cardiac STOP cassette was deleted in the second-generation, the offspring showed a stable MHC-ChR2 expression and was used to maintain cardiac photosensitive colonies. All experiments were done with adult mice of both genders at an age of 36 - 48 weeks. The illumination is achieved using a 3 x 3 micro-LED array, fabricated as described in16,17 except that the silicon-based housing and the short optical glass fibers are not implemented. Its first usage in a cardiac application is found in18. A linear micro-LED array based on a similar fabrication technology has been applied as a penetrating probe for heart pacing19. The micro-LEDs are arranged in a 3 x 3 array at a pitch of 550 &#181;m, providing both a high spatial resolution and a high radiant power on a very small area. The authors demonstrate in this work a versatile local multi-site photostimulation that may open the path for developing novel anti-arrhythmic therapy methods.
The following experimental protocol involves a retrograde Langendorff perfusion ex vivo, for which the cannulated aorta functions as perfusion inlet. Due to the applied perfusion pressure and the cardiac contraction the perfusate is flowing through the coronary arteries, which branch off the aorta. In the presented work, the heart is perfused using a constant pressure setup achieved by elevating the perfusate reservoirs to 1 m height, equivalent to 73.2 mmHg, which yields to a flow rate of 2.633 &#177; 0.583 mL/min. Two kinds of Tyrode&#39;s solution are used as perfusate during the experiment. Regular Tyrode&#39;s solution supports a stable sinus rhythm, whereas Low-K+ Tyrode&#39;s solution is mixed with Pinacidil to enable the induction of arrhythmia in murine hearts. The usage of a hexagonal water bath permits the observation of the heart through six different planar windows, allowing the coupling of several optical components with less distortion by refraction.

<table><tbody><tr><td>Chemical Components</td><td></td><td></td><td></td></tr><tr><td>Blebbistatin</td><td>TargetMol</td><td>T6038</td><td>10 mM stock solution</td></tr><tr><td>BSA/Albumin</td><td>Sigma-Aldrich</td><td>A4919</td><td></td></tr><tr><td>Calcium Chloride</td><td>Sigma-Aldrich</td><td>C1016</td><td>CaCl2</td></tr><tr><td>Carbogen</td><td>Westfalen</td><td></td><td>50 l bottle</td></tr><tr><td>DI-4-ANBDQPQ</td><td>AAT Bioquest</td><td>21499</td><td>Dye for Optical Mapping</td></tr><tr><td>Glucose</td><td>Sigma-Aldrich</td><td>D9434</td><td>C6H12O6</td></tr><tr><td>Heparin</td><td>LEO Pharma</td><td></td><td>Heparin-Natrium Leo 25.000 I.E./5 ml, available only on prescription</td></tr><tr><td>Hydrochlorid Acid</td><td>Merck</td><td>1.09057.1000</td><td>HCl, 1 M stock solution</td></tr><tr><td>Isoflurane</td><td>CP Pharma</td><td></td><td>1 ml/ml, available only on prescription</td></tr><tr><td>Magnesium Chloride</td><td>Merck</td><td>8.14733.0500</td><td>MgCl2</td></tr><tr><td>Monopotassium Phosphate</td><td>Sigma-Aldrich</td><td>30407</td><td>KH2PO4</td></tr><tr><td>Pinacidil monohydrate</td><td>Sigma-Aldrich</td><td>P154-500mg</td><td>10 mM stock solution</td></tr><tr><td>Potassium Chloride</td><td>Sigma-Aldrich</td><td>P5405</td><td>KCl</td></tr><tr><td>Sodium Bicarbonate</td><td>Sigma-Aldrich</td><td>S5761</td><td>NaHCO3</td></tr><tr><td>Sodium Chloride</td><td>Sigma-Aldrich</td><td>S5886</td><td>NaCl</td></tr><tr><td>Sodium Hydroxide</td><td>Merck</td><td>1.09137.1000</td><td>NaOH, 1 M stock solution</td></tr><tr><td>Electrical Setup</td><td></td><td></td><td></td></tr><tr><td>Biopac MP150</td><td>Biopac Systems</td><td>MP150WSW</td><td>data acquisition and analysis system</td></tr><tr><td>Custom-built ECG, alternative ECG100C</td><td>Biopac Systems</td><td>ECG100C</td><td>Electrocardiogram Amplifier</td></tr><tr><td>Custom-built water bath heater using heating cable</td><td>RMS Heating System</td><td>HK-5,0-12</td><td>Heating cable 120W</td></tr><tr><td>Hexagonal water bath</td><td></td><td></td><td></td></tr><tr><td>LED Driver Power supply</td><td>Thorlabs</td><td>KPS101</td><td>15 V, 2.4 A Power Supply Unit with 3.5 mm Jack Connector for One K- or T-Cube.</td></tr><tr><td>LEDD1B LED Driver</td><td>Thorlabs</td><td>LEDD1B</td><td>T-Cube LED Driver, 1200 mA Max Drive Current</td></tr><tr><td>MAP, ECG Electrode</td><td>Hugo Sachs Elektronik</td><td>BS4&amp;nbsp;73-0200</td><td>Mini-ECG Electrode for isoalted hearts</td></tr><tr><td>micro-LED Driver e.g. AFG</td><td>Agilent Instruments</td><td>A-2230</td><td>Arbitrary function generator (AFG)</td></tr><tr><td>Signal Generator</td><td>Agilent Instruments</td><td>A-2230</td><td>AFG</td></tr><tr><td>micro-LED Array Components</td><td></td><td></td><td></td></tr><tr><td>Epoxid glue</td><td>Epoxy Technology</td><td>EPO-TEK 353ND</td><td>Two component epoxy</td></tr><tr><td>Fluoropolymer</td><td>&amp;nbsp;Asahi Glass Co. Ltd.</td><td>Cytop 809M</td><td>Fluoropolymer with high transparency</td></tr><tr><td>Image reversal photoresist</td><td>Merck KGaA</td><td>AZ 5214E</td><td>Image Reversal Resist for High Resolution</td></tr><tr><td>LED chip</td><td>&amp;nbsp;Cree Inc.</td><td>C460TR2227-S2100</td><td>Blue micro-LED</td></tr><tr><td>Photoresist</td><td>Merck KGaA</td><td>AZ 9260</td><td>Thick Positive Photoresists</td></tr><tr><td>Polyimide</td><td>UBE Industries Ltd.</td><td>U-Varnish S</td><td>Polyimide Solution</td></tr><tr><td>Silicone</td><td>NuSil Technology LLC</td><td>MED-6215</td><td>Low viscosity silicone elastomer</td></tr><tr><td>Solvent free adhesive</td><td>John P. Kummer GmbH</td><td>Epo-Tek 301-2</td><td>Epoxy resin with low viscosity</td></tr><tr><td>Optical Mapping</td><td></td><td></td><td></td></tr><tr><td>Blue Filter</td><td>Chroma Technology Corporation</td><td>ET470/40x</td><td>Blue excitation filter</td></tr><tr><td>Camera</td><td>Photometrics</td><td>Cascade 128+</td><td>High performance EMCCD Camera</td></tr><tr><td>Camera Objective</td><td>Navitar</td><td>DO-5095</td><td>Navitar high speed fixed focal length lenses work with CCD and CMOS cameras</td></tr><tr><td>Dichroic Mirror</td><td>Semrock</td><td>FF685-Di02-25x36</td><td>685 nm edge BrightLine&amp;reg; single-edge standard epi-fluorescence dichroic beamsplitter</td></tr><tr><td>Emmision Filter</td><td>Semrock</td><td>FF01-775/140-25</td><td>775/140 nm BrightLine&amp;reg; single-band bandpass filter</td></tr><tr><td>Heatsink</td><td>Advanced Thermal Solutions</td><td>ATSEU-077A-C3-R0</td><td>Heat Sinks - LED STAR LED Heatsink, 45mm dia., 68mm, Black/Silver, Unthreaded Baseplate Hardware</td></tr><tr><td>LED 1 and LED 2</td><td>LED Engin Osram</td><td>LZ4-00B208</td><td>High Power LEDs - Single Colour Blue, 460 nm 130 lm, 700mA</td></tr><tr><td>LED 3</td><td>Thorlabs</td><td>M625L3</td><td>625 nm, 700 mW (Min) Mounted LED, 1000 mA</td></tr><tr><td>Lenses</td><td>LED Engin Osram</td><td>LLNF-2T06-H</td><td>LED Lighting Lenses Assemblies LZ4 LENS NARROW FLOOD BEAM</td></tr><tr><td>Photodiode for power meter</td><td>Thorlabs</td><td>S120VC</td><td>Standard Photodiode Power Sensor</td></tr><tr><td>Power Meter</td><td>Thorlabs</td><td>PM100D</td><td>Compact Power and Energy Meter</td></tr><tr><td>Red Filter</td><td>Semrock</td><td>FF02-628/40-25</td><td>BrightLine&amp;reg; single-band bandpass filter</td></tr></tbody></table>

advanced cardiac rhythm management by applying optogenetic multi-site photostimulation in murine hearts

Advanced Cardiac Rhythm Management by Applying Optogenetic Multi-Site Photostimulation in Murine Hearts

Ventricular tachyarrhythmias are a major cause of mortality and morbidity worldwide. Electrical defibrillation using high-energy electric shocks is currently the only treatment for life-threatening ventricular fibrillation. However, defibrillation may have side-effects, including intolerable pain, tissue damage, and worsening of prognosis, indicating a significant medical need for the development of more gentle cardiac rhythm management strategies. Besides energy-reducing electrical approaches, cardiac optogenetics was introduced as a powerful tool to influence cardiac activity using light-sensitive membrane ion channels and light pulses. In the present study, a robust and valid method for successful photostimulation of Langendorff perfused intact murine hearts will be described based on multi-site pacing applying a 3 x 3 array of micro light-emitting diodes (micro-LED). Simultaneous optical mapping of epicardial membrane voltage waves allows the investigation of the effects of region-specific stimulation and evaluates the newly induced cardiac activity directly on-site. The obtained results show that the efficacy of defibrillation is strongly dependent on the parameters chosen for photostimulation during a cardiac arrhythmia. It will be demonstrated that the illuminated area of the heart plays a crucial role for termination success as well as how the targeted control of cardiac activity during illumination for modifying arrhythmia patterns can be achieved. In summary, this technique provides a possibility to optimize the on-site mechanism manipulation on the way to real-time feedback control of cardiac rhythm and, regarding the region specificity, new approaches in reducing the potential harm to the cardiac system compared to the usage of non-specific electrical shock applications.

The protocol allows the induction of ventricular arrhythmias in intact murine hearts using photostimulation pulses generated by LED 1 and LED 2 (Figure 1) with a frequency find between 25 Hz and 35 Hz and a pulse duration Wind between 2 ms and 10 ms. Please notice that the aim of such rapid light pulses is not to capture the cardiac rhythm but rather to unbalance the cardiac activity so that erratic electrical waves can be generated, which then facilitate an arrhythmia....

This work reports a method for controlling the cardiac rhythm of intact murine hearts of transgenic channelrhodopsin-2 (ChR2) mice using local photostimulation with a micro-LED array and simultaneous optical mapping of epicardial membrane potential.

Ventricular tachyarrhythmias are a major cause of mortality and morbidity worldwide. Electrical defibrillation using high-energy electric shocks is ...

advanced-cardiac-rhythm-management-applying-optogenetic-multi-site

Research

JoVE Journal

Medicine

This article describes a high temporal and spatial resolution technique to optically image action potential movement on the surface of Langendorff-perfused rat hearts using a potentiometric dye (di-8-ANEPPS).

Optical Mapping of Langendorff-perfused Rat Hearts

This article describes the basic procedures for conducting optical mapping experiments in the Langendorff-perfused rabbit heart using the panoramic imaging system, and the dual (voltage and calcium) imaging modality.

Multiparametric Optical Mapping of the Langendorff-perfused Rabbit Heart

This paper details the dissection procedure, instrumental setup, and experimental conditions during optical mapping of transmembrane potential (Vm) and intracellular calcium transient (CaT) in intact isolated Langendorff perfused mouse hearts.

Optical Mapping of Action Potentials and Calcium Transients in the Mouse Heart

This protocol describes the electrophysiological evaluation of murine atria utilizing an optical mapping system with a high temporal and spatial resolution, including dual recordings of the membrane voltage and Ca2+ transient under programmed stimulation through a specialized electrode catheter.

Electrophysiological Assessment of Murine Atria with High-Resolution Optical Mapping

Here, we present a protocol for the modulation of the intracardiac autonomic nervous system and the assessment of its influence on basic electrophysiology, arrhythmogenesis, and cAMP dynamics using an ex vivo Langendorff setup.

Impact of Intracardiac Neurons on Cardiac Electrophysiology and Arrhythmogenesis in an Ex Vivo Langendorff System

The objective of this study was to establish a method for investigating cardiac dynamics using a translational animal model. The described experimental approach incorporates dual-emission optocardiography in conjunction with an electrophysiological study to assess electrical activity in an isolated, intact porcine heart model.

Optocardiography and Electrophysiology Studies of Ex Vivo Langendorff-perfused Hearts

We present a protocol for evaluating the electromechanical effects of GtACR1 activation in rabbit cardiomyocytes. We provide detailed information on cell isolation, culturing and adenoviral transduction, and on functional experiments with the patch-clamp and carbon-fiber techniques.

Electromechanical Assessment of Optogenetically Modulated Cardiomyocyte Activity

The present protocol describes the generation of Drosophila melanogaster expressing eNpHR2.0 or ReaChR opsins in the heart for OCT imaging and optogenetic heart pacing. Detailed instructions for Drosophila OCT imaging and heart beating modulation, including the simulation of restorable heart arrest, bradycardia, and tachycardia in live animals at different developmental stages, are reported.

Developing Drosophila melanogaster Models for Imaging and Optogenetic Control of Cardiac Function

This protocol introduces dual-dye optical mapping of mouse hearts obtained from wild-type and knock-in animals affected by catecholaminergic polymorphic ventricular tachycardia, including electrophysiological measurements of transmembrane voltage and intracellular Ca2+ transients with high temporal and spatial resolution.

Dual-Dye Optical Mapping of Hearts from RyR2R2474S Knock-In Mice of Catecholaminergic Polymorphic Ventricular Tachycardia

This protocol illustrates a method for optogenetic stimulation of intrinsic cardiac neurons in transgenic mouse hearts. The approach described is used for investigating the kinetics of sudden cardiac neuron activation in ex vivo perfused hearts and the interactions between cholinergic and catecholaminergic activity.

Optogenetic Activation of Intrinsic Cardiac Autonomic Neurons in Excised Perfused Mouse Hearts

Using light to control cardiac cells and tissue enables non-contact stimulation, thereby preserving the natural state and function of the cells, making it a valuable approach for both basic research and therapeutic applications.

A Cardiac Microphysiological System for Studying Ca2+ Propagation via Non-genetic Optical Stimulation

Learn optogenetic photostimulation of Langendorff-perfused murine hearts using a micro-LED array for precise cardiac rhythm control. This method enables targeted arrhythmia termination with real-time optical mapping.

Advanced Cardiac Rhythm Management by Applying Optogenetic Multi-Site Photostimulation in Murine Hearts

Explore More Videos