This research was supported by the National Natural Science Foundation of China (31871085), the Natural Science Foundation of Shanghai (21ZR1407300), the Shanghai Municipal Science and Technology Major Project (2018SHZDZX01), ZJ Lab, and Shanghai Center for Brain Science and Brain-Inspired Technology.

This protocol received approval from the Animal Care and Use Committee at Fudan University and was conducted following the guidelines and regulations designed by the National Institutes of Health Guide for Care and Use of Laboratory Animals. All possible measures were implemented to minimize the number of animals utilized in this study. The time required to perform each step is included in the respective steps.
1. Preparation of electrodes (Figure 1...

Flexible Multi-Region Neural Recording Device for Epilepsy Research

<ol>
	<li>Devinsky, O., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Epilepsy.">Epilepsy.</a> Nat Rev Dis Primers. 4, 18024 (2018).</li><li>Piper, R. J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Towards+network-guided+neuromodulation+for+epilepsy.">Towards network-guided neuromodulation for epilepsy.</a> Brain. 145 (10), 3347-3362 (2022).</li><li>Bertram, E. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neuronal+circuits+in+epilepsy:+Do+they+matter.">Neuronal circuits in epilepsy: Do they matter.</a> Exp Neurol. 244, 67-74 (2013).</li><li>Goodman, A. M., Szaflarski, J. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Recent+advances+in+neuroimaging+of+epilepsy.">Recent advances in neuroimaging of epilepsy.</a> Neurotherapeutics. 18 (2), 811-826 (2021).</li><li>Caciagli, L., Bernhardt, B. C., Hong, S. J., Bernasconi, A., Bernasconi, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Functional+network+alterations+and+their+structural+substrate+in+drug-resistant+epilepsy.">Functional network alterations and their structural substrate in drug-resistant epilepsy.</a> Front Neurosci. 8, 411 (2014).</li><li>Dai, Y., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=In+vivo+microelectrode+arrays+for+detecting+multi-region+epileptic+activities+in+the+hippocampus+in+the+latent+period+of+rat+model+of+temporal+lobe+epilepsy.">In vivo microelectrode arrays for detecting multi-region epileptic activities in the hippocampus in the latent period of rat model of temporal lobe epilepsy.</a> Micromachines (Basel). 12 (6), 659 (2021).</li><li>Staba, R. J., Stead, M., Worrell, G. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Electrophysiological+biomarkers+of+epilepsy.">Electrophysiological biomarkers of epilepsy.</a> Neurotherapeutics. 11 (2), 334-346 (2014).</li><li>Grienberger, C., Giovannucci, A., Zeiger, W., Portera-Cailliau, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Two-photon+calcium+imaging+of+neuronal+activity.">Two-photon calcium imaging of neuronal activity.</a> Nat Rev Methods Primers. 2 (1), 67 (2022).</li><li>Akerboom, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Optimization+of+a+gcamp+calcium+indicator+for+neural+activity+imaging.">Optimization of a gcamp calcium indicator for neural activity imaging.</a> J Neurosci. 32 (40), 13819-13840 (2012).</li><li>Zhang, Y., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fast+and+sensitive+gcamp+calcium+indicators+for+imaging+neural+populations.">Fast and sensitive gcamp calcium indicators for imaging neural populations.</a> Nature. 615 (7954), 884-891 (2023).</li><li>Wu, Z., Lin, D., Li, Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pushing+the+frontiers:+Tools+for+monitoring+neurotransmitters+and+neuromodulators.">Pushing the frontiers: Tools for monitoring neurotransmitters and neuromodulators.</a> Nat Rev Neurosci. 23 (5), 257-274 (2022).</li><li>Vierock, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bipoles+is+an+optogenetic+tool+developed+for+bidirectional+dual-color+control+of+neurons.">Bipoles is an optogenetic tool developed for bidirectional dual-color control of neurons.</a> Nat Commun. 12 (1), 4527 (2021).</li><li>Luo, W., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Acquiring+new+memories+in+neocortex+of+hippocampal-lesioned+mice.">Acquiring new memories in neocortex of hippocampal-lesioned mice.</a> Nat Commun. 13 (1), 1601 (2022).</li><li>Armstrong, C., Krook-Magnuson, E., Oijala, M., Soltesz, I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Closed-loop+optogenetic+intervention+in+mice.">Closed-loop optogenetic intervention in mice.</a> Nat Protoc. 8 (8), 1475-1493 (2013).</li><li>Kraeuter, A. K., Guest, P. C., Sarnyai, Z. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+open+field+test+for+measuring+locomotor+activity+and+anxiety-like+behavior.">The open field test for measuring locomotor activity and anxiety-like behavior.</a> Methods Mol Biol. 1916, 99-103 (2019).</li><li>Klapoetke, N. C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Independent+optical+excitation+of+distinct+neural+populations.">Independent optical excitation of distinct neural populations.</a> Nat Methods. 11 (3), 338-346 (2014).</li><li>Vismer, M. S., Forcelli, P. A., Skopin, M. D., Gale, K., Koubeissi, M. Z. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+piriform,+perirhinal,+and+entorhinal+cortex+in+seizure+generation.">The piriform, perirhinal, and entorhinal cortex in seizure generation.</a> Front Neural Circuits. 9, 27 (2015).</li><li>Chen, T. W., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ultrasensitive+fluorescent+proteins+for+imaging+neuronal+activity.">Ultrasensitive fluorescent proteins for imaging neuronal activity.</a> Nature. 499 (7458), 295-300 (2013).</li><li>Birling, M. C., Dierich, A., Jacquot, S., Herault, Y., Pavlovic, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Highly-efficient,+fluorescent,+locus+directed+cre+and+flpo+deleter+mice+on+a+pure+c57bl/6n+genetic+background.">Highly-efficient, fluorescent, locus directed cre and flpo deleter mice on a pure c57bl/6n genetic background.</a> Genesis. 50 (6), 482-489 (2012).</li><li>Huang, T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Identifying+the+pathways+required+for+coping+behaviours+associated+with+sustained+pain.">Identifying the pathways required for coping behaviours associated with sustained pain.</a> Nature. 565 (7737), 86-90 (2019).</li><li>Likhtik, E., Stujenske, J. M., Topiwala, M. A., Harris, A. Z., Gordon, J. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Prefrontal+entrainment+of+amygdala+activity+signals+safety+in+learned+fear+and+innate+anxiety.">Prefrontal entrainment of amygdala activity signals safety in learned fear and innate anxiety.</a> Nat Neurosci. 17 (1), 106-113 (2014).</li><li>Tavares, L. C. S., Tort, A. B. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Hippocampal-prefrontal+interactions+during+spatial+decision-making.">Hippocampal-prefrontal interactions during spatial decision-making.</a> Hippocampus. 32 (1), 38-54 (2022).</li><li>Lu, Y., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Optogenetic+dissection+of+ictal+propagation+in+the+hippocampal-entorhinal+cortex+structures.">Optogenetic dissection of ictal propagation in the hippocampal-entorhinal cortex structures.</a> Nat Commun. 7, 10962 (2016).</li><li>Liu, Q., Wu, Y., Wang, H., Jia, F., Xu, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Viral+tools+for+neural+circuit+tracing.">Viral tools for neural circuit tracing.</a> Neurosci Bull. 38 (12), 1508-1518 (2022).</li><li>Christenson Wick, Z., Krook-Magnuson, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Specificity,+versatility,+and+continual+development:+The+power+of+optogenetics+for+epilepsy+research.">Specificity, versatility, and continual development: The power of optogenetics for epilepsy research.</a> Front Cell Neurosci. 12, 151 (2018).</li><li>Paschen, E., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Hippocampal+low-frequency+stimulation+prevents+seizure+generation+in+a+mouse+model+of+mesial+temporal+lobe+epilepsy.">Hippocampal low-frequency stimulation prevents seizure generation in a mouse model of mesial temporal lobe epilepsy.</a> Elife. 9, e54518 (2020).</li><li>Gummadavelli, A., Zaveri, H. P., Spencer, D. D., Gerrard, J. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Expanding+brain-computer+interfaces+for+controlling+epilepsy+networks:+Novel+thalamic+responsive+neurostimulation+in+refractory+epilepsy.">Expanding brain-computer interfaces for controlling epilepsy networks: Novel thalamic responsive neurostimulation in refractory epilepsy.</a> Front Neurosci. 12, 474 (2018).</li><li>Vandekerckhove, B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Technological+challenges+in+the+development+of+optogenetic+closed-loop+therapy+approaches+in+epilepsy+and+related+network+disorders+of+the+brain.">Technological challenges in the development of optogenetic closed-loop therapy approaches in epilepsy and related network disorders of the brain.</a> Micromachines (Basel). 12 (1), 38 (2020).</li><li>Li, Z. J., Zhang, L. B., Chen, Y. X., Hu, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Advancements+and+challenges+in+neuromodulation+technology:+Interdisciplinary+opportunities+and+collaborative+endeavors.">Advancements and challenges in neuromodulation technology: Interdisciplinary opportunities and collaborative endeavors.</a> Sci Bull (Beijing). 68 (18), 1978-1982 (2023).</li><li>Zhang, Q., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+prototype+closed-loop+brain-machine+interface+for+the+study+and+treatment+of+pain.">A prototype closed-loop brain-machine interface for the study and treatment of pain.</a> Nat Biomed Eng. 7 (4), 533-545 (2023).</li><li>Varela, C., Wilson, M. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mpfc+spindle+cycles+organize+sparse+thalamic+activation+and+recently+active+ca1+cells+during+non-rem+sleep.">Mpfc spindle cycles organize sparse thalamic activation and recently active ca1 cells during non-rem sleep.</a> Elife. 9, e48881 (2020).</li><li>Adaikkan, C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Gamma+entrainment+binds+higher-order+brain+regions+and+offers+neuroprotection.">Gamma entrainment binds higher-order brain regions and offers neuroprotection.</a> Neuron. 102 (5), 929-943.e8 (2019).</li><li>Muthmann, J. O., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Spike+detection+for+large+neural+populations+using+high+density+multielectrode+arrays.">Spike detection for large neural populations using high density multielectrode arrays.</a> Front Neuroinform. 9, 28 (2015).</li></ol>

The authors declare no competing financial interests to disclose.

Here, we employed a self-made optrode device for in vivo neural signal recording across multiple regions. The feasibility of this system for simultaneous optogenetic stimulation, calcium signal recording, and electrophysiological recording has been validated. The electrode preparation method described herein is efficient and cost-effective. According to the experimental design, we could record signals from relevant brain regions. The strategic arrangement of optrodes allows for the potential extraction of abunda...

Epilepsy is a common neurological condition characterized by recurrent seizures, which manifest as convulsions, sensory disturbances, and loss of consciousness1. The pathophysiological mechanisms underlying epilepsy are complex and involve multiple interconnected brain regions2,3. Recent advances in neuroimaging have shed light on the large-scale networks involved in epilepsy4,5. However, understanding of the intricate circuitry and network mechanisms underlying the generation and propagation of epilepsy remains limited, partly due to the insufficient application of multi-region neural recording techniques6. Therefore, developing a flexible, integrated method capable of simultaneously monitoring neural activity across disparate brain regions is imperative.
Electrophysiological recordings are conducted to capture seizures and determine the presence of epilepsy7. Except for recording electrophysiological activity, there is a growing emphasis on the precise calcium activity of specific&#160;neural populations in epilepsy studies8,9. Advancements in calcium indicator synthesis and various probe designs have prompted researchers to adopt fiber photometry for capturing changes in neuronal activity and neural substances within the brain10,11. The two independent methods of detecting neuronal activity, namely electrophysiology and fiber photometry recording, complement each other, facilitating a more comprehensive understanding of dynamic neuronal processes.
Additionally, synchronous recording and modulating neural activity are pivotal for gaining insights into the brain functioning at both network and cellular levels. This approach enables researchers to observe and manipulate the brain&#39;s complex processes in real-time. Optogenetics has emerged as an indispensable tool for probing neural signaling, owing to its distinctive ability for selective stimulation or inhibition12. Despite widespread applications of multi-site electrophysiological recording in neuroscience13, the integration of multi-region electrophysiological recording with fiber photometry and optogenetic manipulation remains limited. More importantly, current methods for fabricating and implanting multi-region electrodes lack flexibility14. These limitations hinder our capacity to dissect specific circuit functions and interactions across multiple regions. Here, we present a cost-effective and convenient multi-region in vivo recording approach that sheds light on neuronal processes across regions in kindling-induced seizures and other neuropsychiatric disorders.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>8-32 adapter</td>
			<td>Plexon</td>
			<td>Custom ordered</td>
			<td>Connect the female connector and headstage</td>
		</tr>
		<tr>
			<td>AAV-CaMKII&#945;-ChrimsonR-mcherry</td>
			<td>Taitool Bioscience</td>
			<td>S0371-9</td>
			<td>4 x 1012 VG/mL&#160;</td>
		</tr>
		<tr>
			<td>AAV-hsyn-Gcamp6m</td>
			<td>Taitool Bioscience</td>
			<td>S0471-9</td>
			<td>4 x 1012 VG/mL&#160;</td>
		</tr>
		<tr>
			<td>DAPI</td>
			<td>Sigma</td>
			<td>236276</td>
			<td>Titered 1:500</td>
		</tr>
		<tr>
			<td>Dental Cement</td>
			<td>New Century Dental</td>
			<td>430205</td>
			<td></td>
		</tr>
		<tr>
			<td>Electrophysiological recordings system</td>
			<td>Plexon</td>
			<td>Omniplex</td>
			<td></td>
		</tr>
		<tr>
			<td>Enameled wire</td>
			<td>N/A</td>
			<td>Custom ordered</td>
			<td>Diameter = 0.2 mm</td>
		</tr>
		<tr>
			<td>Female connector</td>
			<td>N/A</td>
			<td>Custom ordered</td>
			<td>1.25 mm pitch</td>
		</tr>
		<tr>
			<td>Glue</td>
			<td>Loctite</td>
			<td>45282</td>
			<td></td>
		</tr>
		<tr>
			<td>Laser</td>
			<td>Changchun New Industries</td>
			<td>BH81563</td>
			<td>635 nm&#160;</td>
		</tr>
		<tr>
			<td>MATLAB</td>
			<td>MathWorks</td>
			<td>R2021b</td>
			<td></td>
		</tr>
		<tr>
			<td>Microdrill</td>
			<td>RWD</td>
			<td>78001</td>
			<td></td>
		</tr>
		<tr>
			<td>Multichannel fiber photometry</td>
			<td>ThinkerTech</td>
			<td>FPS-SS-MC-LED</td>
			<td></td>
		</tr>
		<tr>
			<td>Optical fiber</td>
			<td>Xi&#39;an Bogao</td>
			<td>L-200UM</td>
			<td>Select the appropriate fiber length based on the depth of the targeted brain regions.</td>
		</tr>
		<tr>
			<td>PFA-Coated Tungsten wire</td>
			<td>A-M System</td>
			<td>795500</td>
			<td>Bare 0.002&#34;; Coated 0.0040&#34;</td>
		</tr>
		<tr>
			<td>Power meter</td>
			<td>Thorlabs</td>
			<td>PM100D</td>
			<td></td>
		</tr>
		<tr>
			<td>Stereotaxic Fxrame</td>
			<td>RWD</td>
			<td>68807</td>
			<td></td>
		</tr>
		<tr>
			<td>Tissue adhesive</td>
			<td>3M</td>
			<td>1469SB</td>
			<td></td>
		</tr>
	</tbody>
</table>

an integrated method for crafting flexible and convenient electrophysiological optrodes for multi-region in vivo recording

Epilepsy is a neurological disorder characterized by synchronized abnormal discharges involving multiple brain regions. Focal lesions facilitate the propagation of epileptic signals through associated neural circuits. Therefore, in vivo recording of local field potential (LFP) from the critical brain regions is&#160;essential for deciphering the circuits involved in seizure propagation. However, current methods for electrode fabrication and implantation lack flexibility. Here, we present a handy device designed for electrophysiological recordings (LFPs and electroencephalography [EEG]) across multiple regions. Additionally, we seamlessly integrated optogenetic manipulation and calcium signaling recording with LFP recording. Robust after-discharges were observed in several separate regions during epileptic seizures, accompanied by increasing&#160;calcium signaling. The approach used in this study offers a convenient and flexible strategy for synchronous neural recordings across diverse regions of the brain. It holds the potential for advancing research on neurological disorders by providing insights into the neural profiles of multiple regions involved in these disorders.

Author Spotlight: Unraveling Neural Communication and Circuit Interactions in Health and Disease

An Integrated Method for Crafting Flexible and Convenient Electrophysiological Optrodes for Multi-Region In Vivo Recording

We investigate brain networks and neural circuits using multimodal approaches which encompass techniques such as multichannel electrophysiological recording, fiber photometry, neuro modulation, and functional magnetic resonance imagery. We are trying to answer how different brain regions communicate and interact in vivo both in healthy and diseased states. We use methods for natural replication and implantation to connect the molecular cellular cycles and network layers.This comprehensive approach allowed us to study neurological disorders more effectively and a deep understanding of their underlying mechanisms. Our results pave the way for new scientific question about how flexible multiregion brain recordings can be, how different brain circuits interact in epilepsy, and how this could apply to other neurological disorders. It offers fringe insights into your neural activities across various brain regions.

We combined optogenetics with multiregional electrophysiological recording and calcium imaging to observe neuronal activity across various brain regions during optogenetic seizures. For this purpose, an adeno-associated virus (AAV) expressing ChrimsonR under the control of the CaMKII&#945; promoter (AAV-CaMKII&#945;-ChrimsonR-mcherry)16 was injected into a classical epileptogenic site, the piriform cortex (ROI 1)17, in rodents. Additionally, AAV-hsyn-Gcamp6m<sup class="xref...

We have developed a simplified and cost-effective approach for electrode fabrication and conducted recordings of signals across multiple regions in freely moving mice. Utilizing optogenetics, alongside multi-region electrophysiology and calcium signal recording, enabled the revelation of neuronal activities across regions in the seizure kindling model.

Epilepsy is a neurological disorder characterized by synchronized abnormal discharges involving multiple brain regions. Focal lesions facilitate the ...

an-integrated-method-for-crafting-flexible-convenient

Research

JoVE Journal

Neuroscience

Designing Flexible Optrode for Electrophysiological Recordings Across Multiple Regions in Freely Moving Mice

To begin, cut an appropriate length of tungsten wire and burn off the insulation layer at one end to expose the tungsten wire. Then, fold the other end of the wire into an L shape. Carefully place the L end of the tungsten wire onto the optical fiber.Dip a drop of adhesive and apply it at the interface of the optical fiber and the tungsten wire. At the corner of the L shape, use adhesive to closely bond the base of the optical fiber and the tungsten wire together. Then, use soldering to establish a connection between the tungsten wire and the pin of the female connector.According to the experimental design, solder the required number of optrodes. Next, cut the enameled wire to the appropriate length and burn off the insulation on both ends. Wrap one end of the enameled wire around the base of the screw and solder it.After that, connect the other end of the enameled wire to the pins of the female connector via soldering iron. Then, apply hot melt adhesive to the female connector's pin array to encapsulate the connector and ensure the solder joints are fully enveloped. Shape the adhesive using a plastic board during its solidification to minimize space occupancy during implantation.Next, use a light intensity meter and digital multimeter to measure luminousness and conductivity. Once these tests are passed, the setup is ready for use. Before use, immerse the prepared electrodes in 75%alcohol for 15 minutes.Then, transfer them to sterile saline.

Procedure for Electrode Implantation Surgery in Mice

After securing the anesthetized animal in the stereotaxic apparatus, trim away the scalp along the edge of the skull. Then wipe the skull surface with 75%alcohol to expose the positions of the bregma and lambda. Using a micro drill, level the mouse and drill holes above the regions of interest.Then inject the relevant viruses with a syringe pump. Now hold the optrodes with a holder and move their tip to the bregma point, setting this position as zero. Using the bregma point as the reference coordinate, move the optrodes to the target regions.Position the optrodes 0.3 millimeters above the virus injection site. After inserting the optic fibers into the corresponding brain areas, apply a small amount of tissue adhesive around them to cover the exposed tissue. Sequentially apply additional adhesive and a little dental cement.After the dental cement solidifies, release the holder carefully. Once all erodes are implanted, drill the holes behind the lambda point for ground and above the neocortex for EEG using a micro drill. Insert skull screws into the hole above the cortex and behind the lambda point.Then organize the tungsten and enameled wires so they are positioned above the implantation area and not exposed outside the skull. Secure the position of the female connector using a holder. Now fill the implantation site with dental cement to ensure a stable connection between the electrode device and the skull.Encapsulate the internal wires, leaving the female connector's insertion holes exposed for connection to the head stage. Once the dental cement hardens, apply iodine to the wound and place the animal on a warming pad.

Recording In Vivo Synchronous Electrophysiological and Calcium Activity in Mice

To begin, implant the recording electrodes and inject the relevant viruses in the mouse skull. After anesthetizing the mice, connect the optical fibers sequentially in the order of the recording site before inserting the head stage. Then, set the 635 nanometer light pulse with a 10%duty cycle at 20 hertz, and set up the light stimulation for 10 seconds during recording.Set the intensity at the optical fiber tips at three milliwatts. Next, use a fiber photometry system to record Gcamp6m signals throughout the behavior testing. Simultaneously, record calcium signals, local field potentials, and electroencephalogram under optogenetic stimulation, along with corresponding behavioral performances.Finally, employ transistor-transistor logic signal synchronization tagging to ensure temporal consistency in recorded data. Brief optogenetic stimulation elicited robust after discharges, along with remarkable increases in calcium activities in the target brain regions. A 30 hertz response was observed in the opto-stimulating regio, induced by the activation of optogenetic proteins via blue light LED.

352 Views.  Fudan University.  We have developed a simplified and cost-effective approach for electrode fabrication and conducted recordings of signals across multiple regions in freely moving mice. Utilizing optogenetics, alongside multi-region electrophysiology and calcium signal recording, enabled the revelation of neuronal activities across regions in the seizure kindling model.

Flexible Multi-Region Neural Recording Device for Epilepsy Research

Summary