This work was supported by Kaken-hi (17H05939; 17H05551), the Nakatomi Foundation, and the Suzuken Memorial Foundation.

All of the procedures involving animal subjects were performed according to the NIH guidelines for the care and use of animals.
1. Preparation of the Integrative Micro-Drive Array
<ol>
	<li>Create a micro-drive array for cortical LFP recordings as described elsewhere1,2,3. Leave at least 6 metal holes open on an electrode interface board (EIB) for use as ECG/EMG/BR channels that are connected to bioflex wires as described in 1.2.</li>
	<li>Cut a bioflex wire into 6 pieces with lengths of 5.0 cm. Peel off the polytetrafluoroethylene (PTFE) coating of both ends of all the wire pieces with lengths of ~5.0 mm. Connect one end of each of the wire pieces to one of the open metallic holes (ECG/EMG/BR channels) on the EIB with a gold pin.</li>
	<li>Cut an enamel wire into two 5.0-cm pieces. Solder one end of each of these wires to the ground/reference (g/r) channels on the EIB (Figure 3, see also previous papers12,13).</li>
	<li>For the preparation of ECG electrodes, cut a bioflex wire into two 16-cm pieces. Peel off the PTFE coating of the ends of these wire pieces at lengths of ~5.0 mm on one end (short end) and ~15 mm on the other end (long end).</li>
	<li>Form a wire ring with a diameter of 2.0 mm by bending the long end of the wire, and fixing the shape of the ring by soldering.</li>
	<li>For the preparation of EMG electrodes, cut a bioflex wire into 2 pieces with lengths of 8 cm. Peel off the PTFE coating from both ends of these wire pieces with lengths of ~5.0 mm.</li>
	<li>For the preparation of BR electrodes, cut a bioflex wire into 2 pieces with lengths of 6.0 cm. Peel off the enamel coating of both ends of these wire pieces with lengths of ~5.0 mm. Solder one end of each of these wire pieces to the head of a stainless- steel screw (stem diameter: 1.0 mm, stem length: 4.0 mm).</li>
	<li>For the preparation of ground/reference (gr) electrodes, cut an enamel wire into 2 pieces with lengths of 6.0 cm. Peel off the enamel coating of both ends of these wire pieces with lengths of ~5.0 mm. Solder one end of each of these wire pieces to the head of a stainless- steel screw (stem diameter: 1.4 mm, stem length: 3.0 mm).</li>
	<li>Gas sterilize all electrodes and stainless screws&#160;and keep these in a clean space.</li>
</ol>
2. Implantation of the ECG/EMG Electrodes
NOTE: Perform all surgical steps with aseptic technique using sterilized gloves and autoclaved instruments. For all steps involving the creation of an incision, sterilize the skin with 70% ethanol before, and cover the incision with surgical drapes.
<ol>
	<li>Fix an anesthetized (1.0-3.0% isoflurane gas) rat on its back on a flat heat pad. Give buprenorphine as an analgesic. Place veterinary ointment on the rat&#39;s eyes to prevent dryness.&#160;Use betadine to clean the surface of the skin.&#160;</li>
	<li>Make an incision of ~2.0 cm in the medial chest area. Expose the intercostal muscles by separating the chest muscles. Suture the rings of the ECG electrodes to the intercostal muscles.</li>
	<li>Fix the stomach of the animal on the heat pad. Make an incision of ~1.0 cm in the dorsal neck area.</li>
	<li>Insert the ECG electrodes subcutaneously through the chest incision. Slide the ends to the dorsal neck area, and pull them out from the neck incision. Suture the chest incision.</li>
	<li>Insert one end of each of the EMG electrodes subcutaneously to a length of ~2.0 cm through the neck incision. Fix the EMG electrodes to the neck muscle by suturing.</li>
</ol>
3. Implantation of the Integrative Micro-Drive Array and the BR Electrodes
<ol>
	<li>Fix the rat on a stereotaxic device. Make an incision of ~3.0 cm on the head along the midline from the point between the eyes to the neck area. Expose the skull.</li>
	<li>Make two circular craniotomies with diameters of 0.7-1.0 mm above the olfactory bulb 11.0 mm anterior and 1 mm bilateral to bregma with a high-speed drill. Implant two BR electrodes in the skull until the tips of the screw stems are attached to the brain surface.</li>
	<li>Make two circular craniotomies with diameters of 0.7-1.0 mm above the frontal cortex 2.7 mm anterior and 2.7 mm bilateral to bregma. Implant two g/r electrodes in the skull until the tip of the screw stem is attached to the brain surface.</li>
	<li>Make six to eight holes with diameters of 1.0 mm in the area surrounding the large craniotomy. Implant anchor screws (stem diameter: 1.4 mm, stem length: 3.0 mm) in the skull.</li>
	<li>Make a large circular craniotomy with a diameter of ~2.0 mm above the hippocampus 3.8 mm posterior and 2.5 mm bilateral to bregma. Place the integrative micro-drive array such that the cannula tip of the drive array is located above the large craniotomy</li>
	<li>Fill the gap space between the cannula tip and the brain surface with ~100 &#181;L of two solutions, i.e., 0.5% (by mass) sodium alginate and 10% (by mass) calcium chloride. 
	NOTE: This process forms a transparent gel in ~5 min, after the two solutions are mixed on the skull.</li>
	<li>Cover the cannula, BR electrodes, g/r electrodes, and anchor screws with dental cement with a thickness of 0.5 cm. Be careful NOT to cover the open ends of BR and g/r electrodes with the cement at this step.</li>
	<li>Solder the open ends of the ECG, EMG, BR and g/r electrodes to the individual wire tips that were previously connected to the EIB (see the steps 1.2 and 1.3).</li>
	<li>Cover the bottom part of the integrative micro-drive array, and all electrode wires, with dental cement. Ensure that all the electrode wires are completely covered so that the rat cannot scratch them out after the implantation.&#160;</li>
	<li>After regaining sufficient consciousness to maintain sternal recumbency, return the animal to its transparent Plexiglas home cag, and keep it on its own with free access to water and food.&#160;After the surgery, treat the animal with antibiotics (gentamicin).</li>
	<li>After the surgery, monitor the animals with&#160;daily observation. Check that they walk properly, and that they do not squeak when the experimenter touches the micro-drive array.&#160; &#160;</li>
</ol>
4. In Vivo Recordings
NOTE: All signals are amplified, sampled at 2 kHz, and band-pass filtered (0.1 - 500 Hz) except for unit activities (sampled at 30 kHz and band-pass filtered (500 - 6 kHz)).
<ol>
	<li>Connect the EIB of the integrative micro-drive array to the headstage of a recording device.</li>
	<li>Advance the tetrodes by turning the screws for a few weeks after surgery. Once the tetrodes are adjacent to the target brain areas, settle the tetrodes into the areas over a period of several days for stable recordings.</li>
	<li>Monitor the electrical signals while the animal freely moves in a recording chamber.</li>
</ol>

<ol>
	<li>Kloosterman, F., 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=Micro-drive+Array+for+Chronic+in+vivo+Recording:+Drive+Fabrication.">Micro-drive Array for Chronic in vivo Recording: Drive Fabrication.</a> JoVE. (26), e1094 (2009).</li><li>Nguyen, D. P., 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=Micro-drive+Array+for+Chronic+in+vivo+Recording:+Tetrode+Assembly.">Micro-drive Array for Chronic in vivo Recording: Tetrode Assembly.</a> Journal of Visualized Experiments : JoVE. (26), e1098 (2009).</li><li>Jog, M. S., 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=Tetrode+technology:+advances+in+implantable+hardware,+neuroimaging,+and+data+analysis+techniques.">Tetrode technology: advances in implantable hardware, neuroimaging, and data analysis techniques.</a> J Neurosci Methods. 117 (2), 141-152 (2002).</li><li>Fenske, S., 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=Comprehensive+multilevel+in+vivo+and+in+vitro+analysis+of+heart+rate+fluctuations+in+mice+by+ECG+telemetry+and+electrophysiology.">Comprehensive multilevel in vivo and in vitro analysis of heart rate fluctuations in mice by ECG telemetry and electrophysiology.</a> Nat Protoc. 11 (1), 61-86 (2016).</li><li>Rossi, S., 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=The+effect+of+aging+on+the+specialized+conducting+system:+a+telemetry+ECG+study+in+rats+over+a+6+month+period.">The effect of aging on the specialized conducting system: a telemetry ECG study in rats over a 6 month period.</a> PLoS One. 9 (11), 112697 (2014).</li><li>Cesarovic, N., Jirkof, P., Rettich, A., Arras, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Implantation+of+radiotelemetry+transmitters+yielding+data+on+ECG,+heart+rate,+core+body+temperature+and+activity+in+free-moving+laboratory+mice.">Implantation of radiotelemetry transmitters yielding data on ECG, heart rate, core body temperature and activity in free-moving laboratory mice.</a> JoVE. 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A., Khazan, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Electromyographic+power+spectral+changes+associated+with+the+sleep-awake+cycle+and+with+diazepam+treatment+in+the+rat.">Electromyographic power spectral changes associated with the sleep-awake cycle and with diazepam treatment in the rat.</a> Pharmacol Biochem Be. 19 (4), 715-718 (1983).</li><li>Oishi, 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=Polygraphic+Recording+Procedure+for+Measuring+Sleep+in+Mice.">Polygraphic Recording Procedure for Measuring Sleep in Mice.</a> JoVE. (107), e53678 (2016).</li><li>Chaput, M. 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The authors have nothing to disclose.

For understanding how the brain modulates peripheral activity levels, and vice versa, large-scale recording methods to simultaneously capture electrical biosignals from multiple body areas are necessary. This study described a surgical procedure, and a recording system for monitoring cerebral local field potentials, heart rates, the magnitude of muscle construction and respiratory rates, which have been improved on a recording system that is used for extracellular recordings in brain tissue. This system collects electrical signals from both the brain and the peripheral organs into a single EIB on an integrative micro-drive array. The preparation of the integrative micro-drive array should be started at least several hours before surgery, as it takes some time. Here, the drive array includes tetrodes for recording brain local field potentials, but the other types of metal electrodes, such as platinum and tungsten electrodes, can be attached to the EIB if the ends of these electrodes are properly soldered. When following the surgical procedures in the protocol, experienced experimenters have been able to complete all the procedures within 2-3 h. 
A critical step within this protocol is the positioning of the electrodes on the tissue, especially for the ECG and EMG electrodes. Several training repetitions might be required to obtain stable recordings. To date, all recordings have been stable at least one month after surgery. Experimenters should note that, if the signal-to-noise ratios of the recorded data become low, this problem is frequently due to loose fixations of the ground/reference electrodes, or insufficient soldering of the ends of wires between the EIBs or the other ends of wires. Compared with a conventional electrophysiological recording using some independent devices, the advantages of this method are that (1) it is technically simple to conduct, if trained several times, (2) it does not require a communication system to synchronize multiple devices, (3) it reduces the overall experimental costs, as only one recording device is required, and (4) all recorded data files can be handled by the same processing method and program, which increases the efficiency of data analysis. Furthermore, the methodological concept is applicable to many combinations of multiple brain regions and peripheral organs, including the respiratory system, circulatory system, and autonomic nervous system. It is also applicable to any commercially available electrical recording device. Simultaneous monitoring of systemic physiological activity patterns using this method will be helpful in elucidating neuronal activity patterns in various physiological states against emotional challenges, external sensory modulation, and pathological diseases, leading to an increased understanding of the biological mechanisms underlying the brain-body association.

The central nervous system controls body states in response to various environmental changes, and this control is typically represented as changes in heart rate, breathing rate, and muscle contractions. However, few studies have tested how such peripheral physiological factors are associated with cortical activity. To address this issue, a large-scale recording method for monitoring electrical biosignals from both central and peripheral tissues is necessary. In the cerebral cortex, local field potential (LFP) signals are extracellularly recorded by electrodes that are inserted into the cortical tissues1,2,3. To simultaneously record multiple LFP signals from the cortical regions of small mammals, such as rats and mice, a number of studies have developed various types of custom-made electrode assemblies that are termed micro-drives. A conventional micro-drive is composed of metal screws attached to the middle parts of the electrodes (which are typically tetrodes), a core body that accommodates the screws and electrodes, and an electrical interface board (EIB) that accommodates metal holes to connect the open ends of the electrodes (Figure 1, Figure 2, and Figure 3). This electrode assembly enables the operator to control the depth of many electrodes inserted into the brain over the course of days to weeks, and allows the conducting of long-term chronic recordings of neuronal activity as the animal is challenged with various behavioral tasks. In the peripheral organs, heartbeat signals are recorded as electrocardiograms (ECGs) by a pair of electrodes that are implanted on or around the heart area4,5,6, and skeletal muscle signals are recorded as electromyograms (EMGs) with electrodes that are inserted into the muscle tissue7,8,9. The relationship between electrical signals of the olfactory bulb and breathing (BR) rhythm has been studied with single unit recordings10,11. In conventional recording systems, these signals from different tissues have been captured by independent recording devices, which means that an additional experimental system is required to precisely synchronize these multiple devices for simultaneous recordings of brain-body signals. This system was developed to overcome this issue. In this system, all electrical signals recorded from the peripheral organs, including ECGs, EMGs, and electrical signals from the olfactory bulb that reflect the breathing rhythm, are integrated into a single micro-drive array1,2,3, here termed an integrative micro-drive array. This system requires only one multi-channel recording device, and is applicable to any conventional micro-drive array. The advantages of this technique are that it does not require any special devices or trigger signals to match the recording time of multiple devices, and it allows for more convenient data processing, since all of the signals are recorded as similar data types. This technique will aid the understanding of the neurophysiological correlates of the associations between central and peripheral organs. This paper describes the procedures associated with the technique and presents representative datasets obtained from a rat.

<table><tbody><tr><td>FEP Hookup Wire Stranded Stainless Steel&amp;nbsp;</td><td>Cooner Wire Company, Chatsworth, CA</td><td>AS 633</td><td>Bioflex wire</td></tr><tr><td>EIB-36-PTB</td><td>Neuralynx, Inc., Bozeman, MT</td><td>EIB-36-PTB</td><td>EIB</td></tr><tr><td>Cereplex&amp;nbsp; M</td><td>Blackrock&amp;nbsp; Microsystems, Salt Lake City, UT</td><td></td><td>Digital headstage</td></tr><tr><td>Cereplex Direct&amp;nbsp;</td><td>Blackrock&amp;nbsp; Microsystems, Salt Lake City, UT</td><td></td><td>Data acquisition system</td></tr><tr><td>UEW polyurethane magnet wire</td><td>Oyaide.com, Tokyo, Japan</td><td>UEW 0.14mm 20m&amp;nbsp;</td><td>Enamel wire</td></tr><tr><td>SD-102</td><td>Narishige, Tokyo, Japan</td><td>SD-102</td><td>High-speed drill</td></tr><tr><td>Minimo ONE SERIES ver.2</td><td>Minitor Co.,Ltd, Tokyo, Japan</td><td>C2012</td><td>High-peed drill Power Supply&amp;nbsp;</td></tr><tr><td>Provinice 250 mL</td><td>Shofu Inc., Kyoto, Japan</td><td>213620136</td><td>Dental cement</td></tr><tr><td>Small Animal Anesthetizer&amp;nbsp;</td><td>Biomachinery, Chiba, Japan</td><td>TK-7</td><td>Anesthetizer&amp;nbsp;</td></tr><tr><td>Buprenorphine hydrochloride</td><td>Sigma-Aldrich, St. Louis, MO</td><td>B7536-1ML</td><td>Analgesic</td></tr><tr><td>Isoflurane</td><td>DS Pharma Animal Health, Osaka, Japan&amp;nbsp;</td><td>Isoflu 250mL</td><td></td></tr><tr><td>Vaseline, White&amp;nbsp;</td><td>Wako Pure Chemical Industries, Ltd., Osaka, Japan</td><td>224-00165&amp;nbsp;</td><td>Vet ointment&amp;nbsp;</td></tr><tr><td>&amp;nbsp;Sodium alginate</td><td>Nacalai tesque, Kyoto, Japan</td><td>31131-85</td><td></td></tr><tr><td>Calcium Chloride Dihydrate</td><td>Wako Pure Chemical Industries, Ltd., Osaka, Japan</td><td>031-00435&amp;nbsp;</td><td></td></tr><tr><td>Stainless steel screw M1.0&amp;times;4.0&amp;nbsp;</td><td>MonotaRO, Hyogo, Japan</td><td>42617504</td><td>Stainless steel screw for BR electrodes</td></tr><tr><td>Stainless steel screw M1.4&amp;times;3.0</td><td>MonotaRO, Hyogo, Japan</td><td>42617687</td><td>Stainless steel screw for g/r electrodes and anchors</td></tr></tbody></table>

simultaneous recordings of cortical local field potentials, electrocardiogram, electromyogram, and breathing rhythm from a freely moving rat

Monitoring the physiological dynamics of the brain and peripheral tissues is necessary for addressing a number of questions about how the brain controls body functions and internal organ rhythms when animals are exposed to emotional challenges and changes in their living environments. In general experiments, signals from different organs, such as the brain and the heart, are recorded by independent recording systems that require multiple recording devices and different procedures for processing the data files. This study describes a new method that can simultaneously monitor electrical biosignals, including tens of local field potentials in multiple brain regions, electrocardiograms that represent the cardiac rhythm, electromyograms that represent awake/sleep-related muscle contraction, and breathing signals, in a freely moving rat. The recording configuration of this method is based on a conventional micro-drive array for cortical local field potential recordings in which tens of electrodes are accommodated, and the signals obtained from these electrodes are integrated into a single electrical board mounted on the animal's head. Here, this recording system was improved so that signals from the peripheral organs are also transferred to an electrical interface board. In a single surgery, electrodes are first separately implanted into the appropriate body parts and the target brain areas. The open ends of all of these electrodes are then soldered to individual channels of the electrical board above the animal's head so that all of the signals can be integrated into the single electrical board. Connecting this board to a recording device allows for the collection of all of the signals into a single device, which reduces experimental costs and simplifies data processing, because all data can be handled in the same data file. This technique will aid the understanding of the neurophysiological correlates of the associations between central and peripheral organs.

This method can simultaneously capture bioelectrical signals from multiple organs that represent the neuronal activity of the brain, heart rate, breathing rhythm, and skeletal muscle contractions (Figure 1). Figure 4 provides representative recording data from a freely moving rat that was freely foraging in a rectangular box (25 &#215; 40 cm2). The example dataset includes typical behavioral transitions between moving and resting states. A power spectrum was computed from a hippocampal LFP trace by wavelet analysis. The BR signal recorded from the surface area of the olfactory bulb was used to roughly estimate the relative changes in breathing frequencies, such as those that occur during exploratory sniffing behavior.
<img alt="Figure 1" class="xfigimg" src="/files/ftp_upload/56980/56980fig1.jpg" /> 
Figure 1: Illustration of the recording system for monitoring multiple brain-body signals from a freely moving rat. All bioelectrical signals (LFP, ECG, EEG, BR signals) from a freely moving rat are collected into the integrative-microdrive array mounted on the head. <a href="https://cloudflare-jove-com.bdigitaluss.remotexs.co/files/ftp_upload/56980/56980fig1large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 2" class="xfigimg" src="/files/ftp_upload/56980/56980fig2.jpg" /> 
Figure 2: An integrative micro-drive array. All LFP, ECG, EEG, and BR signals are transmitted to the holes on the EIB as indicated by the arrows. The dotted area is magnified in the right panel and displays some of the tetrodes protruding from the micro-drive array that are inserted into the brain tissue. <a href="https://cloudflare-jove-com.bdigitaluss.remotexs.co/files/ftp_upload/56980/56980fig2large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 3" class="xfigimg" src="/files/ftp_upload/56980/56980fig3.jpg" /> 
Figure 3: A top view of the EIB. The EIB includes 24 cortical LFP (LFP) channels that are connected to the tetrodes, 2 ECG channels, 2 EMG channels, 2 BR channels, and 2 ground (Gr) channels. All channels except the LFP channels were connected to insulated wires. <a href="https://cloudflare-jove-com.bdigitaluss.remotexs.co/files/ftp_upload/56980/56980fig3large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 4" class="xfigimg" src="/files/ftp_upload/56980/56980fig4v2.jpg" /> 
Figure 4: An example of the simultaneous multi-channel recording of bioelectrical signals. 
(From top to bottom) LFP signals in the somatosensory cortex (scale bar: 250 &#181;V). LFP signals in the hippocampal CA1 region (scale bar: 500 &#181;V). A color-coded power spectrum of the hippocampal LFP trace. An ECG signal (band-pass filtered at 20-200 Hz, scale bar: 500 &#181;V). An EMG signal (band-pass filtered at 100-500 Hz, scale bar: 100 &#181;V). A BR signal (scale bar: 500 &#181;V). A color-coded power spectrum of the BR signal that indicates sniffing behavior as defined by a transient increase in breathing rate. <a href="https://cloudflare-jove-com.bdigitaluss.remotexs.co/files/ftp_upload/56980/56980fig4v2large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>

Watch this Scientific Journal Video about Simultaneous Recordings of Cortical Local Field Potentials, Electrocardiogram, Electromyogram, and Breathing Rhythm from a Freely Moving Rat at JoVE.com

Simultaneous Recordings of Cortical Local Field Potentials, Electrocardiogram, Electromyogram, and Breathing Rhythm from a Freely Moving Rat

This study introduces a method for the simultaneous recording of local field potentials in the brain, electrocardiograms, electromyograms, and breathing signals of a freely moving rat. This technique, which reduces experimental costs and simplifies data analysis, will contribute to the understanding of the interactions between the brain and peripheral organs.

Monitoring the physiological dynamics of the brain and peripheral tissues is necessary for addressing a number of questions about how the brain controls ...

simultaneous-recordings-cortical-local-field-potentials

Universidad San Sebastian

Research

JoVE Journal

Neuroscience

10.8K Views.  The University of Tokyo. This study introduces a method for the simultaneous recording of local field potentials in the brain, electrocardiograms, electromyograms, and breathing signals of a freely moving rat. This technique, which reduces experimental costs and simplifies data analysis, will contribute to the understanding of the interactions between the brain and peripheral organs.

Video: Simultaneous Recordings of Cortical Local Field Potentials, Electrocardiogram, Electromyogram, and Breathing Rhythm from a Freely Moving Rat

Simultaneous fMRI and Electrophysiology in the Rodent Brain

To examine the neural basis of the blood oxygenation level dependent (BOLD) magnetic resonance imaging (MRI) signal, we have developed a rodent model in which functional MRI data and in vivo intracortical recording can be performed simultaneously. The combination of MRI and electrical recording is technically challenging because the electrodes used for recording distort the MRI images and the MRI acquisition induces noise in the electrical recording. To minimize the mutual interference of the two modalities, glass microelectrodes were used rather than metal and a noise removal algorithm was implemented for the electrophysiology data. In our studies, two microelectrodes were separately implanted in bilateral primary somatosensory cortices (SI) of the rat and fixed in place. One coronal slice covering the electrode tips was selected for functional MRI. Electrode shafts and fixation positions were not included in the image slice to avoid imaging artifacts. The removed scalp was replaced with toothpaste to reduce susceptibility mismatch and prevent Gibbs ringing artifacts in the images. The artifact structure induced in the electrical recordings by the rapidly-switching magnetic fields during image acquisition was characterized by averaging all cycles of scans for each run. The noise structure during imaging was then subtracted from original recordings. The denoised time courses were then used for further analysis in combination with the fMRI data. As an example, the simultaneous acquisition was used to determine the relationship between spontaneous fMRI BOLD signals and band-limited intracortical electrical activity. Simultaneous fMRI and electrophysiological recording in the rodent will provide a platform for many exciting applications in neuroscience in addition to elucidating the relationship between the fMRI BOLD signal and neuronal activity.

We have developed a method for simultaneous functional magnetic resonance imaging and electrophysiological recording in the rodent brain, providing a platform for the investigation of the relationship between neural activity and the blood oxygenation level dependent (BOLD) MRI signal.

To examine the neural basis of the blood oxygenation level dependent (BOLD) magnetic resonance imaging (MRI) signal, we have developed a rodent model in ...

Simultaneous Recordings of Cortical Local Field Potentials and Electrocorticograms in Response to Nociceptive Laser Stimuli from Freely Moving Rats

Electrocortical responses, elicited by laser heat pulses that selectively activate nociceptive free nerve endings, are widely used in many animal and human studies to investigate the cortical processing of nociceptive information. These laser-evoked brain potentials (LEPs) consist of several transient responses that are time-locked to the onset of laser stimuli. However, the functional properties of the LEP responses are still largely unknown, due to the lack of a sampling technique that can simultaneously record neural activities at the surface of the cortex (i.e., electrocorticogram [ECoG] and scalp electroencephalogram [scalp EEG]) and inside the brain (i.e., local field potential [LFP]). To address this issue, we present here an animal protocol using freely moving rats. This protocol is composed of three main procedures: (1) animal preparation and surgical procedures, (2) a simultaneous recording of ECoG and LFP in response to nociceptive laser stimuli, and (3) data analysis and feature extraction. Specifically, with the help of a 3D-printed protective shell, both ECoG and LFP electrodes implanted on the rat&#39;s skull were securely held together. During data collection, laser pulses were delivered on the rat&#39;s forepaws through gaps in the bottom of the chamber when the animal was in spontaneous stillness. Ongoing white noise was played to avoid the activation of the auditory system by the laser-generated ultrasounds. As a consequence, only nociceptive responses were selectively recorded. Using the standard analytical procedures (e.g., band-pass filtering, epoch extraction, and baseline correction) to extract stimulus-related brain responses, we obtained results showing that LEPs with a high signal-to-noise ratio were simultaneously recorded from ECoG and LFP electrodes. This methodology makes the simultaneous recording of ECoG and LFP activities possible, which provides a bridge of electrocortical signals at the mesoscopic and macroscopic levels, thereby facilitating the investigation of nociceptive information processing in the brain.

We developed a technique that simultaneously records both electrocorticography and local field potentials in response to nociceptive laser stimuli from freely moving rats. This technique helps establish a direct relationship of electrocortical signals at the mesoscopic and macroscopic levels, which facilitates the investigation of nociceptive information processing in the brain.

Electrocortical responses, elicited by laser heat pulses that selectively activate nociceptive free nerve endings, are widely used in many animal and ...

Behavior

Recording EEG in Freely Moving Neonatal Rats Using a Novel Method

EEG is a useful method to detect electrical activity in the brain. Moreover, it is a widely used diagnostic tool for various neurological conditions, such as epilepsy and neurodegenerative disorders. However, it is technically difficult to obtain EEG recordings in neonates as it requires specialized handling and great care. Here, we present a novel method to record EEG in neonatal rat pups (P8-P15). We designed a simple and reliable electrode using computer pin loci; it can be easily implanted into the skull of a rat pup to record high-quality EEG signals in the normal and epileptic brain. Pups were given an intraperitoneal (i.p.) injection of the neurotoxin kainic acid (KA) to induce epileptic seizures. The surgical implantation performed in this procedure is less expensive than other EEG procedures for neonates. This method allows one to record high-quality and stable EEG signals for more than 1 week. Furthermore, this procedure can also be applied to adult rats and mice to study epilepsy or other neurological disorders.

Here, we introduce a novel technique designed to record electroencephalography (EEG) in freely moving neonatal epileptic pups and describe its procedures, features, and applications. This method allows one to record EEG for more than 1 week.

EEG is a useful method to detect electrical activity in the brain. Moreover, it is a widely used diagnostic tool for various neurological conditions, such ...

Concurrent Recording of Co-localized Electroencephalography and Local Field Potential in Rodent

Although electroencephalography (EEG) is widely used as a non-invasive technique for recording neural activities of the brain, our understanding of the neurogenesis of EEG is still very limited. Local field potentials (LFPs) recorded via a multi-laminar microelectrode can provide a more detailed account of simultaneous neural activity across different cortical layers in the neocortex, but the technique is invasive. Combining EEG and LFP measurements in a pre-clinical model can greatly enhance understanding of the neural mechanisms involved in the generation of EEG signals, and facilitate the derivation of a more realistic and biologically accurate mathematical model of EEG. A simple procedure for acquiring concurrent and co-localized EEG and multi-laminar LFP signals in the anesthetized rodent is presented here. We also investigated whether EEG signals were significantly affected by a burr hole drilled in the skull for the insertion of a microelectrode. Our results suggest that the burr hole has a negligible impact on EEG recordings.

This protocol describes a simple method for concurrent recording of co-localized electroencephalography (EEG) and multi-laminar local field potential in an anesthetized rat. A burr hole drilled in the skull for the insertion of a microelectrode is shown to produce negligible distortion of the EEG signal.

Although electroencephalography (EEG) is widely used as a non-invasive technique for recording neural activities of the brain, our understanding of the ...

Simultaneous Electrocardiography Recording and Invasive Blood Pressure Measurement in Rats

For studies related to cardiovascular physiology or pathophysiology, blood pressure (BP) and electrocardiography are basic observational parameters. Research focusing on cardiovascular disease models, potential cardiovascular therapeutic targets or pharmaceutical agents requires assessment of systemic arterial pressure and heart rhythm changes. In situations where radio telemetry systems are not available or affordable, the technique of femoral artery cannulation is an alternative way to obtain intra-arterial pressure waveform recordings and systemic BP measurements. This technique is economical and can be performed with standard equipment in animal facilities. However, invasive arterial pressure recording requires cannulation of small arteries, which can be a challenging surgical skill. Here, we present step-by-step protocols for femoral artery cannulation procedures. Key procedures include the calibration of the data acquisition system, tissue dissection and femoral artery cannulation, and setup of the arterial cannulation system for pressure recording. Surface electrocardiography recording procedures are also included. We also present examples of BP recordings from normotensive and hypertensive rats. This protocol allows reliable direct recordings of systemic BP with simultaneous electrocardiography.

Here, we describe a setup for simultaneous recording of electrocardiography and intra-arterial blood pressure (BP) in experimental rats, which can be done with standard equipment in animal facilities and can be applied to physiological or pharmacological studies to investigate pathogenic or therapeutic mechanisms in cardiovascular medicine.

For studies related to cardiovascular physiology or pathophysiology, blood pressure (BP) and electrocardiography are basic observational parameters. ...

Medicine

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

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.

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 ...

Multisystem Monitoring of Epileptic Abnormalities in Rabbit: A Method for Simultaneous Video EEG, ECG, Capnography, and Oximetry Recording During Induced Seizures

Source: Bosinski, C., et al. <a href="https://www-jove-com.bdigitaluss.remotexs.co/v/62256">Multi-system Monitoring for Identification of Seizures, Arrhythmias and Apnea in Conscious Restrained Rabbits.</a> J. Vis. Exp. (2021).
In this video, we demonstrate a multisystem monitoring method to evaluate neurological and cardiorespiratory abnormalities during intravenous injection of a seizure-inducing drug. The simultaneous monitoring of video EEG, ECG, oximetry, and capnography parameters helps to understand the real-time physiological changes associated with epileptic seizures.

Source: Bosinski, C., et al. Multi-system Monitoring for Identification of Seizures, Arrhythmias and Apnea in Conscious Restrained Rabbits. J. Vis. Exp. ...

JoVE Encyclopedia of Experiments

Biology

Large Animal Models

Leporine

In Vivo Electrophysiological Recordings of Brain Activities from Multiple Sites in Rats

Source: Haumesser, J. K., et al. <a href="https://app-jove-com.bdigitaluss.remotexs.co/v/55940">Acute In Vivo Electrophysiological Recordings of Local Field Potentials and Multi-unit Activity from the Hyperdirect Pathway in Anesthetized Rats.</a> J. Vis. Exp.(2017).
This video demonstrates the detailed procedure for preparing an anesthetized rat for electrophysiological recordings of brain activity from multiple sites. The scalp is dissected to access the skull, and holes are drilled at precise locations to install and secure electrodes. Tungsten microwire electrodes are then inserted into the targeted location to record the data.

All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board.
1. ...

Neurophysiology

Electrophysiology Techniques

Electroencephalographic Recording of Epileptic Seizures in Epilepsy-Induced Rats

Source: Soukupov&#225;, M., et al. <a href="https://app-jove-com.bdigitaluss.remotexs.co/v/58455">Microdialysis of Excitatory Amino Acids During EEG Recordings in Freely Moving Rats.</a> J. Vis. Exp. (2018)
This video demonstrates electroencephalographic (EEG) recording of epileptic seizures in epilepsy-induced rats. A rat with electrodes implanted in the ventral hippocampus is connected to the EEG recording system and placed in a plexiglass chamber. The rat is allowed to move freely while the brain activity is recorded to monitor the occurrence of seizures.

Stimulation and Modulation Techniques

A Micro-drive Array Method for Electrophysiological Recording from Multiple Brain Regions

Source: Shikano, Y., et al. <a href="https://app-jove-com.bdigitaluss.remotexs.co/v/56980">Simultaneous Recordings of Cortical Local Field Potentials, Electrocardiogram, Electromyogram, and Breathing Rhythm from a Freely Moving Rat.</a> J. Vis. Exp. (2018).
The video demonstrates simultaneous electrophysiological recording from multiple brain regions in a rat using an integrative micro-drive array. Recording electrodes are placed in the olfactory bulb, reference electrodes in the frontal cortex, and micro-drive array electrodes in the hippocampus. The rat is allowed to forage freely, and simultaneous recording from all the implanted electrodes is recorded using the array.