This work was supported by the National Institute of Health (RO1 NS120945, R37NS119012 to JK) and the UVA Brain Institute.

The present work is approved by the University of Virginia Animal Care and Use Committee. C57Bl/6 mice of both sexes (7-12 weeks) were used for the experiments. The animals were maintained on a 12 h light/12 h dark cycle and had ad libitum access to food and water.
1. Microelectrode construction
<ol>
	<li>To construct the microelectrodes, use 50 &#181;m (diameter) diamel-coated nickel-chromium wire (see Table of Materials). Tape one end of the wire at the back of the platform and wrap the wire three times around the nearest knob on the platform (Figure 1A,C). 
	NOTE: An acrylic platform with two knobs (2 x 5 inches) was used here, but any platform can be used.
	<ol>
		<li>Stretch the wire around the farthest second knob to make two loops between the knobs. Wrap the wire three more times around the first knob to fix the wire in place and tape the end again at the back of the platform. 
		NOTE: After the wires are separated (steps 1.2-1.3.1), there must be two wires on each side (four wires in total, Figure 1B).</li>
	</ol>
	</li>
	<li>Place the tension bars under the wires with the tape wrapped around them (sticky side up) (Figure 1C). 
	NOTE: Triangular acrylic pieces were used for the tension bars, with tape wrapped around them (sticky side outside to attach the wires). The sticky side of the tape outside of the tension bars will keep the wires in place to adjust the distance between them. The tension bars must be ~2.5 cm away from the knobs, and the wires must not be loose.</li>
	<li>Using a microscope and fine forceps, make either a 3 mm or 4.5 mm gap between the wires (3 mm gap between the wires to make cortical (Ctx) - ventrolateral thalamic nucleus (VL) microelectrodes; 4.5 mm gap to make striatal (Str) - nigral (SNR) microelectrodes) (Figure 1B).
	<ol>
		<li>If magnification is used on the microscope, ensure to calculate and adjust for the difference in magnification and the actual distance between the wires. 
		NOTE: If microelectrodes are constructed for structures other than those used here, the distance between the wires needs to be adjusted to the stereotaxic distance between the structures. Figure 2B provides an example of how the wires will be organized; accordingly, the stereotaxic coordinates for other structures must be adjusted.</li>
	</ol>
	</li>
	<li>Cut four small pieces of plastic (0.5 mm thick) ~6 mm (width) x 3 mm (height) (Figure 1C). 
	NOTE: Any plastic pieces can be used as long as they are 0.5 mm thick; here, square tubing was used in which the pins were sold (Pins, see Table of Materials). If a different thickness is used, please add more or fewer pieces of plastic to fit the required stereotaxic coordinates.</li>
	<li>Apply glue (see Table of Materials) on the plastic and place them on the wires (Figure 1C). Place plastic pieces ~1.0 cm away from the middle of the wire, which is 1.0 cm away from the tension bar. Remove the excess of superglue with a cotton swab.</li>
	<li>After the superglue dries, cut the wires using fine scissors, in the order indicated in Figure 1C.</li>
	<li>Cut four 7 mm glass tubes using a commercially available kit (see Table of Materials) and insert the electrode wires into the glass tubes as indicated in Figure 2A.
	<ol>
		<li>Insert VL and SNR electrode pairs into the glass tubes. 
		NOTE: Only the wires for deep structures need to be inserted into the glass tubes to support surgical implantation. Ensure not to insert cortical electrodes into the glass tube.</li>
	</ol>
	</li>
	<li>Put the glue at the base of the glass tubes to connect them to the plastic. Wait some time until the glue dries.</li>
	<li>Cut the glass tubes and wires using a scalpel as indicated in Table 1; ensure that the lengths of the microelectrodes are correct. If the microelectrodes target different structures, adjust the cutting distance according to the required stereotaxic coordinates.</li>
</ol>
<img alt="Figure 1" class="xfigimg" src="/files/ftp_upload/63633/63633fig01.jpg" /> 
Figure 1: Schematic of the microelectrode construction. (A) Set up of wires on the platform with tension bars below the wires. (B) The gap between the wires. (C) Four pieces of plastic are glued to the wires. <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/63633/63633fig01large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td></td>
			<td>Ctx</td>
			<td>Str</td>
			<td>VL</td>
			<td>SNR</td>
		</tr>
		<tr>
			<td>AP (Anterior/Posterior)</td>
			<td>2.2</td>
			<td>1.2</td>
			<td>-1.3</td>
			<td>-3.3</td>
		</tr>
		<tr>
			<td>ML (Medial/Lateral)</td>
			<td>1.8</td>
			<td>1.5</td>
			<td>1</td>
			<td>1.5</td>
		</tr>
		<tr>
			<td>DV (Dorsal/Ventral)</td>
			<td>0.5</td>
			<td>3.5</td>
			<td>4</td>
			<td>4.75</td>
		</tr>
		<tr>
			<td>Electrode length</td>
			<td>4</td>
			<td>4.75</td>
			<td>5.25</td>
			<td>6</td>
		</tr>
	</tbody>
</table>
Table 1: Stereotaxic implantation coordinates and dimensions of the microelectrodes.
2. Microelectrode array assembly
<ol>
	<li>Use the glue to attach plastics in the desired order of target regions. An example for cortical, thalamic, striatal, and nigral electrodes is shown in Figure 2B,C.
	<ol>
		<li>Place Ctx-VL electrode pair face down (side with electrode wires has to face down) and connect two 6 mm x 3 mm empty pieces of plastic on top with glue.</li>
		<li>On top of the three pieces of plastic, place the second Ctx-VL electrode pair with the electrodes facing upwards (use a microscope and ensure that the VL electrodes are aligned). 
		NOTE: Alignment of bilateral electrodes (here, the alignment of left and right VL electrodes) is essential to target desired bilateral structures appropriately.</li>
		<li>Use the glue to attach the SNR electrodes on top with the SNR electrodes 2.0 mm away from the VL electrodes and ~5.0 mm away from the cortical electrodes (the SNR electrode wires have to face upwards).</li>
		<li>Repeat step 2.1.3. for the other side (the SNR electrode wires have to face outside of the microelectrode array).</li>
	</ol>
	</li>
	<li>Apply epoxy resin around the plastic to bind the electrodes together. Avoid putting epoxy resin on the electrodes.</li>
	<li>Take a thick wire and make a loop on one end. Dip the loop in the epoxy solution and place it on the plastic, ensuring the thick wire is lying flat (Figure 2D) so that for the next steps, this wire could be used as a handle. Wait until the electrodes are fully dry.</li>
	<li>Cut the wires to 2 cm, as shown in Figure 2E.</li>
</ol>
<img alt="Figure 2" class="xfigimg" src="/files/ftp_upload/63633/63633fig02v2.jpg" /> 
Figure 2: Microelectrode construction and dimensions. (A) Four pairs of electrodes formed after the wires were cut with scissors, as indicated in Figure 1C (2 pairs of Ctx-VL electrodes and 2 pairs of Str-SNR electrodes). Insert deep structure electrodes (VL and SNR) into the glass tubes and glue their bases to plastic (red dots). (B) Top view: The electrode pairs from (A) are glued in a stack to create the microelectrode core. Red lines indicate glue lines. (C) Front side view of (B). (D) The thick wire was attached to the microelectrodes. (E) The wires are grouped as indicated, and the isolated ends are scraped off and cut into 2 cm. <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/63633/63633fig02largev2.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
3. Microelectrode connection to the headset
<ol>
	<li>Group the wires as indicated in Figure 2E and scrape away 1 mm of the isolated ends with a scalpel.</li>
	<li>Bend the cortical electrodes as shown in Figure 3A. Separate the wires as shown in Figure 3B. Using fine forceps, make a loop at the end of each wire (Figure 3B).</li>
	<li>Hold a 10-pin headset with a hemostat (see Table of Materials) and use the wooden end of a cotton swab to apply minimal amounts of flux on the pins (Figure 3C). Ensure not to put flux outside the pins to prevent short-circuit among the pins.</li>
	<li>Using the wooden end of a cotton swab, apply flux to the wire loops.</li>
	<li>Solder the wire loops to the 10-pin headset as shown in Figure 3C. After soldering, dry the headset to prevent short-circuit among the pins.</li>
	<li>Take a thin wire (0.005-0.008 inch) for the reference and ground wires and strip off the plastic from one end. Make a loop on the other end of the wire.</li>
	<li>Solder the stripped side of the reference and ground wires to their respective pins (Figure 3A,C).</li>
	<li>Holding on the thick wire (Figure 2D), apply cranioplasty cement around the microelectrodes, especially where the wires connect to the pins. Avoid touching the actual electrode ends with the cement.</li>
	<li>After the cement dries, put epoxy resin at the base of the glass tubes, striatal microelectrode wires, and the whole electrode. Avoid touching the actual electrode ends with epoxy resin. Wait until the electrodes are fully dry.</li>
	<li>The electrodes are ready. Drill holes in the mouse&#39;s skull (as per the required stereotaxic coordinates) using a dental drill and implant the headset as shown in Figure 3D by lowering the headset with microelectrodes facing the skull and the appropriate holes. The headset could be attached to the stereotaxic arm for support during the implantation.</li>
</ol>
<img alt="Figure 3" class="xfigimg" src="/files/ftp_upload/63633/63633fig03.jpg" /> 
Figure 3: Microelectrode implantation. (A) The cortical electrodes are bent as indicated. (B) The wires are separated to make loops at the ends. (C) The flux (at the red dots) and looped wires are soldered to the 10-pin headset, ensuring that each wire goes to its appropriate pin. (D) The headset is implanted to record LFPs. <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/63633/63633fig03large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
4. Marking electrode location after recordings
<ol>
	<li>At the end of the LFP recordings, confirm the correct position of the electrodes in the target region by applying a current at the electrode tips to make a lesion and waiting for 30 min before perfusing the mouse. 
	NOTE: Lesion settings to confirm the location of the electrode tips: Single burst, 40 &#181;A, 0.75 ms monophasic square wave pulse, 50 Hz, 30 s.</li>
	<li>Anesthetize the mice with isoflurane (until the mouse fell asleep) and perfuse10&#160;transcardially with 4% paraformaldehyde (PFA) in 0.1 M sodium phosphate buffer. Section the brains (40 &#181;m thick) on a cryostat (see Table of Materials) and stain with DAPI (0.02% in PBS). Confirm the correct location of the electrodes by the presence of the electrode tip lesions as shown in Figure 4B,C. 
	NOTE: The percentage of the isoflurane needs to be applied following the individual institution&#39;s guidelines.</li>
</ol>
5. Measuring the electrode resistance
<ol>
	<li>Measure the resistance of the electrodes and check the short-circuit among the electrodes using a multi-range ohmmeter (see Table of Materials). Set the resistance scale at R x 10,000, indicating that a unit deflection in the pointer corresponds to 1 k&#937; resistance. 2 cm long electrodes need to have 60-70 k&#937; resistance.</li>
	<li>Customize the headset by taking ten individual pins (see Table of Materials). Solder each pin with a thin, multi-stranded copper wire in a cable.
	<ol>
		<li>Press-fit the soldered pins with their corresponding cable tails into the 10-pin double-row mating socket (matching sockets). Press-fit the open-ended pins of the mating socket into the LFP headset. In this way, each LFP electrode has a designated cable wire in the assembly.</li>
	</ol>
	</li>
	<li>Dip the tip of each LFP electrode (the resistance of which is to be measured) into 0.9% NaCl saline solution (NaCl concentration in the blood). Connect the cable wire end that corresponds to the LFP electrode to the positive terminal of the ohmmeter.
	<ol>
		<li>Use a low resistance ( &#820;100 &#937;) wire with one side in the saline water and the other side as the open end. Connect the open end of the low-resistance wire to the ground of the ohmmeter pointer. 
		NOTE: This arrangement completes the circuit and prompts a deflection of the ohmmeter pointer.</li>
	</ol>
	</li>
	<li>Ensure that there is no electrical connection between any two electrodes on the headset. Check the electrical insulation of pairwise LFP electrodes (ground: one cable corresponds to one electrode; positive: another cable corresponds to another electrode). Discard the electrodes if any deflection is observed in this case.</li>
</ol>

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The authors have nothing to disclose.

Historically, microelectrode arrays have been extensively used to record neuronal activity from a specific brain region of interest2,3,4,5,6,7,8,9,13. However, our easy microelectrode design allows recording from multiple structures simultaneously11,12. Here,&#160;the construction of the cortical, thalamic, striatal, and nigral microelectrodes are described as an example. Investigators can modify the microelectrode design to fit any desired structure by calculating the necessary stereotaxic distances and adjusting the construction accordingly.
For example, we have previously modified the design of these microelectrode arrays to record LFPs in the lamellar and septotemporal direction in the hippocampus12. A 50 &#181;m spacing wire separated adjacent electrodes as four microelectrodes recorded along the hippocampal lamina to prevent cross-contamination of the signal. Although those were not single-unit recordings, each electrode represented a small group of neurons as indicated by the variability of a spike waveform as a function of distance from the cell body.
During the construction, insertion of the thalamic and nigral microelectrode wires into glass tubes was necessary to provide stability during implantation surgery to target those deep structures. There were eight bilateral microelectrodes, four of which had glass tubes (2 VL and 2 SNR), which were a limit before elevating intracranial pressure and increasing mortality. Generally, glass tubes are needed when the desired insertion depth is at least 2 mm.
Also, 0.5 mm thick plastic was needed, limiting the minimum distance separation between the electrodes to 0.5 mm, but other plastics could be used. In the present case, plastics were placed along the major axis of the headset. Plastics can also be placed across the headset axis where several electrodes have identical anterior-posterior (AP) but different medial-lateral (ML) coordinates. This method offers a wide range of possible configurations for specific brain regions.
The number of pins on a headset limits the number of microelectrodes. A headset containing 12 pins covers the anterior-posterior extent of an adult mouse head completely. Each pin should be isolated from the other pins during soldering. An ohmmeter and 0.9% saline water were needed to test the electrical isolation for each pair of electrode terminals. The 12-pin headset limits the recording to 10 regions (2 are reserved for the ground and reference).

Local field potentials (LFPs) are the electric potentials recorded from the extracellular space in the brain. They are generated by ion concentration imbalances outside of neurons and represent the activity of a small, localized population of neurons, allowing to precisely monitor the activity of a specific brain region compared to the macroscale EEG recordings1. As an estimate, the LFP microelectrodes separated by 1 mm correspond to two completely different populations of neurons. While EEG signal is filtered by brain tissue, cerebrospinal fluid, skull, muscle, and skin, LFP signal is a reliable marker of local neuronal activity1.
Researchers often need to simultaneously record LFPs from several brain structures, but commercially available microelectrode arrays often do not offer such flexibility. Here, the present protocol describes fully customizable, easily constructed microelectrodes to simultaneously record LFPs from any desired brain region at different depths. Although LFPs have extensively been used to record the neuronal activity of a specific brain region2,3,4,5,6,7,8,9, the current easy customizable design allows recording LFPs from any multiple superficial or deep brain regions11,12. The protocol can also be modified to construct any desired microelectrode array by determining stereotaxic coordinates of the brain regions and assembling the array accordingly. These microelectrodes with a 10 kHz sampling rate and 60-70 k&#8486; resistance (2 cm length) allow us to record LFPs with millisecond precision. The data can then be amplified by a 16-channel amplifier, filtered (low pass 1 Hz, high pass 5 kHz), and digitized.

<table><tbody><tr><td>Amplifier 16-Channel</td><td>A-M Systems</td><td>Model 3600</td><td>Amplifier</td></tr><tr><td>Cranioplasty cement</td><td>Coltene</td><td>Perm Reeline/Repair Resin Type II Class I Shade - Clear</td><td>Cement to hold microelectrodes</td></tr><tr><td>Cryostat Microtome</td><td>Precisionary</td><td>CF-6100</td><td>To slice brain</td></tr><tr><td>Diamel-coatednickel-chromium wire</td><td>Johnson Matthey Inc.</td><td>50 &amp;micro;m</td><td>Microelectrode wire</td></tr><tr><td>Dremel</td><td>Dremel</td><td>300 Series</td><td>To drill holes in mouse skull</td></tr><tr><td>Epoxy</td><td>CEC Corp</td><td>C-POXY 5</td><td>Fast setting adhesive</td></tr><tr><td>Hemostat</td><td>Any</td><td></td><td>To hold the headset</td></tr><tr><td>Forceps</td><td>Any</td><td></td><td>To hold microelectrodes</td></tr><tr><td>Light microscope</td><td>Nikon</td><td>SMZ-10</td><td>To see alignment</td></tr><tr><td>Ohmmeter</td><td>Any</td><td></td><td>To measurre resistance</td></tr><tr><td>Pins (Headers and matching Sockets)</td><td>Mill-Max</td><td>Interconnects, 833 series, 2 mm grid gull wing surface mount headers and sockets</td><td>To attach microelectrodes to</td></tr><tr><td>Polymicro Tubing Kit</td><td>Neuralynx</td><td>ID 100 &amp;plusmn; 04 &amp;micro;m, OD 164 &amp;plusmn; 06 &amp;micro;m, coating thickness 12 &amp;micro;m</td><td>Glass tubes</td></tr><tr><td>Pulse Stimulator</td><td>A-M Systems</td><td>Model 2100</td><td>To mark the microelectrode location at the end of the recordings</td></tr><tr><td>Scissors</td><td>Any</td><td></td><td>To cut microelectrodes</td></tr><tr><td>Superglue</td><td>Gorilla</td><td></td><td>Adhesive</td></tr><tr><td>Thick wire 0.008 in. &amp;ndash; 0.011 in.</td><td>A-M Systems</td><td>791900</td><td>Tick wire to hold the microelectrode array</td></tr><tr><td>Thin wire 0.005 in. - 0.008 in.</td><td>A-M Systems</td><td>791400</td><td>Thin wire for reference and ground</td></tr></tbody></table>

construction of local field potential microelectrodes for in vivo recordings from multiple brain structures simultaneously

Researchers often need to record local field potentials (LFPs) simultaneously from several brain structures. Recording from multiple desired brain regions requires different microelectrode designs, but commercially available microelectrode arrays often do not offer such flexibility. Here, the present protocol outlines the straightforward design of custom-made microelectrode arrays to record LFPs from multiple brain structures simultaneously at different depths. This work describes the construction of the bilateral cortical, striatal, ventrolateral thalamic, and nigral microelectrodes as an example. The outlined design principle offers flexibility, and the microelectrodes can be modified and customized to record LFPs from any structure by calculating stereotaxic coordinates and quickly changing the construction accordingly to target different brain regions in either freely moving or anesthetized mice. The microelectrode assembly requires standard tools and supplies. These custom microelectrode arrays allow investigators to easily design microelectrode arrays in any configuration to track neuronal activity, providing LFP recordings with millisecond resolution.

In this work, the LFP microelectrodes were used to map the seizure spread through the basal ganglia11. Simultaneous LFP recordings were performed from the right premotor cortex (where the seizure focus was) and the left VL, striatum, and SNR (Figure 4). Seizure start was identified as deflection of the voltage trace at least twice the baseline (Figure 4A, red arrow). The power spectrum plot11&#160;shows frequency distributions for the recorded LFPs (Figure 4A). Seizure onset latencies (red bars) could be compared between each structure with millisecond precision (Figure 4A). A current pulse was applied at the end of the recordings to mark and confirm the location of the electrode tips, forming a lesion (Figure 4B,C).
<img alt="Figure 4" class="xfigimg" src="/files/ftp_upload/63633/63633fig04.jpg" /> 
Figure 4: Representative LFP recordings. (A) A seizure was recorded from the right premotor cortex and left VL, striatum, and SNR using LFP microelectrodes with the corresponding power spectrums. The red arrow indicates seizure onset. The red horizontal bars indicate seizure onset delay in each structure. The brain schematic shows the position of the microelectrodes (red dots). (B,C) The structures were lesioned after the recordings to mark the location of the microelectrode tips in the VL and SNR. <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/63633/63633fig04large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>

Watch this Scientific Journal Video about Construction of Local Field Potential Microelectrodes for in vivo Recordings from Multiple Brain Structures Simultaneously at JoVE.com

Construction of Local Field Potential Microelectrodes for in vivo Recordings from Multiple Brain Structures Simultaneously

The present protocol describes the construction of custom-made microelectrode arrays to record local field potentials in vivo from multiple brain structures simultaneously.

Construction of Local Field Potential Microelectrodes for in vivo Recordings from Multiple Brain Structures Simultaneously

Researchers often need to record local field potentials (LFPs) simultaneously from several brain structures. Recording from multiple desired brain regions ...

construction-local-field-potential-microelectrodes-for-vivo

Nanyang Technological University

Research

JoVE Journal

Neuroscience

3.0K Views.  University of Virginia. Onderzoekers moeten vaak lokale veldpotentialen van meerdere structuren tegelijkertijd vastleggen. Ons eenvoudige micro-elektrodeontwerp stelt ons in staat om LFP's van verschillende hersenstructuren op verschillende diepten tegelijkertijd op te nemen. Commercieel verkrijgbare micro-elektroden missen vaak flexibiliteit en laten niet toe om op te nemen van meerdere hersenstructuren.Ons micro-elektrodeontwerp maakt het mogelijk om de constructie eenvoudig aan te passen aan elke gewenste ...

Video: Construction of Local Field Potential Microelectrodes for in vivo Recordings from Multiple Brain Structures Simultaneously

Examining Local Network Processing using Multi-contact Laminar Electrode Recording

Cortical layers are ubiquitous structures throughout neocortex1-4 that consist of highly recurrent local networks. In recent years, significant progress has been made in our understanding of differences in response properties of neurons in different cortical layers5-8, yet there is still a great deal left to learn about whether and how neuronal populations encode information in a laminar-specific manner.
Existing multi-electrode array techniques, although informative for measuring responses across many millimeters of cortical space along the cortical surface, are unsuitable to approach the issue of laminar cortical circuits. Here, we present our method for setting up and recording individual neurons and local field potentials (LFPs) across cortical layers of primary visual cortex (V1) utilizing multi-contact laminar electrodes (Figure 1; Plextrode U-Probe, Plexon Inc).
The methods included are recording device construction, identification of cortical layers, and identification of receptive fields of individual neurons. To identify cortical layers, we measure the evoked response potentials (ERPs) of the LFP time-series using full-field flashed stimuli. We then perform current-source density (CSD) analysis to identify the polarity inversion accompanied by the sink-source configuration at the base of layer 4 (the sink is inside layer 4, subsequently referred to as granular layer9-12). Current-source density is useful because it provides an index of the location, direction, and density of transmembrane current flow, allowing us to accurately position electrodes to record from all layers in a single penetration6, 11, 12.

A fundamental issue in our understanding of cortical circuitry is how networks in different cortical layers encode sensory information. Here we describe electrophysiological techniques utilizing multi-contact laminar electrodes to record single-units and local field potentials and present analyses to identify cortical layers.

Cortical layers are ubiquitous structures throughout neocortex1-4 that consist of highly recurrent local networks. In recent years, significant progress ...

Recording Large-scale Neuronal Ensembles with Silicon Probes in the Anesthetized Rat

Large scale electrophysiological recordings from neuronal ensembles offer the opportunity to investigate how the brain orchestrates the wide variety of behaviors from the spiking activity of its neurons. One of the most effective methods to monitor spiking activity from a large number of neurons in multiple local neuronal circuits simultaneously is by using silicon electrode arrays1-3.
Action potentials produce large transmembrane voltage changes in the vicinity of cell somata. These output signals can be measured by placing a conductor in close proximity of a neuron. If there are many active (spiking) neurons in the vicinity of the tip, the electrode records combined signal from all of them, where contribution of a single neuron is weighted by its 'electrical distance'. Silicon probes are ideal recording electrodes to monitor multiple neurons because of a large number of recording sites (+64) and a small volume. Furthermore, multiple sites can be arranged over a distance of millimeters, thus allowing for the simultaneous recordings of neuronal activity in the various cortical layers or in multiple cortical columns (Fig. 1). Importantly, the geometrically precise distribution of the recording sites also allows for the determination of the spatial relationship of the isolated single neurons4. Here, we describe an acute, large-scale neuronal recording from the left and right forelimb somatosensory cortex simultaneously in an anesthetized rat with silicon probes (Fig. 2).

Extracellular recordings of neuronal activity using silicon probes in the anesthetized rat will be described. This technique allows information to be obtained across multiple brain areas from more than 100 neurons simultaneously. It provides information with single cell resolution about neuronal ensembles dynamics in multiple local circuits.

Large scale electrophysiological recordings from neuronal ensembles offer the opportunity to investigate how the brain orchestrates the wide variety of ...

Acute In Vivo Electrophysiological Recordings of Local Field Potentials and Multi-unit Activity from the Hyperdirect Pathway in Anesthetized Rats

Converging evidence shows that many neuropsychiatric diseases should be understood as disorders of large-scale neuronal networks. To better understand the pathophysiological basis of these diseases, it is necessary to precisely characterize in which way the processing of information is disturbed between the different neuronal parts of the circuit. Using extracellular in vivo electrophysiological recordings, it is possible to accurately delineate neuronal activity within a neuronal network. The application of this method has several advantages over alternative techniques, e.g., functional magnetic resonance imaging and calcium imaging, as it allows a unique temporal and spatial resolution and does not rely on genetically engineered organisms. However, the use of extracellular in vivo recordings is limited since it is an invasive technique that cannot be universally applied. In this article, a simple and easy to use method is presented with which it is possible to simultaneously record extracellular potentials such as local field potentials and multiunit activity at multiple sites of a network. It is detailed how a precise targeting of subcortical nuclei can be achieved using a combination of stereotactic surgery and online analysis of multi-unit recordings. Thus, it is demonstrated, how a complete network such as the hyperdirect cortico-basal ganglia loop can be studied in anesthetized animals in vivo.

Acute In Vivo Electrophysiological Recordings of Local Field Potentials and Multi-unit Activity from the Hyperdirect Pathway in Anesthetized Rats

In this study, the methodology is presented on how to perform multi-site in vivo electrophysiological recordings from the hyperdirect pathway under urethane anesthesia.

Converging evidence shows that many neuropsychiatric diseases should be understood as disorders of large-scale neuronal networks. To better understand the ...

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 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 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 ...

Construction and Implementation of Carbon Fiber Microelectrode Arrays for Chronic and Acute In Vivo Recordings

Multichannel electrode arrays offer insight into the working brain and serve to elucidate neural processes at the single-cell and circuit levels. Development of these tools is crucial for understanding complex behaviors and cognition and for advancing clinical applications. However, it remains a challenge to densely record from cell populations stably and continuously over long time periods. Many popular electrodes, such as tetrodes and silicon arrays, feature large cross-diameters that produce damage upon insertion and elicit chronic reactive tissue responses associated with neuronal death, hindering the recording of stable, continuous neural activity. In addition, most wire bundles exhibit broad spacing between channels, precluding simultaneous recording from a large number of cells clustered in a small area. The carbon fiber microelectrode arrays described in this protocol offer an accessible solution to these concerns. The study provides a detailed method for fabricating carbon fiber microelectrode arrays that can be used for both acute and chronic recordings in vivo. The physical properties of these electrodes make them ideal for stable and continuous long-term recordings at high cell densities, enabling the researcher to make robust, unambiguous recordings from single units across months.

Construction and Implementation of Carbon Fiber Microelectrode Arrays for Chronic and Acute In Vivo Recordings

This protocol describes a procedure for constructing carbon fiber microelectrode arrays for chronic and acute in vivo electrophysiological recordings in mouse (Mus musculus) and ferret (Mustela putorius furo) from multiple brain regions. Each step, following the purchase of raw carbon fibers to microelectrode array implantation, is described in detail, with emphasis on microelectrode array construction.

Multichannel electrode arrays offer insight into the working brain and serve to elucidate neural processes at the single-cell and circuit levels. ...

Extracellular Electrophysiological Recording in the Motor Cortex of Mice

Multichannel Extracellular Recording in Freely Moving Mice

The protocol aims to uncover the properties of neuronal firing and network local field potentials (LFPs) in behaving mice carrying out specific tasks by correlating the electrophysiological signals with spontaneous and/or specific behavior. This technique represents a valuable tool in studying the neuronal network activity underlying these behaviors. The article provides a detailed and complete procedure for electrode implantation and consequent extracellular recording in free-moving conscious mice. The study includes a detailed method for implanting the microelectrode arrays, capturing the LFP and neuronal spiking signals in the motor cortex (MC) using a multichannel system, and the subsequent offline data analysis. The advantage of multichannel recording in conscious animals is that a greater number of spiking neurons and neuronal subtypes can be obtained and compared, which allows the evaluation of the relationship between a specific behavior and the associated electrophysiological signals. Notably, the multichannel extracellular recording technique and the data analysis procedure described in the present study can be applied to other brain areas when conducting experiments in behaving mice.

Author Spotlight: Advancements in Multichannel Extracellular Recording for Studying Neuronal Activity in Freely Moving Mice 

The protocol describes the methodology of extracellular recording in the motor cortex (MC) to reveal extracellular electrophysiological properties in freely moving conscious mice, as well as the data analysis of local field potentials (LFPs) and spikes, which is useful for evaluating the network neural activity underlying behaviors of interest.

The protocol aims to uncover the properties of neuronal firing and network local field potentials (LFPs) in behaving mice carrying out specific tasks by ...

Open&#45;Source Implant for Multi&#45;Region Electrophysiological Monitoring in Rats

The TD Drive: A Parametric, Open-Source Implant for Multi&#45;Area Electrophysiological Recordings in Behaving and Sleeping Rats

Intricate interactions between multiple brain areas underlie most functions attributed to the brain. The process of learning, as well as the formation and consolidation of memories, are two examples that rely heavily on functional connectivity across the brain. In addition, investigating hemispheric similarities and/or differences goes hand in hand with these multi-area interactions. Electrophysiological studies trying to further elucidate these complex processes thus depend on recording brain activity at multiple locations simultaneously and often in a bilateral fashion. Presented here is a 3D-printable implant for rats, named TD Drive, capable of symmetric, bilateral wire electrode recordings, currently in up to ten distributed brain areas simultaneously. The open-source design was created employing parametric design principles, allowing prospective users to easily adapt the drive design to their needs by simply adjusting high-level parameters, such as anterior-posterior and mediolateral coordinates of the recording electrode locations. The implant design was validated in n = 20 Lister Hooded rats that performed different tasks. The implant was compatible with tethered sleep recordings and open field recordings (Object Exploration) as well as wireless recording in a large maze using two different commercial recording systems and headstages. Thus, presented here is the adaptable design and assembly of a new electrophysiological implant, facilitating fast preparation and implantation.

Author Spotlight: Deciphering Memory and Learning Through Neural Implants for Multi&#45;Region Brain Studies

Here, we present a unique, 3D-printable implant for rats, named TD Drive, capable of symmetric, bilateral wire electrode recordings, currently in up to ten distributed brain areas simultaneously.

Intricate interactions between multiple brain areas underlie most functions attributed to the brain. The process of learning, as well as the formation and ...

Flexible Multi-Region Neural Recording Device for Epilepsy Research

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

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 ...

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

Source: Haumesser, J. K., et al. <a href="https://app-jove-com.remotexs.ntu.edu.sg/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.
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Construction of Local Field Potential Microelectrodes for in vivo Recordings from Multiple Brain Structures Simultaneously

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