The authors would like to acknowledge Dr. Claire Warriner for contributing to the development of this method. Mark Agrios and Sajishnu Savya assisted with preparing figures. This research was supported by a Searle Scholar Award, a Sloan Research Fellowship, a Simons Collaboration on the Global Brain Pilot Award, a Whitehall Research Grant Award, The Chicago Biomedical Consortium with support from the Searle Funds at The Chicago Community Trust, NIH grant DP2 NS120847 (A.M.), and NIH grant 2T32MH067564 (A.K.).

Fabrication and Surgical Implantation of Electromyographic Electrodes in Mice

<ol>
	<li>Dombeck, D. A., Khabbaz, A. N., Collman, F., Adelman, T. L., Tank, D. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Imaging+large+scale+neural+activity+with+cellular+resolution+in+awake+mobile+mice.">Imaging large scale neural activity with cellular resolution in awake mobile mice.</a> Neuron. 56 (1), 43-57 (2007).</li><li>Guo, Z. V., 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=Flow+of+cortical+activity+underlying+a+tactile+decision+in+mice.">Flow of cortical activity underlying a tactile decision in mice.</a> Neuron. 81 (1), 179-194 (2014).</li><li>Guo, J. Z., 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=Cortex+commands+the+performance+of+skilled+movement.">Cortex commands the performance of skilled movement.</a> eLife. 4, e10774 (2015).</li><li>Morandell, K., Huber, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+role+of+forelimb+motor+cortex+areas+in+goal+directed+action+in+mice.">The role of forelimb motor cortex areas in goal directed action in mice.</a> Sci Rep. 7 (1), 15759 (2017).</li><li>Gali&#241;anes, G. L., Bonardi, C., Huber, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Directional+reaching+for+water+as+a+cortex-dependent+behavioral+framework+for+mice.">Directional reaching for water as a cortex-dependent behavioral framework for mice.</a> Cell Rep. 22 (10), 2767-2783 (2018).</li><li>Evarts, E. 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R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+role+of+cutaneous+afferents+in+the+control+of+gamma-motoneurones+during+locomotion+in+the+decerebrate+cat.">The role of cutaneous afferents in the control of gamma-motoneurones during locomotion in the decerebrate cat.</a> J Physiol. 434, 529-547 (1991).</li><li>Manuel, M., Chardon, M., Tysseling, V., Heckman, C. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Scaling+of+motor+output,+from+mouse+to+humans.">Scaling of motor output, from mouse to humans.</a> Physiol Bethesda Md. 34 (1), 5-13 (2019).</li><li>Sauerbrei, B. A., 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=Cortical+pattern+generation+during+dexterous+movement+is+input-driven.">Cortical pattern generation during dexterous movement is input-driven.</a> Nature. 577 (7790), 386-391 (2020).</li><li>Barrett, J. M., Raineri Tapies, M. G., Shepherd, G. M. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Manual+dexterity+of+mice+during+food-handling+involves+the+thumb+and+a+set+of+fast+basic+movements.">Manual dexterity of mice during food-handling involves the thumb and a set of fast basic movements.</a> PLoS One. 15 (1), e0226774 (2020).</li><li>Serradj, N., 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=Task-specific+modulation+of+corticospinal+neuron+activity+during+motor+learning+in+mice.">Task-specific modulation of corticospinal neuron activity during motor learning in mice.</a> Nat Commun. 14, 2708 (2023).</li><li>Scholle, H. 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G., Acharya, H., Fouad, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+new+electrode+configuration+for+recording+electromyographic+activity+in+behaving+mice.">A new electrode configuration for recording electromyographic activity in behaving mice.</a> J Neurosci Methods. 148 (1), 36-42 (2005).</li><li>Akay, T., Acharya, H. J., Fouad, K., Pearson, K. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Behavioral+and+electromyographic+characterization+of+mice+lacking+EphA4+receptors.">Behavioral and electromyographic characterization of mice lacking EphA4 receptors.</a> J Neurophysiol. 96 (2), 642-651 (2006).</li><li>Akay, T., Tourtellotte, W. G., Arber, S., Jessell, T. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Degradation+of+mouse+locomotor+pattern+in+the+absence+of+proprioceptive+sensory+feedback.">Degradation of mouse locomotor pattern in the absence of proprioceptive sensory feedback.</a> Proc Natl Acad Sci USA. 111 (47), 16877-16882 (2014).</li><li>Miri, A., 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=Behaviorally+selective+engagement+of+short-latency+effector+pathways+by+motor+cortex.">Behaviorally selective engagement of short-latency effector pathways by motor cortex.</a> Neuron. 95 (3), 683-696 (2017).</li><li>Osborne, J., Dudman, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=RIVETS:+A+mechanical+system+for+in+vivo+and+in+vitro+electrophysiology+and+imaging.">RIVETS: A mechanical system for in vivo and in vitro electrophysiology and imaging.</a> PloS One. 9 (2), e89007 (2014).</li><li>Santuz, A., Laflamme, O. 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This protocol enables stable muscle activity recordings from head-fixed mice performing a variety of behaviors for several weeks. Recently, this method has been employed to examine neural control of limb musculature during behaviors such as treadmill locomotion18,20, a joystick pulling task18, and a co-contraction task21.&#160;While the protocol described here is specific for mouse elbow and wrist muscles, it is easily modified to record from different muscles or a different number of muscles by changing the length and/or total number of electrode pairs. The method described here was adapted from those used previously to record forelimb and hindlimb muscle activity in mice without head restraint15,16,17.
Electrode fabrication requires significant practice to master. Daily practice for 1-2 h is recommended while learning. Stripping the electrodes is the most challenging step due to the precise level of force required to cut the insulation without damaging the underlying wire. This level of force is dependent on the sharpness of the blade, so frequently replacing the scalpel blade can help ensure reproducibility during learning. Soldering the wires to the brass blades of the connector can also be difficult because stainless steel does not readily solder. Applying a liberal amount of stainless steel-compatible flux&#160;helps promote the connection.
The main challenge during the implantation surgery is tying the distal knot without disturbing the implanted wire or proximal knot. The proximal knot must be large enough to resist slipping into muscle at the insertion site - thus, avoid tying the knot too tight in step 2 of electrode set fabrication. If the proximal knot migrates after implantation, use carbon fiber-tipped forceps to reposition it carefully. Tighten the distal knot slowly while maintaining a firm grip on the wire with forceps to avoid pulling the entire electrode through. This step is critical to ensure the longevity of implanted electrodes: too much tension placed on the electrode can cause it to break when the animal moves, while a loose electrode can shift during recovery and lose contact with its associated muscle as the tissue heals.
Animals recover remarkably well from the surgery, though there are potential complications to note. First, mice will chew on their sutures and electrodes if given the chance. While the Elizabethan collar prevents this, it also prevents the animal from grooming itself. Some mice develop a mucus-like build-up around their eyes. Occasional male mice, particularly older ones, experience urethra blockages that can be distressing to the animal. Allowing the animal to groom itself for 20 min each day before inspecting sutures should give the animal enough time to prevent these issues.
There are important limitations of this method to note. First, these custom electrodes generally cannot resolve single motor unit activity. Furthermore, the electrical signal is not guaranteed to emanate exclusively from a specific muscle (i.e., biceps), as it is difficult to rule out crosstalk from activity in nearby synergist muscles. Therefore, in publications, researchers commonly refer to the recorded muscles by their synergy group (i.e., elbow flexor). It is recommended to perform post-mortem dissections after each experiment to verify the position of each electrode, as they could shift in the tissue during recovery.
Researchers interested in single motor unit activity should consider trying newly developed EMG electrodes by the Center for Advanced Motor Bioengineering Research (CAMBER) at Emory University. These electrodes are still being developed, but CAMBER will provide the latest electrode design. The main drawback of these electrodes is longevity: the hand-fabricated electrodes described in this protocol generally allow recordings for several weeks, whereas CAMBER electrodes work best for short-term experiments. Researchers selecting an EMG recording method can contact CAMBER directly to determine if their electrodes will be suitable for a given experiment.

In recent decades, mice have become an attractive model organism for studying the mammalian motor system. Common experimental approaches involve head-fixed mice performing motor tasks alongside the monitoring and/or perturbation of neural activity1,2,3,4,5. Motor system studies in larger species (such as cats and primates) have traditionally relied upon electromyography (EMG) to measure motor output directly during such experiments6,7,8. However, recording muscle activity in mice is challenging because their musculature is too small for commercially available EMG electrodes used in large mammal experiments9. Many researchers opt to track limb kinematics through video4,10,11 and/or behavioral performance2,4,12 to probe motor output indirectly, but these methods lack the resolution to detect the millisecond-timescale influence of neural activity and perturbation thereof on muscles. Thus, recording EMG is desirable for researchers interested in direct neural control of muscles.
EMG involves measuring the voltage between two points, typically separated by a short distance roughly parallel to the fibers of the muscle being recorded. EMG electrodes come in surface (or &#34;patch&#34;) and intramuscular (or &#34;needle&#34;) varieties. Surface electrodes are placed atop the skin or overlaid on the muscle tissue and secured with adhesive or suturing. As such, surface electrodes are less invasive than intramuscular electrodes and are most popular with humans, cats, and primates due to their relative ease of use. Surface electrodes have also been used successfully with rats and mice13,14; however, they must be hand-fabricated and surgically implanted under the skin due to the tendency of rodents to try to remove foreign objects while grooming. Intramuscular EMG electrodes, on the other hand, are surgically implanted within the muscle tissue. Because they are engulfed in muscle tissue, they provide high spatial resolution and remain fixed in position indefinitely. Thus, implanted intramuscular EMG electrodes are ideal over surface electrodes for long-term experiments using rodents. To record intramuscular EMG reliably in mice, researchers have developed a method for hand fabricating and implanting EMG electrodes in muscles as small as those in an adult mouse&#39;s forearm. These electrodes enable chronic muscle recording during motor behavior in rodents over several weeks.
The protocol described here is the result of a decade-long refinement of established methods15,16,17,18, which has yielded a procedure for hand fabricating, implanting, and recording from wire EMG electrodes chronically implanted in flexor/extensor muscle pairs of the elbow and wrist in behaving mice. The first section describes the hand fabrication of an electrode set with four electrode pairs and an 8-pin connector&#160;for the head stage interface. The next section details the surgical implantation of the electrodes intramuscularly in upper and lower arm muscles in the same surgery as headplate implantation. Finally, representative recordings from mice performing a variety of behaviors are discussed. Overall, this method is a cost-effective and customizable way to include muscle activity measurements in head-fixed behavior experiments that is ideal for labs with some electrode fabrication experience.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>#11 Scalpel Blades</td>
			<td>World Precision Instruments</td>
			<td>504170</td>
			<td>For EMG electrode fabrication</td>
		</tr>
		<tr>
			<td>#3 Scalpel Handle</td>
			<td>Fine Science Tools</td>
			<td>10003-12</td>
			<td>For EMG electrode fabrication</td>
		</tr>
		<tr>
			<td>1 mL Sub-Q Syringe (100 pack)&#160;</td>
			<td>Becton Dickinson</td>
			<td>309597</td>
			<td>For administering injectable drugs</td>
		</tr>
		<tr>
			<td>12-pin connector</td>
			<td>Newark</td>
			<td>33AC2371</td>
			<td>12-pin connector with brass fittings; for EMG electrode fabrication</td>
		</tr>
		<tr>
			<td>18 G Needles</td>
			<td>Exel International</td>
			<td>26419</td>
			<td>For EMG electrode fabrication</td>
		</tr>
		<tr>
			<td>27 G Needles&#160;</td>
			<td>Exel International</td>
			<td>26426</td>
			<td>For EMG electrode fabrication</td>
		</tr>
		<tr>
			<td>3 M Transpore Surgical Tape</td>
			<td>3M</td>
			<td>1527-0</td>
			<td>For taping animal&#39;s limbs out during surgery</td>
		</tr>
		<tr>
			<td>6-0 silk sutures</td>
			<td>Henry Schein</td>
			<td>101-2636</td>
			<td>These sutures work well with delicate skin around the wrists&#160;</td>
		</tr>
		<tr>
			<td>C&#38;B Metabond Complete Kit</td>
			<td>Pearson Dental</td>
			<td>P16-0126</td>
			<td>Dental cement to affix connector to headplate</td>
		</tr>
		<tr>
			<td>C57BL6/J Mice&#160;</td>
			<td>Jackson Laboratories</td>
			<td>#000664</td>
			<td>Wild type mice&#160;</td>
		</tr>
		<tr>
			<td>Carbofib 5-CF Tweezers (2)</td>
			<td>Aven tools&#160;</td>
			<td>18762</td>
			<td>Carbon fiber tipped forceps, used to manipulate delicate parts of electrode (stripped or inserted sections)&#160;</td>
		</tr>
		<tr>
			<td>Carprodyl (Carprofen) 50 mg/mL Injection</td>
			<td>Ceva Animal Health, LLC</td>
			<td>G43010B</td>
			<td>Injectable analgesic for pain management during and after surgery</td>
		</tr>
		<tr>
			<td>Castroviejo Micro Needle Holder</td>
			<td>Fine science tools</td>
			<td>12060-01</td>
			<td>For suturing</td>
		</tr>
		<tr>
			<td>Castroviejo Needle Holder (large)</td>
			<td>Fine science tools</td>
			<td>12565-14</td>
			<td>For inserting needle into muscle&#160;</td>
		</tr>
		<tr>
			<td>Delicate Bone Scraper</td>
			<td>Fine science tools</td>
			<td>10075-16</td>
			<td>To separate skin from underlying tissue&#160;</td>
		</tr>
		<tr>
			<td>Dietgel 76A Dietary Supplement</td>
			<td>Clear H2O</td>
			<td>72-07-5022</td>
			<td>For post-operative care</td>
		</tr>
		<tr>
			<td>Dumont #5/45 Forceps</td>
			<td>Fine science tools</td>
			<td>11251-35</td>
			<td>To remove fascia overlying muscle&#160;</td>
		</tr>
		<tr>
			<td>Elizabethan collar for mouse</td>
			<td>Kent Scientific Corporation</td>
			<td>EC201V-10</td>
			<td>For post-operative care</td>
		</tr>
		<tr>
			<td>Enrofloxacin 2.27%</td>
			<td>Covetrus</td>
			<td>#074743</td>
			<td>Injectable antibiotic for use during and after surgery</td>
		</tr>
		<tr>
			<td>Epoxy gel</td>
			<td>Devcon</td>
			<td>14265</td>
			<td>For EMG electrode fabrication</td>
		</tr>
		<tr>
			<td>Hopkins Bulldog Clamp (4)</td>
			<td>Stoelting</td>
			<td>10-000-481</td>
			<td>Tissue clamps for headplate implantation</td>
		</tr>
		<tr>
			<td>Isoflurane Solution</td>
			<td>Covetrus</td>
			<td>11695067771</td>
			<td>Inhalable anesthesia</td>
		</tr>
		<tr>
			<td>Lidocaine Hydrochloride Injectable - 2%</td>
			<td>Covetrus</td>
			<td>#002468</td>
			<td>Topical analgesic for pain management during surgery</td>
		</tr>
		<tr>
			<td>Medical Grade Oxygen</td>
			<td>Airgas</td>
			<td>OX USP200</td>
			<td>For administering isoflurane during surgery</td>
		</tr>
		<tr>
			<td>MetriCide 1 Gallon</td>
			<td>Metrex</td>
			<td>10-1400</td>
			<td>Glutaraldehyde solution for cold-sterilization of headplate and electrodes&#160;</td>
		</tr>
		<tr>
			<td>MetriTest Strips 1.5%</td>
			<td>Metrex</td>
			<td>10-303</td>
			<td>Test strips for monitoring glutaraldehyde solution (recommended)</td>
		</tr>
		<tr>
			<td>Model 900LS Small Animal Stereotaxic Instrument</td>
			<td>Kopf Instruments&#160;</td>
			<td>900LS</td>
			<td>Stereotax with lazy susan feature that allows platform rotation during surgery&#160;</td>
		</tr>
		<tr>
			<td>PFA-coated 0.0055&#34; braided stainless steel wire&#160;</td>
			<td>A-M systems&#160;</td>
			<td>793200</td>
			<td>For EMG electrode fabrication</td>
		</tr>
		<tr>
			<td>Povidone-iodine prep pads</td>
			<td>Dynarex</td>
			<td>1108</td>
			<td>For cleaning skin&#160;</td>
		</tr>
		<tr>
			<td>Puralube Vet Ointment&#160;</td>
			<td>Dechra</td>
			<td>37327</td>
			<td>Eye ointment for surgery&#160;</td>
		</tr>
		<tr>
			<td>Sterile alcohol prep pads</td>
			<td>Dynarex</td>
			<td>1113</td>
			<td>For cleaning skin&#160;</td>
		</tr>
		<tr>
			<td>Straight fine #5 forceps</td>
			<td>Fine science tools</td>
			<td>11295-10</td>
			<td>For curling wire after insertion&#160;</td>
		</tr>
		<tr>
			<td>Straight fine scissors</td>
			<td>Fine science tools</td>
			<td>14060-11</td>
			<td>For cutting wire&#160;</td>
		</tr>
		<tr>
			<td>Student Vannas Spring Scissors</td>
			<td>Fine science tools</td>
			<td>91500-09</td>
			<td>For making incisions, trimming fat and fascia, and suturing&#160;</td>
		</tr>
		<tr>
			<td>Technik Tweezers 7B-SA (2)</td>
			<td>Aven tools</td>
			<td>18074USA</td>
			<td>Curved blunt forceps, for general use during surgery</td>
		</tr>
		<tr>
			<td>Triple Antibiotic Ointment</td>
			<td>Walgreens</td>
			<td>975863</td>
			<td>Topical antibiotic for surgery</td>
		</tr>
		<tr>
			<td>V-1 Tabletop Laboratory Animal Anesthesia System</td>
			<td>VetEquip</td>
			<td>901806</td>
			<td>Contains all necessary equipment for anesthesia induction and scavenging including vaporizer, induction chamber, moveable plastic nose cone, and tubing&#160;</td>
		</tr>
	</tbody>
</table>

recording forelimb muscle activity in head&#45;fixed mice with chronically implanted emg electrodes

Powerful genetic and molecular tools available in mouse systems neuroscience research have enabled researchers to interrogate motor system function with unprecedented precision in head-fixed mice performing a variety of tasks. The small size of the mouse makes the measurement of motor output difficult, as the traditional method of electromyographic (EMG) recording of muscle activity was designed for larger animals like cats and primates. Pending commercially available EMG electrodes for mice, the current gold-standard method for recording muscle activity in mice is to make electrode sets in-house. This article describes a refinement of established procedures for hand fabrication of an electrode set, implantation of electrodes in the same surgery as headplate implantation, fixation of a connector on the headplate, and post-operative recovery care. Following recovery, millisecond-resolution EMG recordings can be obtained during head-fixed behavior for several weeks without noticeable changes in signal quality. These recordings enable precise measurement of forelimb muscle activity alongside in vivo neural recording and/or perturbation to probe mechanisms of motor control in mice.

Author Spotlight: Investigating Mouse Motor Cortex Interactions from Muscle Activity to Neural Dynamics

Recording Forelimb Muscle Activity in Head&#45;Fixed Mice with Chronically Implanted EMG Electrodes

I'm interested in how parts of mouse motor cortex, called primary and secondary motor cortex, interact with each other to influence muscle activity during different kinds of movements. So we know that both primary and secondary motor cortex project to the spinal cord, but it's unclear how they differentially influence movement. So I'm using this protocol to compare the effects of inactivating primary and secondary motor cortex on forelimb muscle activity throughout movement to understand how they differentially control movement.The use of forelimb muscle EMG has enabled us to measure the impact of optogenetic inactivation of different motor system regions on motor output. For example, we've been able to show that direct motor cortical influence on muscles is specific to certain motor behaviors and particular muscle activity states during those behaviors. New large-scale multi-electrode arrays like Neuropixels now enable us to record from large neuronal populations across multiple motor system regions simultaneously.Performing EMG recordings during these neural recordings will enable us to characterize how the interactions between these motor system regions depend on muscle activity state.

Figure 2, Figure 3, and Figure 4 show normalized muscle activity recorded from the forelimb muscles of mice performing different behaviors: treadmill walking without head-fixation (Figure 2), climbing a rotating wheel under head-fixation (Figure 3), and reaching for water droplets under head-fixation (Figure 4). Figure 2 shows 1.5 s of treadmill locomotion with an approximate step cycle estimated from the time between two elbow flexor activations. Figure 3 shows 5 s of EMG data from an animal that had the wrist extensor electrode fail 6 weeks after implantation. In Figure 3A, all four electrodes produce a clean EMG signal that aligns with the turning of the wheel (which indicates climbing). Figure 3B shows the signal from the same electrodes after failure: the wrist extensor electrode produces a noisy signal that does not change with the animal&#39;s movement. Figure 4 shows 1 s of EMG from the four forelimb muscle groups during a task in which the mouse transitioned from immobility to reaching for a water droplet.
In Figure 2, Figure 3, and Figure 4, the voltage signals were amplified and bandpass filtered (250-20,000 Hz) using a differential amplifier. Raw voltage was then subsampled to 1 kHz and z-scored for comparison across datasets. Note again that while electrodes were implanted in the four muscles specified in the protocol (biceps, triceps, ECR, and PL), it is not guaranteed that adjacent synergistic muscles did not influence EMG signal; therefore, each recording is assigned to its synergy group (elbow flexor, etc.) for accuracy. Verifying isolated recordings from single muscles would require simultaneous recordings in multiple synergists to assay for crosstalk between muscle recordings, which may be prohibitively difficult, especially in the lower arm of mice.
<img alt="Figure 1" class="xfigimg" src="/files/ftp_upload/66584/66584fig01.jpg" /> 
Figure 1: Schematics of the electrode set fabrication. (A) Diagram of a single electrode pair. Gray areas indicate where to strip. (B) Diagram of the connector assembly with a single completed electrode pair inserted into the connector. The diagram in (B) is not to scale. <a href="https://www.jove.com/files/ftp_upload/66584/66584fig01large.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/66584/66584fig02.jpg" /> 
Figure 2: Representative EMG recording from four muscles of a freely moving (not head-fixed) mouse walking on a treadmill. The total duration is 1.5 s. The step cycle was estimated from the time between sequential elbow extensor activations. <a href="https://www.jove.com/files/ftp_upload/66584/66584fig02large.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/66584/66584fig03.jpg" /> 
Figure 3: Representative EMG recording from four muscles of a head-fixed mouse performing a naturalistic climbing behavior. The 5th row shows the position of the climbing wheel read out by a rotary encoder; changes in this value indicate that the wheel is turning and the animal is actively climbing. The total duration is 5 s. (A) Recording 36 days after implantation during climbing. (B) Recording 72 days after implantation in the same mouse after the wrist extensor electrode failed. <a href="https://www.jove.com/files/ftp_upload/66584/66584fig03large.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/66584/66584fig04.jpg" /> 
Figure 4: Representative EMG recording from four muscles of a head-fixed mouse transitioning from immobility to performing a reaching movement. The total duration is 1 s. <a href="https://www.jove.com/files/ftp_upload/66584/66584fig04large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>

Read this Scientific Article Protocol about Author Spotlight: Investigating Mouse Motor Cortex Interactions from Muscle Activity to Neural Dynamics at JoVE.com

This protocol describes the hand fabrication and surgical implantation of electromyographic (EMG) electrodes in the forelimb muscles of mice to record muscle activity during head-fixed behavior experiments.

Powerful genetic and molecular tools available in mouse systems neuroscience research have enabled researchers to interrogate motor system function with ...

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1.1K Views.  Northwestern University. I'm interested in how parts of mouse motor cortex, called primary and secondary motor cortex, interact with each other to influence muscle activity during different kinds of movements. So we know that both primary and secondary motor cortex project to the spinal cord, but it's unclear how they differentially influence movement. So I'm using this protocol to compare the effects of inactivating primary and secondary motor cortex on forelimb muscle activity throughout movement to understand how ...

Video: Author Spotlight: Investigating Mouse Motor Cortex Interactions from Muscle Activity to Neural Dynamics

Fabrication of an Electromyographic Set of Electrodes for Implantation in Murine Muscles

To begin the fabrication of the electrode set, first tie two pieces of PFA coated braided stainless steel wires together with a single knot. Insert an 18 gauge needle into a piece of corrugated cardboard. Position the knot six centimeters from the needles insertion end.Then pull the electrode strands against the cardboard to tighten it around the needle. Carefully remove the needle to maintain the knot's integrity, and pull the knot tight two times to tighten it further. Next, tape the electrode taut against a piece of flat cardboard.Ensure that the two connector end wires are together, and the insertion end wires are spread apart. With a scalpel, make precise nicks in the insulation, marking the ends where the insulation will be removed. Make a series of approximately six cuts at each marked location with two cuts each on top, at the sides, and underneath the wire.Angle the scalpel blade to cut along the loosened insulation length wise, and use a pair of forceps to carefully remove it from the wire. Strip one millimeter insulation off the end of each wire at the connector end, and five millimeters off at the insertion end. Twist the wire segments at the insertion end together, and crimp the exposed ends in the shaft of a 0.5 inch 27 gauge needle.Next, use a pair of straight forceps to carefully reinsert each soldered brass fitting back into the connector. Make sure that the wires of each electrode pair are adjacent and untangled. Coat the pin slots with epoxy to insulate all metal or wire near the connector from the tissue.

Electrode Implantation Surgery to Record Forelimb Muscle Activity in Head&#45;Fixed Mice

To begin, extend the incision on the back of the neck of an anesthetized mouse with a head plate implantation. With a blunt bone scraper, separate the skin under the neck incision from the underlying tissue. Work the scraper under the skin and behind the neck incision to clear a path for the electrodes.Insert the tip of a large needle driver through the triceps incision and out of the neck incision. Clamp the needle driver around the electrode needle lengthwise, then pull it through to the triceps incision. Next, bend a 27 gauge needle to give it a slight curve.Hold the needle with the needle driver. Then press it against the handle of a pair of forceps to result in an additional bend of five to 10 degrees. Add three total bends at different positions along the length of the needle.Next, identify the entry and exit points on the muscle. With a pair of fine forceps, remove any fat in fascia, obscuring the entry and exit site. After submerging three to five millimeters of wire parallel to the muscle fibers, use the needle driver to insert the needle into the proximal end of the muscle.Now push the needle through the muscle to the exit site. Once the needle exits the muscle, grab the tip with blunt forceps and pull the needle through. With a pair of forceps, tie a loose distal knot at the exit site.Tighten the knot down to a one centimeter loop. Push the loop with the forceps and position it over the exit site. After identifying the location over which the distal knot is to be closed, gently grasp the loop with a pair of fine bent forceps and pull the loop tight over the forceps.Remove the fine forceps from the knot. Push the knot toward the exit site with the fine bent forceps, and pull the needle end with fingers to finish tightening it. Use the straight fine forceps to grasp the exit knot, then tightly curl the distal wire around the forceps to bend the wire around the knot and toward the muscle.Finally, cut the wire off about 0.5 millimeters distal to the distal knot, leaving a small nub curled around the knot. All the implanted electrodes produced a clean electromyographic signal 36 days after implantation. However, after 72 days, the wrist extensor electrode failed.