We are grateful for the assistance from Martina Lienert and Fabienne Schaufelberger during exercise testing, as well as from P. D. Claudio Perret, PhD for his scientific advice.

The study was approved by the local ethical committee (Ethikkommission Nordwest- und Zentralschweiz, Basel, Switzerland), and written informed consent was obtained from the participants before starting the study.
1. Test Preparation and Participant Instruction
<ol>
	<li>Arm crank ergometer
	<ol>
		<li>Turn the power on the rotation speed-dependent arm crank ergometer before opening the software.</li>
		<li>Choose the test protocol for the 3 min, all-out ergometer test.
		<ol>
			<li>Insert a new protocol with a 120 s warmup, 180 s of test duration, and a 720 s cooldown period. Choose this test protocol and open a new participant sheet.</li>
		</ol>
		</li>
		<li>For every new test, determine the participant&#39;s body mass beforehand.</li>
		<li>Set the relative torque factor to 0.2 for able-bodied and paraplegic individuals (e.g., for a 100-kg participant with a relative torque factor of 0.2, a torque of 20 Nm results)12.
		<ol>
			<li>Apply a lower torque factor for tetraplegic participants depending on the lesion level of the spinal cord injury; two or more familiarization trials are needed to determine the optimal relative torque factor for the relevant participant.</li>
			<li>Perform a familiarization trial in the same manner as described in section 2. If no peak appears after printing the data from the familiarization trial, or if the participant was not able to crank for the whole 3 min, perform a second familiarization trial with a lower torque factor. Give participants a rest of at least two days between each trial.</li>
		</ol>
		</li>
	</ol>
	</li>
	<li>Exercise test settings
	<ol>
		<li>Adjust the height of the arm crank and record it to replicate identical test settings in the next test session. Adjust and record the distance between the arm crank ergometer and the participant.
		<ol>
			<li>To determine the height, measure the distance between the floor and the fixation of the crank. To record the distance between the crank and the participant, measure and record the distance between the wall and the chair fixation. Adjust the arm crank axis to a height horizontal to the shoulder joint.</li>
		</ol>
		</li>
		<li>Record either the distance between the wall fixation and the chair or the distance between the ergometer and the chair fixation. Adjust the chair settings according to whether the participant is a) able-bodied, b) paraplegic ,or c) tetraplegic.
		<ol>
			<li>If the participant is able-bodied, have the participant sit in the chair provided by the distributor.</li>
			<li>If the participant is paraplegic and needs to sit in their own wheelchair, use a fixation set to fix the wheelchair to the arm crank ergometer. If the participant does not need their own wheelchair, have the participant sit in the chair provided by the distributor.</li>
			<li>If the participant is tetraplegic, fix their upper body to the chair provided by the distributor or to their own wheelchair and possibly fix their hands to the pedals. To fix the upper body, use a strap with hook-and-loop fastener. For hand fixation, use a wristband in tetraplegic patients.</li>
		</ol>
		</li>
	</ol>
	</li>
	<li>Additional measurements
	<ol>
		<li>Make sure that the lactate system solution is refilled before using the lactate analyzer. Insert a new chip sensor every six months. Use the quality control solution (12 mM) every day and the 3 mM quality control solution every two weeks.
		<ol>
			<li>Put the 12 mM quality control solution into the &#34;STD 1&#34; slot every morning.</li>
			<li>To further enhance the quality, add 3 mM quality control solutions into gap &#34;1&#34; and &#34;2&#34; and run a measurement by pressing &#34;start&#34; every two weeks. The measurement should result in a range between 2.96 and 3.10 mM.</li>
		</ol>
		</li>
		<li>To determine the whole-blood lactate concentration before and after the 3 min, all-out arm crank ergometer test, obtain a baseline lactate concentration. Disinfect the earlobe with a disinfectant before drawing a blood sample from the earlobe using a 10 &#181;L capillary. Use a lancet to get the whole blood sample.
		<ol>
			<li>When the capillary is completely full of blood, put it into the hemolysis cup. 
			NOTE: These cups are commercially available and prefilled with a hemolyzing solution. Shake the solution until the blood is completely mixed before putting it into the tray of the lactate analyzer.</li>
			<li>Run a calibration before analyzing the lactate concentration. Place the quality control cup into the lactate analyzer (see step 1.3.1.1.). Ensure that the calibration results in a 12-mM lactate concentration; otherwise, replace the chip sensor.</li>
			<li>Place the samples into the numbered slots, starting with &#34;1&#34; for the sample that was taken first. 
			NOTE: After the calibration is completed, the samples are measured automatically by the chip sensor system.</li>
		</ol>
		</li>
		<li>To determine the heartrate, place a heartrate belt around the chest of the participant and fix the heartrate monitor to the arm crank ergometer. Start the measurement by pressing on the red start button on the monitor. If no heartrate is displayed on the watch, wet the heartrate belt with water to ensure a good recording of the heartrate.</li>
		<li>To determine oxygen consumption during warmup and during the 3 min all-out test, calibrate the metabolic cart before the test. Run automatic volume and gas calibration immediately before the test and before putting the mask on.
		<ol>
			<li>Open the automatic volume calibration in the software and press the start button. Store the results if the error is below 3% on the screen.</li>
			<li>Open the gas calibration in the software, as well as the calibration gas, and start with the automatic calibration. 
			NOTE: The calibration gas consists of 5% CO2, 16% O2, and 79% N2. When 8 green buttons are displayed on the screen at the end of the calibration, the calibration is successful and the results can be stored. Close the gas bottle to ensure no leakage of gas.</li>
			<li>Ensure that the participant&#39;s actual body mass is inserted into the computer program. After the participant is chosen by the search engine on the computer, choose &#34;ergospirometry&#34; in the software and start with the measurement of room air concentration by pressing the start button.</li>
			<li>To run this calibration, take the sensor out of the spirometer and press the start button. Calibration is finished when &#34;ok&#34; is displayed on the computer.</li>
			<li>Meanwhile, during the calibration, put the oxygen mask on the participant.</li>
			<li>When the measurement of room air concentration is finished and the program is ready to measure, put the sensor back into the spirometer. Then, put the whole spirometer into the cavity of the mask; the device is now ready to measure oxygen consumption.</li>
			<li>In addition, fix the hose of the spirometer somewhere (e.g., on the shoulder with an adhesive tape) so that it does not interfere during the arm crank exercise.</li>
		</ol>
		</li>
	</ol>
	</li>
</ol>
2. Execution of the Exercise Protocol
<ol>
	<li>Warmup
	<ol>
		<li>1 min before starting the warmup, start measuring the oxygen consumption at rest when the participant sits at the arm crank ergometer without moving or talking. Press the start button in the software program.</li>
		<li>At the same time, start the measurement of the heartrate by pressing the red button. Measure the heartrate during the warmup, as well as during and after the test.</li>
		<li>Perform a standardized warmup over 2 min at 20 W before the start of the test. During the last 30 s of the warmup, keep the cadence constant at 60 rpm. Count down the last 10 s of the 30-s warmup.</li>
	</ol>
	</li>
	<li>3 min all-out exercise test
	<ol>
		<li>At the end of the countdown, make sure to give a clear starting signal by shouting &#34;go.&#34; After the start signal is given, allow the participant to accelerate.</li>
		<li>Instruct the participant to accelerate the arm crank ergometer to the maximum possible speed right at the beginning of the test. Keep the cadence at the maximum possible speed during the whole test. For standardization reasons, do not encourage the participants during the tests.</li>
		<li>Give information on duration every 30 s. Finish the test after a duration of 3 min.</li>
	</ol>
	</li>
	<li>Cooldown and post analysis
	<ol>
		<li>After finishing the 3 min all-out test, measure the end lactate concentration, if desired, and thereafter every 2 min for the next 10 min. Re-use the same puncture site for blood sampling as was used before the test.</li>
		<li>Stop the measurement of oxygen consumption after finishing these 3 min by pressing the stop button. Remove the oxygen mask. Save the measurement of oxygen consumption on the computer by pressing the exit button and by clicking &#34;yes&#34; when the software asks for data storage. 
		NOTE: The data is stored in the software program and can easily be converted to a csv document later.</li>
		<li>To export the data, press the &#34;Export&#34; button to convert the file to a csv document for later analysis. Stop the heartrate measurement by pressing the stop button on the left side of the heartrate monitor after all blood samples have been drawn from the earlobe.</li>
	</ol>
	</li>
</ol>
3. Data Analysis and Interpretation of the Results
<ol>
	<li>Performance parameters
	<ol>
		<li>Analyze several different parameters after finishing this performance test. 
		First, save the test and export it to a spreadsheet.</li>
		<li>Calculate the mean power (Pmean = <img alt="Equation" src="/files/ftp_upload/55485/55485eq1.jpg" /><img alt="Equation" src="/files/ftp_upload/55485/55485eq2.jpg" /> over 3 min, the peak power, and the minimal power in between these 3-min12. 
		NOTE: the peak power (Ppeak) is the maximal power during the whole 3 min. The power is measured in 0.2-s intervals. The peak power is the highest and the minimal power (Pmin) the lowest single-power measurement.</li>
		<li>Calculate the fatigue index as the power decline per second from the peak power to the end power ((Ppeak [W] &#8211; Pmin [W])/(tmin [s] &#8211; tpeak [s])).</li>
		<li>Calculate the total work over the whole 3 min by adding the work done every second (Work [J]= resistance [kg] * revolutions per min * flywheel distance [m] * time [min]).</li>
		<li>Calculate the time from the start to the peak power (time to peak power = tpeak [s]). Furthermore, calculate the relative peak (relative Ppeak = Ppeak/kg body mass) and the mean power (relative Pmean = Pmean/kg body mass) by dividing the absolute values by the body mass of the participant.</li>
		<li>Divide the 3-min all-out test into 30-s segments to check the pacing strategy and fatigue over these 3 min. Calculate the mean power for every 30-s segment (Pmean = <img alt="Equation" src="/files/ftp_upload/55485/55485eq3.jpg" /><img alt="Equation" src="/files/ftp_upload/55485/55485eq4.jpg" />.</li>
	</ol>
	</li>
	<li>Other measurements
	<ol>
		<li>Place all blood samples into the numbered slots of the blood lactate analyzer and run the measurements automatically by pressing &#34;analyze.&#34; Print blood lactate concentrations for later analysis by turning on the printer.</li>
		<li>Transmit the heartrate measurements to the computer using an infrared device from the manufacturer. Open the software of the heartrate monitor and import the data from the heartrate monitor to the software. Store the data locally and, if desired, export it to a spreadsheet for later analysis (e.g., segment analysis)13.</li>
		<li>Set a marker right at the beginning of the 3 min and at the end of the 3 min, allowing the average, maximal, and minimal heartrate to be calculated automatically for this segment. 
		NOTE: The heartrate is averaged automatically by the software over 5 s intervals.</li>
		<li>Export the data for oxygen consumption to a csv file (step 2.3) and open it in a spreadsheet for analysis14. Calculate the average oxygen consumption at rest: (VO2_rest = <img alt="Equation" src="/files/ftp_upload/55485/55485eq5.jpg" /><img alt="Equation" src="/files/ftp_upload/55485/55485eq6.jpg" /> and during the 3 min (VO2_180s = <img alt="Equation" src="/files/ftp_upload/55485/55485eq1.jpg" /><img alt="Equation" src="/files/ftp_upload/55485/55485eq7.jpg" />, as well as the peak oxygen consumption and oxygen consumption during the 30-s segments: (VO2_30s = <img alt="Equation" src="/files/ftp_upload/55485/55485eq3.jpg" /><img alt="Equation" src="/files/ftp_upload/55485/55485eq8.jpg" />. 
		NOTE: The data for oxygen consumption are measured breath-by-breath and then averaged automatically over a duration of 15 s per segment. The peak oxygen consumption is the highest value over a 15-s interval during the 3-min exercise test.</li>
	</ol>
	</li>
	<li>Statistics
	<ol>
		<li>Use the Shapiro-Wilk test, the Q-Q-plot, and the Kolmogorov Smirnov tests to check the normal distribution of the data. If the data is normally distributed, present it as the mean and the standard deviation (SD).</li>
		<li>Analyze the test-retest reliability using the intra-class correlation coefficient (ICC; 3,1 model)15.</li>
		<li>Calculate the absolute and relative reliability using the standard error of the measurement (SEM), coefficient of variation (CV), smallest real difference (SRD), and 95% confidence interval of the ICC16. 
		NOTE: The ICC should be interpreted according to Munro`s classification17: 0.26 to 0.49 reflects a low correlation; 0.50 to 0.69 reflects a moderate correlation; 0.70 to 0.89 reflects a high correlation; and 0.90 to 1.0 indicates a very high correlation. The absolute reliability should be presented as the SRD, CV, and SEM, and the relative reliability should be in the form of the ICC16,18.</li>
		<li>Analyze significant changes between the two test sessions using a paired t-test. To show the agreement of the data sets of both test sessions, use Bland-Altman19 plots. Use statistical software to perform data analysis; set a statistical significance level of 0.05 throughout.</li>
	</ol>
	</li>
</ol>

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The authors have nothing to disclose.

Exercise testing in spinal-cord injured athletes is crucial to tracking exercise performance over several months or years of training. Only a few exercise tests exist to check short-term, high-intensity exercise performance on the arm crank ergometer. This method describes in detail how an exercise test that was already examined for its reliability in cycling5 and rowing7 might be applied to the arm crank ergometer. To collect reliable and meaningful results, two factors ar...

There are several exercise tests that accurately determine the increase in exercise performance in elite athletes over the course of a training period1,2,3,4,5. One of these tests is the reliable 3-min all-out exercise test on a braked cycling ergometer3,4,5,6. This test was used to determine critical power, but it was also applied to exercise testing with athletes, as well as to research7,8,9. As this test was mainly used for lower-extremity performance, such as in rowing7 and cycling3,5, a similar testing protocol for upper-body exercise was needed. Sport disciplines that mainly use the upper body might be possible beneficiaries for such a new test protocol, in addition to athletes or individuals with an impairment of lower body muscles (e.g., an amputation or an impairment of limbs due to a spinal cord injury). Hence, a test protocol on the arm crank ergometer is a good tool to easily test upper-body exercise performance in a variety of athletes from different sport disciplines.
The existence of a very similar 30 s Wingate arm crank ergometer test10,11 helped with the development of a protocol for a 3 min, all-out arm crank ergometer test. Its duration is very similar to that of a 1,500 m wheelchair race. Therefore, this new test protocol of the 3 min, all-out arm crank ergometer test was tested for its test-retest reliability12. Overall, the reliability of this test protocol was excellent, so it could be a future testing tool in the field of upper-body exercise testing. Nevertheless, the use of this exercise test requires attention, especially when testing individuals with a spinal cord injury. Therefore, the aim of this experimental article is to demonstrate a detailed protocol that describes not only the test settings and analysis of the test results, but that also indicates the differences between testing able-bodied individuals and athletes with a spinal cord injury.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>Angio V2 arm crank ergometer</td>
			<td>Lode BV, Groningen, NL</td>
			<td>N/A</td>
			<td>arm crank ergometer</td>
		</tr>
		<tr>
			<td>Lode Ergometry Manager Software</td>
			<td>Lode BV, Groningen, NL</td>
			<td>N/A</td>
			<td>Software</td>
		</tr>
		<tr>
			<td>10ul end-to-end capillary</td>
			<td>EKF-diagnostics GmbH, Barleben, Germany</td>
			<td>0209-0100-005</td>
			<td>Capillaries</td>
		</tr>
		<tr>
			<td>haemolysis cup</td>
			<td>EKF-diagnostics GmbH, Barleben, Germany</td>
			<td>0209-0100-006</td>
			<td>hemolysis cup</td>
		</tr>
		<tr>
			<td>lactate analyzer</td>
			<td>Biosen C line, EKF-diagnostics GmbH</td>
			<td>5213-0051-6200</td>
			<td>lactate analyzer</td>
		</tr>
		<tr>
			<td>Heart rate monitor, Polar 610i</td>
			<td>Polar, Kempele, Finland</td>
			<td>P610i</td>
			<td>heart rate monitor</td>
		</tr>
		<tr>
			<td>metabolic cart, Oxygen Pro</td>
			<td>Jaeger GmbH</td>
			<td>N/A</td>
			<td>metabolic cart</td>
		</tr>
		<tr>
			<td>oxygen mask, Hans Rudolph</td>
			<td>Hans Rudolph Inc. , USA</td>
			<td>113814</td>
			<td>oxygen mask</td>
		</tr>
		<tr>
			<td>statistical software, PSAW Software</td>
			<td>SPSS Inc., Chicago USA</td>
			<td>N/A</td>
			<td>statistical software</td>
		</tr>
		<tr>
			<td>desinfectant, Soft-Zellin</td>
			<td>Hartmann GmbH, Austria</td>
			<td>999979</td>
			<td>desinfectant</td>
		</tr>
		<tr>
			<td>Quality control cup, EasyCon Norm</td>
			<td>EKF-diagnostics GmbH, Barleben, Germany</td>
			<td>0201-005.012P6</td>
			<td>quality control</td>
		</tr>
		<tr>
			<td>Quality control cup 3mmol/L</td>
			<td>EKF-diagnostics GmbH, Barleben, Germany</td>
			<td>5130-6152</td>
			<td>control cup</td>
		</tr>
		<tr>
			<td>Chip sensor lactate analyzer</td>
			<td>EKF-diagnostics GmbH, Barleben, Germany</td>
			<td>5206-3029</td>
			<td>chip sensor</td>
		</tr>
		<tr>
			<td>Lactate system solution</td>
			<td>EKF-diagnostics GmbH, Barleben, Germany</td>
			<td>0201-0002-025</td>
			<td>lactate system solution</td>
		</tr>
		<tr>
			<td>lancet, Mediware Blutlanzetten</td>
			<td>medilab</td>
			<td>54041</td>
			<td>lancet</td>
		</tr>
		<tr>
			<td>Calibration gas,&#160;</td>
			<td>Jaeger GmbH</td>
			<td>36-MC G020</td>
			<td>calibration gas</td>
		</tr>
		<tr>
			<td>chair provided by distributor (ergoselect)</td>
			<td>ergoline GmbH, Germany</td>
			<td>N/A</td>
			<td>chair provided by distributor</td>
		</tr>
	</tbody>
</table>

experimental protocol of a three-minute, all-out arm crank exercise test in spinal-cord injured and able-bodied individuals

Reliable exercise protocols are required to test changes in exercise performance in elite athletes. Performance improvements in these athletes may be small; therefore, sensitive tools are fundamental to exercise physiology. There are currently many exercise tests that allow for the examination of exercise capacity in able-bodied athletes, with protocols mainly for lower-body or whole-body exercise. There is a trend to test athletes in a sport-specific setting that closely resembles the actions that the participants are used to performing. Only a few protocols test short-term, high-intensity exercise capacity in participants with an impairment of the lower body. Most of these protocols are very sport-specific and are not applicable to a wide range of athletes. One well-known test protocol is the 30 s Wingate test, which is well-established in cycling and in arm crank exercise testing. This test analyzes high-intensity exercise performance over a 30 s time duration. In order to monitor exercise performance over a longer duration, a different method was modified for application to the upper body. The 3 min, all-out arm crank ergometer test allows athletes to be tested in a manner specific to 1,500 m wheelchair racing (in terms of exercise duration), as well as to upper body exercises such as rowing or hand-cycling. In order to increase the reliability with identical test conditions, it is crucial to precisely replicate settings such as the resistance (i.e., torque factor) and the position of the participants (i.e., the height of the crank, the distance between the crank and the participant, and the fixation of the participant). Another important issue concerns the beginning of the exercise test. Fixed revolutions per minute are required to standardize the test conditions for the start of the exercise test. This exercise protocol shows the importance of accurate operations to reproduce identical test conditions and settings.

The test-retest reliability was checked in 21 recreationally trained (but not specifically upper-body trained), non-smoking individuals (9 males, 12 females; age: 34 &#177; 11 years; body mass: 69.6 &#177; 11.1 kg; and height: 175.5 &#177; 6.9 cm). Table 1 shows the results for the relative and absolute test-retest reliabilities12. The peak power compared between the test and retest is presented in Figure 112</sup...

Watch this Scientific Journal Video about Experimental Protocol of a Three-minute, All-out Arm Crank Exercise Test in Spinal-cord Injured and Able-bodied Individuals at JoVE.com

Experimental Protocol of a Three-minute, All-out Arm Crank Exercise Test in Spinal-cord Injured and Able-bodied Individuals

We present a protocol to test the aerobic and anaerobic power of the upper body muscles over a duration of 3 min in able-bodied as well as in paraplegic and tetraplegic individuals. The protocol presents specific modifications in its application for upper-body exercise in individuals with or without disability.

Reliable exercise protocols are required to test changes in exercise performance in elite athletes. Performance improvements in these athletes may be ...

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9.6K Views.  Swiss Paraplegic Center Nottwil. We present a protocol to test the aerobic and anaerobic power of the upper body muscles over a duration of 3 min in able-bodied as well as in paraplegic and tetraplegic individuals. The protocol presents specific modifications in its application for upper-body exercise in individuals with or without disability. 

Video: Experimental Protocol of a Three-minute, All-out Arm Crank Exercise Test in Spinal-cord Injured and Able-bodied Individuals

Acute and Chronic Tactile Sensory Testing after Spinal Cord Injury in Rats

Spinal cord injury (SCI) impairs sensory systems causing allodynia1-8. To identify cellular and molecular causes of allodynia, sensitive and valid sensory testing in rat SCI models is needed. However, until recently, no single testing approach had been validated for SCI so that standardized methods have not been implemented across labs. Additionally, available testing methods could not be implemented acutely or when severe motor impairments existed, preventing studies of the development of SCI-induced allodynia3. Here we present two validated sensory testing methods using von Frey Hair (VFH) monofilaments which quantify changes in tactile sensory thresholds after SCI4-5. One test is the well-established Up-Down test which demonstrates high sensitivity and specificity across different SCI severities when tested chronically5. The other test is a newly-developed dorsal VFH test that can be applied acutely after SCI when allodynia develops, prior to motor recovery4-5. Each VFH monofilament applies a calibrated force when touched to the skin of the hind paw until it bends. In the up-down method, alternating VFHs of higher or lower forces are used on the plantar L5 dermatome to delineate flexor withdrawal thresholds. Successively higher forces are applied until withdrawal occurs then lower force VFHs are used until withdrawal ceases. The tactile threshold reflects the force required to elicit withdrawal in 50% of the stimuli. For the new test, each VFH is applied to the dorsal L5 dermatome of the paw while the rat is supported by the examiner. The VFH stimulation occurs in ascending order of force until at least 2 of 3 applications at a given force produces paw withdrawal. Tactile sensory threshold is the lowest force to elicit withdrawal 66% of the time. Acclimation, testing and scoring procedures are described. Aberrant trials that require a retest and typical trials are defined. Animal use was approved by Ohio State University Animal Care and Use Committee.

We describe two tactile sensory testing methods for acute or chronic periods of spinal cord injury in rats. These validated procedures can detect the development and maintenance of allodynia-like sensations.

Spinal cord injury (SCI) impairs sensory systems causing allodynia1-8. To identify cellular and molecular causes of allodynia, sensitive and valid sensory ...

Evaluation of Respiratory Muscle Activation Using Respiratory Motor Control Assessment (RMCA) in Individuals with Chronic Spinal Cord Injury

During breathing, activation of respiratory muscles is coordinated by integrated input from the brain, brainstem, and spinal cord. When this coordination is disrupted by spinal cord injury (SCI), control of respiratory muscles innervated below the injury level is compromised1,2 leading to respiratory muscle dysfunction and pulmonary complications. These conditions are among the leading causes of death in patients with SCI3. Standard pulmonary function tests that assess respiratory motor function include spirometrical and maximum airway pressure outcomes: Forced Vital Capacity (FVC), Forced Expiratory Volume in one second (FEV1), Maximal Inspiratory Pressure (PImax) and Maximal Expiratory Pressure (PEmax)4,5. These values provide indirect measurements of respiratory muscle performance6. In clinical practice and research, a surface electromyography (sEMG) recorded from respiratory muscles can be used to assess respiratory motor function and help to diagnose neuromuscular pathology. However, variability in the sEMG amplitude inhibits efforts to develop objective and direct measures of respiratory motor function6. Based on a multi-muscle sEMG approach to characterize motor control of limb muscles7, known as the voluntary response index (VRI)8, we developed an analytical tool to characterize respiratory motor control directly from sEMG data recorded from multiple respiratory muscles during the voluntary respiratory tasks. We have termed this the Respiratory Motor Control Assessment (RMCA)9. This vector analysis method quantifies the amount and distribution of activity across muscles and presents it in the form of an index that relates the degree to which sEMG output within a test-subject resembles that from a group of healthy (non-injured) controls. The resulting index value has been shown to have high face validity, sensitivity and specificity9-11. We showed previously9 that the RMCA outcomes significantly correlate with levels of SCI and pulmonary function measures. We are presenting here the method to quantitatively compare post-spinal cord injury respiratory multi-muscle activation patterns to those of healthy individuals.

Evaluation of Respiratory Muscle Activation Using Respiratory Motor Control Assessment RMCA in Individuals with Chronic Spinal Cord Injury

The purpose of this publication is to present our original work on a multi-muscle surface electromyographic approach to quantitatively characterize respiratory muscle activation patterns in individuals with chronic spinal cord injury using vector-based analysis.

During breathing, activation of respiratory muscles is coordinated by integrated input from the brain, brainstem, and spinal cord. When this coordination ...

Implanting Glass Spinal Cord Windows in Adult Mice with Experimental Autoimmune Encephalomyelitis

Experimental autoimmune encephalomyelitis (EAE) in adult rodents is the standard experimental model for studying autonomic demyelinating diseases such as multiple sclerosis. Here we present a low-cost and reproducible glass window implantation protocol that is suitable for intravital microscopy and studying the dynamics of spinal cord cytoarchitecture with subcellular resolution in live adult mice with EAE. Briefly, we surgically expose the vertebrae T12-L2 and construct a chamber around the exposed vertebrae using a combination of cyanoacrylate and dental cement. A laminectomy is performed from T13 to L1, and a thin layer of transparent silicone elastomer is applied to the dorsal surface of the exposed spinal cord. A modified glass cover slip is implanted over the exposed spinal cord taking care that the glass does not directly contact the spinal cord. To reduce the infiltration of inflammatory cells between the window and spinal cord, anti-inflammatory treatment is administered every 2 days (as recommended by ethics committee) for the first 10 days after implantation. EAE is induced only 2-3 weeks after the cessation of anti-inflammatory treatment.
Using this approach we successfully induced EAE in 87% of animals with implanted windows and, using Thy1-CFP-23 mice (blue axons in dorsal spinal cord), quantified axonal loss throughout EAE progression. Taken together, this protocol may be useful for studying the recruitment of various cell populations as well as their interaction dynamics, with subcellular resolution and for extended periods of time. This intravital imaging modality represents a valuable tool for developing therapeutic strategies to treat autoimmune demyelinating diseases such as EAE.

We describe a method for implanting and maintaining glass windows over the exposed spinal cords of adult mice for studying experimental autoimmune encephalomyelitis. These windows provide chronic optical access to the spinal cord for monitoring cell dynamics at the subcellular level in live animals using two-photon microscopy.

Experimental autoimmune encephalomyelitis (EAE) in adult rodents is the standard experimental model for studying autonomic demyelinating diseases such as ...

Controlled Cortical Impact Model for Traumatic Brain Injury

Every year over a million Americans suffer a traumatic brain injury (TBI). Combined with the incidence of TBIs worldwide, the physical, emotional, social, and economical effects are staggering. Therefore, further research into the effects of TBI and effective treatments is necessary. The controlled cortical impact (CCI) model induces traumatic brain injuries ranging from mild to severe. This method uses a rigid impactor to deliver mechanical energy to an intact dura exposed following a craniectomy. Impact is made under precise parameters at a set velocity to achieve a pre-determined deformation depth. Although other TBI models, such as weight drop and fluid percussion, exist, CCI is more accurate, easier to control, and most importantly, produces traumatic brain injuries similar to those seen in humans. However, no TBI model is currently able to reproduce pathological changes identical to those seen in human patients. The CCI model allows investigation into the short-term and long-term effects of TBI, such as neuronal death, memory deficits, and cerebral edema, as well as potential therapeutic treatments for TBI.

Traumatic brain injuries (TBIs) remain a serious health problem. Using the controlled cortical impact surgery model, research on the effects of TBI and possible treatment methods may be performed.

Every year over a million Americans suffer a traumatic brain injury (TBI).  Combined with the incidence of TBIs worldwide, the physical, emotional, ...

A Radio-telemetric System to Monitor Cardiovascular Function in Rats with Spinal Cord Transection and Embryonic Neural Stem Cell Grafts

High thoracic or cervical spinal cord injury (SCI) can lead to cardiovascular dysfunction. To monitor cardiovascular parameters, we implanted a catheter connected to a radio transmitter into the femoral artery of rats that underwent a T4 spinal cord transection with or without grafting of embryonic brainstem-derived neural stem cells expressing green fluorescent protein. Compared to other methods such as cannula insertion or tail-cuff, telemetry is advantageous to continuously monitor blood pressure and heart rate in freely moving animals. It is also capable of long term multiple data acquisitions. In spinal cord injured rats, basal cardiovascular data under unrestrained condition and autonomic dysreflexia in response to colorectal distension were successfully recorded. In addition, cardiovascular parameters before and after SCI can be compared in the same rat if a transmitter is implanted before a spinal cord transection. One limitation of the described telemetry procedure is that implantation in the femoral artery may influence the blood supply to the ipsilateral hindlimb.

We present a protocol for using a radio-telemetric system to record cardiovascular parameters in T4 spinal cord transected rats eight weeks after embryonic brainstem neural stem cell grafting into the lesion site. Telemetry is an advanced technique to accurately evaluate cardiovascular function in conscious freely moving spinal cord injured rats.

High thoracic or cervical spinal cord injury (SCI) can lead to cardiovascular dysfunction. To monitor cardiovascular parameters, we implanted a catheter ...

Synergetic Use of Neural Precursor Cells and Self-assembling Peptides in Experimental Cervical Spinal Cord Injury

Spinal cord injuries (SCI) cause serious neurological impairment and psychological, economic, and social consequences for patients and their families. Clinically, more than 50% of SCI affect the cervical spine1. As a consequence of the primary injury, a cascade of secondary mechanisms including inflammation, apoptosis, and demyelination occur finally leading to tissue scarring and development of intramedullary cavities2,3. Both represent physical and chemical barriers to cell transplantation, integration, and regeneration. Therefore, shaping the inhibitory environment and bridging cavities to create a supportive milieu for cell transplantation and regeneration is a promising therapeutic target4. Here, a contusion/compression model of cervical SCI using an aneurysm clip is described. This model is more clinically relevant than other experimental models, since complete transection or ruptures of the cord are rare. Also in comparison to the weight drop model, which in particular damage the dorsum columns, circumferential compression of the spinal cord appears advantageous. Clip closing force and duration can be adjusted to achieve different injury severity. A ring spring facilitates precise calibration and constancy of clip force. Under physiological conditions, synthetic self-assembling peptides (SAP) self-assemble into nanofibers and thus, are appealing for application in SCI5. They can be injected directly into the lesion minimizing damage to the cord. SAPs are biocompatible structures erecting scaffolds to bridge intramedullary cavities and thus, equip the damaged cord for regenerative treatments. K2(QL)6K2 (QL6) is a novel SAP introduced by Dong et al.6 In comparison to other peptides, QL6 self-assembles into &#946;-sheets at neutral pH6.14 days after SCI, after the acute stage, SAPs are injected into the center of the lesion and neural precursor cells (NPC) are injected into adjacent dorsal columns. In order to support cell survival, transplantation is combined with continuous subdural administration of growth factors by osmotic micro pumps for 7 days.

Treating cervical spinal cord injury with both self-assembling peptides (SAP) and neural precursor cells (NPC), together with growth factors, is a promising approach to promote regeneration and recovery. A contusion/compression aneurysm clip rat model of cervical SCI and combined treatment involving SAP injection and NPC transplantation is established.

Spinal cord injuries (SCI) cause serious neurological impairment and psychological, economic, and social consequences for patients and their families. ...

Neural Stem Cell Transplantation in Experimental Contusive Model of Spinal Cord Injury

Spinal cord injury is a devastating clinical condition, characterized by a complex of neurological dysfunctions. Animal models of spinal cord injury can be used both to investigate the biological responses to injury and to test potential therapies. Contusion or compression injury delivered to the surgically exposed spinal cord are the most widely used models of the pathology. In this report the experimental contusion is performed by using the Infinite Horizon (IH) Impactor device, which allows the creation of a reproducible injury animal model through definition of specific injury parameters. Stem cell transplantation is commonly considered a potentially useful strategy for curing this debilitating condition. Numerous studies have evaluated the effects of transplanting a variety of stem cells. Here we demonstrate an adapted method for spinal cord injury followed by tail vein injection of cells in CD1 mice. In short, we provide procedures for: i) cell labeling with a vital tracer, ii) pre-operative care of mice, iii) execution of a contusive spinal cord injury, and iv) intravenous administration of post mortem neural precursors. This contusion model can be utilized to evaluate the efficacy and safety of stem cell transplantation in a regenerative medicine approach.

Spinal cord injury is a traumatic condition that causes severe morbidity and high mortality. In this work we describe in detail a contusion model of spinal cord injury in mice followed by a transplantation of neural stem cells.

Spinal cord injury is a devastating clinical condition, characterized by a complex of neurological dysfunctions. Animal models of spinal cord injury can ...

Contrast Enhanced Ultrasound Imaging for Assessment of Spinal Cord Blood Flow in Experimental Spinal Cord Injury

Reduced spinal cord blood flow (SCBF) (i.e., ischemia) plays a key role in traumatic spinal cord injury (SCI) pathophysiology and is accordingly an important target for neuroprotective therapies. Although several techniques have been described to assess SCBF, they all have significant limitations. To overcome the latter, we propose the use of real-time contrast enhanced ultrasound imaging (CEU). Here we describe the application of this technique in a rat contusion model of SCI. A jugular catheter is first implanted for the repeated injection of contrast agent, a sodium chloride solution of sulphur hexafluoride encapsulated microbubbles. The spine is then stabilized with a custom-made 3D-frame and the spinal cord dura mater is exposed by a laminectomy at ThIX-ThXII. The ultrasound probe is then positioned at the posterior aspect of the dura mater (coated with ultrasound gel). To assess baseline SCBF, a single intravenous injection (400 &#181;l) of contrast agent is applied to record its passage through the intact spinal cord microvasculature. A weight-drop device is subsequently used to generate a reproducible experimental contusion model of SCI. Contrast agent is re-injected 15 min following the injury to assess post-SCI SCBF changes. CEU allows for real time and in-vivo assessment of SCBF changes following SCI. In the uninjured animal, ultrasound imaging showed uneven blood flow along the intact spinal cord. Furthermore, 15 min post-SCI, there was critical ischemia at the level of the epicenter while SCBF remained preserved in the more remote intact areas. In the regions adjacent to the epicenter (both rostral and caudal), SCBF was significantly reduced. This corresponds to the previously described &#8220;ischemic penumbra zone&#8221;. This tool is of major interest for assessing the effects of therapies aimed at limiting ischemia and the resulting tissue necrosis subsequent to SCI.

Contrast Enhanced Ultrasound imaging is a reliable in-vivo tool for quantifying spinal cord blood flow in an experimental rat spinal cord injury model. This paper contains a comprehensive protocol for application of this technique in association with a contusion model of thoracic spinal cord injury.

Reduced spinal cord blood flow (SCBF) (i.e., ischemia) plays a key role in traumatic spinal cord injury (SCI) pathophysiology and is accordingly an ...

Determining The Electromyographic Fatigue Threshold Following a Single Visit Exercise Test

Theoretically, the electromyographic (EMG) fatigue threshold is the exercise intensity an individual can maintain indefinitely without the need to recruit more motor units which is associated with an increase in the EMG amplitude. Although different protocols have been used to estimate the EMG fatigue threshold they require multiple visits which are impractical for a clinical setting. Here, we present a protocol for estimating the EMG fatigue threshold for cycle ergometry which requires a single visit. This protocol is simple, convenient, and completed within 15-20 min, therefore, has the potential to be translated into a tool that clinicians can use in exercise prescription.

This protocol describes the electromyographic fatigue threshold which demarcates between nonfatiguing and fatiguing exercise workloads. This information could be used to develop a more individualized training program.

Theoretically, the electromyographic (EMG) fatigue threshold is the exercise intensity an individual can maintain indefinitely without the need to recruit ...

A Neonatal Mouse Spinal Cord Compression Injury Model

Spinal cord injury (SCI) typically causes devastating neurological deficits, particularly through damage to fibers descending from the brain to the spinal cord. A major current area of research is focused on the mechanisms of adaptive plasticity that underlie spontaneous or induced functional recovery following SCI. Spontaneous functional recovery is reported to be greater early in life, raising interesting questions about how adaptive plasticity changes as the spinal cord develops. To facilitate investigation of this dynamic, we have developed a SCI model in the neonatal mouse. The model has relevance for pediatric SCI, which is too little studied. Because neural plasticity in the adult involves some of the same mechanisms as neural plasticity in early life1, this model may potentially have some relevance also for adult SCI. Here we describe the entire procedure for generating a reproducible spinal cord compression (SCC) injury in the neonatal mouse as early as postnatal (P) day 1. SCC is achieved by performing a laminectomy at a given spinal level (here described at thoracic levels 9-11) and then using a modified Yasargil aneurysm mini-clip to rapidly compress and decompress the spinal cord. As previously described, the injured neonatal mice can be tested for behavioral deficits or sacrificed for ex vivo physiological analysis of synaptic connectivity using electrophysiological and high-throughput optical recording techniques1. Earlier and ongoing studies using behavioral and physiological assessment have demonstrated a dramatic, acute impairment of hindlimb motility followed by a complete functional recovery within 2 weeks, and the first evidence of changes in functional circuitry at the level of identified descending synaptic connections1.

This article describes a method for generating a reproducible spinal cord compression injury (SCI) in the neonatal mouse. The model provides an advantageous platform for studying mechanisms of adaptive plasticity that underlie spontaneous functional recovery.

Spinal cord injury (SCI) typically causes devastating neurological deficits, particularly through damage to fibers descending from the brain to the spinal ...