Ten able-bodied (6 females, 4 males; 24.3 ± 2.58 years old) and one SCI subject (male; 15 years old; T-10 level of injury; ASIA A; 8 years post injury) with no history of lower extremity orthopedic disorders voluntarily participated in this study. All 10 able-bodied subjects participated in the main experiments, which evaluated the effects of stimulation frequency and fatigue on the human quadriceps muscle force-intensity relationship. The SCI subject was recruited for the case study. Able-bodied subjects were recruited from the general population of students at the University of Delaware and individuals in the surrounding community; the SCI subject was recruited at Shriners Hospital for Children in Philadelphia. Each subject was made aware of the nature of the research, the procedures and the potential risks involved. All able-bodied subjects signed an informed consent that was approved by the Human Subjects Review Board of University of Delaware. The subject with SCI and his parent signed an informed consent and an assent form that was approved both by the Human Subjects Review Board of University of Delaware and the Institutional Review Board of Temple University.
The experimental setup was the same for the able-bodied subjects and the subject with SCI. Subjects were seated on a computer-controlled dynamometer (KinCom III 500–11 (able-bodied); KinCom II (SCI), Chattecx Corp., Chattanooga, TN) with their hips flexed to approximately 85° and knees flexed to 90°. The axis of the dynamometer was aligned with the axis of the subject’s knee joint. The trunk, waist and thigh were stabilized using inelastic straps with Velcro closures. Isometric quadriceps muscle forces were measured by a force transducer placed at the anterior aspect of the tibia, with the lower edge of the transducer pad positioned 2.5 cm proximal to the lateral malleolus. The right quadriceps muscle was stimulated using a voltage regulated Grass S8800 stimulator with an SIU8T stimulus isolation unit (Grass Instrument Co., Quincy, MA). A personal computer equipped with a PCI-6Q24FDAQ board, a PCI6602 counter-timer board, and custom written LabVIEW software (National Instruments, Austin, TX) was used to control the timing of all stimulation pulses during testing. A custom-made pulse duration control switch was connected in series with the Grass stimulator for the modulation of pulse duration. For the able-bodied subjects, two, 7.6 × 12.7-cm, self-adhesive electrodes (Versa-stim, Conmed Corporation, Utica, NY) were used to deliver electrical stimulation. For the subjects with SCI, two, 7.5 × 10-cm, self-adhesive electrodes (Pals Platinum, Axelgaard Manufacturing Co., Ltd, Fallbrook, CA) were used. The electrode connected to the anode of the stimulator was placed over the motor point of rectus femoris and the cathode was placed over the motor point of vastus medialis (Barnett et al., 1991
). The force data were sampled at a rate of 200 Hz and stored on the computer’s hard disc.
Experimental procedure for able-bodied subjects
Each able-bodied subject participated in one testing session. Subjects were asked to refrain from strenuous exercise 24 hours before the testing session. At the beginning of the testing session, each subject’s maximal voluntary isometric contraction (MVIC) was determined using the burst superimposition technique (Snyder-Mackler et al., 1993
). The burst superimposition technique used an 11-pulse (600 μs pulse duration, 135 V), 100-Hz train to stimulate the muscle while the subjects performed a maximum voluntary contraction of their quadriceps muscles. The MVIC was accepted if the volitional contraction was ≥ 95 % of the “superimposed” tetanic force. If the peak force of volitional contraction was < 95% of the superimposed tetanic force, subjects rested for 5 minutes before attempting to perform another MVIC testing. All subjects were able to perform successful MVICs within 3 attempts. Subjects rested for 5 minutes after MVIC testing.
Quadriceps muscles were potentiated before setting the stimulation amplitude. One, 12-pulse (600 μs pulse duration), 14-Hz train was delivered to the quadriceps muscles every 5 seconds over a ~ 60 second period to potentiate the muscles (240 total pulses) (Binder-Macleod et al., 2002
). The stimulation intensity was then set using 60-Hz (600 μs pulse duration), 300 ms long trains. The stimulation amplitude was increased gradually until the muscle peak force responses reached 20 % of the subject’s MVIC. Previous work showed that if rather than using a 300-ms train, the train duration was extended to ~ 1 second, this stimulation intensity would produce ~ 40 % of subject’s MVIC (Ding et al., 2003
). Thus, the current intensity was recruiting approximately 40 % of the quadriceps femoris muscle. The amplitude was then held constant throughout the remainder of the session. Although stimulation intensity can be modulated by varying either pulse amplitude or duration, pulse duration modulation was chosen in the present study as it was easier to control and may require less charge per stimulus pulse compared to stimulation amplitude modulation (Crago et al., 1974
Testing trains were delivered right after setting the stimulation intensity. Each subject received a testing protocol consisting of a pre-fatigue portion, a fatiguing portion and a post-fatigue portion. For the pre-fatigue portion, a series of 300-ms long testing trains with different frequency (10, 12.5, 20, 30, 40 and 60 Hz) and pulse duration (100, 200, 300, 400 and 600 μs) combinations were delivered to the quadriceps muscles to examine the force responses to different stimulation frequencies and intensities before muscle fatigue. These testing trains were delivered in a random order at the rate of 1 train every 10 seconds to avoid fatigue, and then repeated in a reversed order (N.B., the same random order was used for all subjects). The force responses to the same train were averaged for later analysis. Next, a modified Burke’s fatigue protocol (Burke et al., 1973
) consisting of a series of 40-Hz, 300-ms trains, with pulse duration at 600 μs, was delivered to the muscle at a rate of one train every second for a total of 180 trains to fatigue the quadriceps muscles. The post-fatigue portion was delivered to subject’s quadriceps muscle immediately after the fatigue portion. Stimulation in this portion was continued at a rate of 1 train per second. Post-fatigue testing included the same sequence of testing trains as used in the pre-fatigue portion, but each of the testing trains was separated by two fatiguing trains (e.g., 40-Hz, 300-ms train with pulse duration at 600 μs) to maintain a steady state of fatigue. The force responses to the same train were averaged for later analysis.
A simple test was performed to determine if fatigue was produced during the pre-fatigue portion of testing. Because the first and last stimulation trains (20 Hz; 300 μs pulse duration) of the pre-fatigue protocol were identical (see above), the peak forces produced by the first and last trains were compared. Analysis of these data using paired t-test showed no significant difference (first trains’ peak force=99.5±11.3 N; last trains’ peak force=100.2±10.7 N; P=0.72). Similarly, a comparison of the peak forces produced by the first and last testing trains of the post-fatigue protocol showed no significant difference (first trains’ peak force=28.19±3.5 N; last trains’ peak force=26.88±4.1 N; P=0.31). Thus, we concluded that there was no fatigue produced during the pre-fatigue portion and a steady level of fatigue was maintained during the post-fatigue portion.
Experimental procedure for the subject with SCI
Quadriceps femoris muscle’s maximal twitch force (MTF) was first determined using single pulses (600 μs pulse duration). Stimulation amplitude was gradually increased until a plateau in the MTF was reached. Next, the stimulation voltage was adjusted to produce tetanic force equal to MTF level using a 1-s, 100-Hz stimulation train (pulse duration of 600 μs). The MTF of paralyzed skeletal muscles are about 15 to 25 % of the maximal tetanic force (Gerrits et al., 1999
; Scott et al., 2006
). Next, a series of 1-sec long stimulation trains with different frequency (12.5, 20, 33.3, 50 and 80 Hz) and intensity (150, 250, 350 and 600 μs) combinations were delivered to the quadriceps femoris muscles. These 20 testing trains were delivered in a random order at the rate of one train every 10 seconds to avoid fatigue, and then repeated in a reversed order. The force responses to the train with the same frequency and intensity were averaged and analyzed to obtain each subject’s force-intensity relationship and intensity modulation steps (Details in the next section). The stimulation amplitude was readjusted using a series of 60-Hz (600-μs pulse duration), 300-ms long trains. The stimulation amplitude was gradually increased until the muscle force responses reached subject’s 2×MTF. The stimulation amplitude was then held constant for the remainder of the session. Quadriceps muscle was then stimulated repetitively at the rate of one train every 1.1 second using a series of 300-ms long, 30-Hz trains with a pulse duration (226 μs) that produced a peak force equal to the subject’s MTF. To precisely maintain muscle force output, the stimulation pulse duration was increased in a stepwise manner every time the peak forces declined more than 10% from the targeted force level (i.e., MTF) due to muscle fatigue. The experimental protocol was terminated when muscle peak forces could not be maintained above 90% MTF at the maximum pulse duration (600 μs).
The pulse duration modulation steps were calculated based on the force-intensity relationship of the subject. Five calculation steps were needed to determine each pulse duration step ():
Fig. 1 Example for the determination of stimulation pulse duration modulation steps based on the force-intensity relationship curve for a typical subject. (1) represents the initial force-intensity relationship for the subject. (2) Identifying the starting pulse (more ...)
Step 1: The force-intensity relationship curve (black curve) obtained for the subject was fitted with equation 1
, and the parameter values for A
and τ for the force-intensity relationship curve were then determined.
Step 2: The starting pulse duration was identified by locating the pulse duration that produced peak forces equal to the targeted force level (dark point). In the case study, stimulation trains were delivered once every second; therefore, muscle peak force output gradually declined due to muscle fatigue.
Step 3: We predetermined that the stimulation pulse duration would be modulated when the peak force drops 10% from the targeted force level (gray point).
Step 4: When the peak force dropped 10% from the targeted force level, a new curve that represented the relationship between force output and stimulation intensity at that point of time could then be identified. Based on the results of the study with able-bodied subjects (See “Results” section), only parameter A changed with fatigue. Thus, with all other parameter values known (determined from Step 1), the new A value for the new force-intensity relationship curve (gray curve) could then be determined.
Step 5: By locating the pulse duration that produced peak forces equal to the targeted force level from the new force-intensity relationship curve, the next modulation step for pulse duration could be identified.
By repeating these steps, a family of the force-intensity relationship curves for the subject was identified (). As expected, the force-intensity relationship scaled down as the muscle fatigues. All the pulse duration modulation steps for the subject were determined by locating the intersections between the curves and the targeted force level. Calculation was stopped when the longest pulse duration (600 μs) was reached.
Fig. 2 Family of force-intensity relationship curves for the subject with SCI. The force-intensity relationship shifted down with fatigue. The intensity modulation steps (arrows) were determined by locating the intersections between the force-intensity curves (more ...)
Data management and analysis
To analyze the force-intensity relationship, the peak force responses to each of the testing trains were measured and plotted as a function of pulse duration for each frequency tested for both pre- and post-fatigue conditions. Each force-intensity relationship curve was normalized to the peak force produced at the longest pulse duration (600 μs).
An exponential equation,
was used to fit each of the force-intensity relationship curves, where parameter A is the scaling factor for the force (F), PD represents the duration of the stimulation pulse (μs). PD0 represents the threshold pulse duration (μs), above which we predict there is a measurable force. τ is the time constant controlling the rise of the force with increasing pulse duration. Because muscles cannot generate negative force, F will equal to zero when PD ≤ PD0. R2 value between the actual force-intensity relationship curves and the fitting curves was calculated to evaluate the quality of the fit. Two-way (conditions versus frequencies) analyses of variance (ANOVAs) with repeated measures were used to determine if the parameter values for A, PD0, or τ changed with frequency or fatigue. Pairwise comparison with a Bonferroni correction was performed if a significant main effect was observed. Statistical significance was accepted at P ≤ 0.05.