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Combined effects of fatigue and temperature manipulation on skeletal muscle electrical and mechanical characteristics during isometric contraction
Journal of Electromyography and Kinesiology 22 (2012) 348–355
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Combined effects of fatigue and temperature manipulation on skeletal muscle electrical and mechanical characteristics during isometric contraction Emiliano Cè a,b,⇑, Susanna Rampichini a, Luca Agnello b, Eloisa Limonta a, Arsenio Veicsteinas a,b, Fabio Esposito a,c a b c
Department of Sport, Nutrition and Health Sciences, University of Milan, Via G. Colombo 71, 20133 Milan, Italy Center of Sport Medicine, Don Gnocchi Foundation, Via Capecelatro 66, 20148 Milan, Italy Centro Sicurezza nelle Attività Sportive e Corretti Stili di Vita (SAS), University of Milan, Via G. Colombo 71, 20133 Milan, Italy
a r t i c l e
i n f o
Article history: Received 19 December 2011 Received in revised form 19 January 2012 Accepted 19 January 2012
Keywords: EMG Isometric contraction Muscle temperature Neuromuscular efficiency Conduction velocity
a b s t r a c t Peripheral fatigue and muscle cooling induce similar effects on sarcolemmal propagation properties. The aim of the study was to assess the combined effects of muscle temperature (Tm) manipulation and fatigue on skeletal muscle electrical and mechanical characteristics during isometric contraction. After maximum voluntary contraction (MVC) assessment, 16 participants performed brief and sustained isometric tasks of different intensities in low (TmL), high (TmH) and neutral (TmN) temperature conditions, before and after a fatiguing exercise (6 s on/4 s off at 50% MVC, to the point of fatigue). During contraction, the surface electromyogram (EMG) and force were recorded from the biceps brachii muscle. The root mean square (RMS) and conduction velocity (CV) were calculated off-line. After the fatiguing exercise: (i) MVC decreased similarly in all Tm conditions (P < 0.05), while EMG RMS did not change; and (ii) CV decreased to a further extent in TmL compared to TmN and TmH in all brief and sustained contractions (P < 0.05). The larger CV drop in TmL after fatigue suggests that TmL and fatigue have a combined and additional effect on sarcolemmal propagation properties. Despite these changes, force generating capacity was not affected by Tm manipulation. A compensatory mechanism has been proposed to explain this phenomenon. Ó 2012 Elsevier Ltd. All rights reserved.
1. Introduction Temperature is an important determinant of skeletal muscle contractile and metabolic properties (Gray et al., 2006; Rall and Woledge, 1990; Ranatunga, 1998). Several investigations have dealt with the role of local temperature on skeletal muscle performance, yet its effects on neuromuscular activation still remain poorly understood. While some studies did not retrieve any significant alteration in neuromuscular function due to changes in muscle temperature (Tm) (Mitchell et al., 2008; Mito et al., 2007), the manipulation of Tm has been found to modify significantly neuromuscular activation during both voluntary (Cheung and Sleivert, 2004; de Ruiter et al., 1999; Dewhurst et al., 2010; Holewijn and Heus, 1992; Petrofsky and Laymon, 2005; Petrofsky and Lind, 1980; Racinais and Oksa, 2010; Ranatunga et al., 1987) and electrically-evoked contractions (Bigland-Ritchie et al., 1992; de Ruiter et al., 1999; Hopf and Maurer, 1990; Racinais and Oksa, 2010; Ranatunga et al., ⇑ Corresponding author. Address: Department of Sport, Nutrition and Health Sciences, Division of Human Physiology, University of Milan, Via G. Colombo 71, 20133 Milan, Italy. Tel.: +39 02 5031 4644; fax: +39 02 5031 4630. E-mail address: [email protected] (E. Cè). 1050-6411/$ - see front matter Ó 2012 Elsevier Ltd. All rights reserved. doi:10.1016/j.jelekin.2012.01.012
1987). In particular, despite no changes have been reported with high Tm (TmH) (Cheung and Sleivert, 2004; Dewhurst et al., 2010; Holewijn and Heus, 1992; Petrofsky and Laymon, 2005), a significant decrease in force generating capacity (Bigland-Ritchie et al., 1992; Cheung and Sleivert, 2004; de Ruiter et al., 1999; Holewijn and Heus, 1992; Hopf and Maurer, 1990; Petrofsky and Laymon, 2005; Ranatunga et al., 1987), in surface electromyogram (EMG) frequency content (Dewhurst et al., 2010; Holewijn and Heus, 1992; Petrofsky and Lind, 1980; Ranatunga et al., 1987) and in mean fiber conduction velocity (CV) (Merletti et al., 1984; Mucke and Heuer, 1989) have been previously described with low Tm (TmL). In TmL, indeed, a change in the action potential architecture and a decrease in its propagation velocity occur (Hicks and McComas, 1989; Hodgkin and Huxley, 1952) together with a reduction in Ca2+ release from the sarcoplasmic reticulum (Caputo, 1972a,b). This leads to a decline in adenosine triphosphate availability and cross-bridge cycling rate (Godt and Lindley, 1982). Also muscle fatigue, which is generally defined as a temporary loss in force generating capacity due to previous muscle contractions (Fitts, 1994), shows similar effects on the neuromuscular activation to those induced by TmL. Fatigue, indeed, decreases
E. Cè et al. / Journal of Electromyography and Kinesiology 22 (2012) 348–355
the motor unit action potential amplitude, its propagation across the sarcolemma, the Ca2+ efflux from sarcoplasmic reticulum and the cross-bridge cycling rate (Allen et al., 1989, 1995; Fitts, 2008). Surface EMG detection by linear electrode arrays is a non-invasive and reliable tool to provide information on muscle electrical activation (Merletti et al., 2003). The alterations in sarcolemmal propagation properties, action potential duration and muscle activation strategies can be evaluated indirectly by EMG time and frequency domain analysis and CV calculation (Farina et al., 2005; Gray et al., 2006; Merletti et al., 2003). While the separate effects of muscle fatigue and TmL alone have been already investigated during isometric contraction (Merletti et al., 1984, 2003; Mucke and Heuer, 1989), to the best of our knowledge, no studies have examined the effect of cooling on the electrical activation and force generating capacity of a previously fatigued muscle. TmL and fatigue, indeed, may have a combined and additional effect on the sarcolemmal propagation properties and force production. Thus, the aim of the present study was to assess the effects of Tm manipulation on the neuromuscular activation characteristics and maximum voluntary contraction (MVC) of a previously fatigued muscle. We hypothesize that, given the partly shared consequences of cooling and fatigue on muscle characteristics, the changes in the fatigued muscle would be more pronounced in TmL than in TmH or in thermo-neutral condition (TmN). To this purpose, we assessed the maximum force production of the elbow flexors and the electrical activation of the biceps brachii muscle at different contraction intensities and durations, before and after a fatiguing exercise. 2. Methods 2.1. Participants After a full explanation of the purpose of the study and experimental procedures, 16 male participants gave their written informed consent to be enrolled in the study. Their physical and anthropometric characteristics are given in Table 1. Participants were all clinically healthy, with no previous history of upper limbs, shoulder joint and muscle injuries. At the time of the study, none of them was involved in any specific training program. The study was approved by the local University Ethics Committee and performed in accordance with the principles of the 1975 Declaration of Helsinki. 2.2. Experimental design Before the experimental sessions, participants were allowed to familiarize with the ergometer and to test their ability to reach and maintain different contraction levels. Then, participants reported to the laboratory six times on different days, with at least 72 h of rest in between, during which they were invited to avoid fatiguing efforts involving the muscles of the upper limbs. The first three sessions were devoted to assess test–retest reliability of force and EMG parameters in TmN. In the other three sessions, the effects of TmN, TmH and TmL were evaluated in random order. All experiments were carried out in a laboratory at a constant temperature of 22 ± 1 °C and relative humidity of 50 ± 5%.
Table 1 Physical and anthropometric characteristics of the participants (n = 16). Data are expressed as mean ± SD. Age (yrs) Body mass (kg) Stature (m) Arm circumference (cm) Biceps brachii skin fold (mm)
23 ± 6 72.5 ± 5.8 1.73 ± 0.04 31.3 ± 2.4 5.1 ± 1.9
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2.3. Reliability assessment As previously described, the first three sessions were devoted to assess the reliability of the measurements. The maximum voluntary contraction (MVC) of each subject was the highest value of three maximum efforts of the elbow flexor muscles of the dominant arm. Then, contractions at 20%, 40%, 60%, 80% and 100% MVC of 5 s each were carried out in a random order. During contractions, the EMG signal was detected from the biceps brachii muscle. From the two central seconds of each contraction, the mean value of force, EMG root mean square (RMS) and CV were calculated. Before the first session, a map with some skin identification points (moles, angiomas and scars) and the positioning of the EMG probe was drawn on a transparency, to allow repeated measurements from the same muscle area during all tests. Each participant was positioned on an ergometer with the hand kept in a position midway between pronation and supination at an elbow angle of 115°. The whole detection apparatus had a resonant frequency >200 Hz. The skin area under the electrodes was carefully cleaned with ethyl alcohol and gently abraded with fine sand paper. A conductive gel was also applied to achieve an inter-electrode impedance below 2000 X. Surface EMG signal was detected by a 16-channel linear array, which included 16 silver bar electrodes (diameter 1 mm, length 5 mm, inter-electrode distance 10 mm) for differential EMG detection. The array was placed along muscle fibers direction, with the EMG electrodes positioned perpendicularly to the major axes of the fibers, in accordance with the European recommendations about surface EMG (Hermens et al., 2000). The force output was detected by a calibrated load cell (mod SM-1000 N, Interface, Crowthorne, UK), operating linearly between 0 and 1000 N. EMG and force signals were amplified and filtered (mod. ASE16, LiSin, Turin, Italy; gain 1000 and 50 for EMG and force, respectively; bandwidth of 10–500 Hz and 2– 20 Hz for EMG and force, respectively), converted to digital data by a 12-bit acquisition board (mod. DAQcard-6024E, National Instruments, Austin, USA; sample rate of 2048 Hz) and stored on a personal computer for further analysis. Contraction intensity was maintained within ±3% of the requested force target with the aid of a visual feedback (both target and force output shown on a computer screen). 2.4. Temperature effects assessment The assessment of the effects of temperature manipulation on neuromuscular activation characteristics has been performed, in a random order, during the other three experimental sessions. A schematic representation of the experimental procedures during the last three sessions is given in Fig. 1. After a warm-up routine (two series of six contractions at 60% MVC with 6 s on and 4 s off), warming or cooling maneuvers started about 30 min before contractions. In both procedures, two packs (mod. Thermogel, Artsana, Milan, Italy) were applied to the lateral surface of the investigated muscle by elastic straps. To avoid a direct application on the skin, packs were covered by two antiallergen sleeves. In TmL session, packs were kept in a refrigerator at 4 °C for 10 h. Once applied, packs reduced the temperature of the skin (Ts) over the biceps brachii from 32 ± 1 °C to 20 ± 2 °C in 30 min. In TmH session, packs were placed in a water bath (mod. TW8, Julabo, Seelbach, Germany) at 60 °C, and a raise in Ts from 31 ± 2 °C to 40 ± 2 °C was obtained in 30 min. Sessions in TmN condition were conducted with a Ts of 31 ± 2 °C. After positioning of the EMG probe in the same place as during reliability tests by the use of a transparency, the experimental protocol started with MVC assessment. Participants were then required to perform, in randomized order, five isometric contractions at 20%, 40%, 60%, 80% and 100% MVC, each interspersed by a 5 min rest.
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100
MVC 3s
5s
20 s
20 s
80 5s
60
5s
where d is the inter-electrode distance and s0 is the time delay between the two double differential signals (Farina et al., 2005). Only the epochs with a correlation coefficient >0.70 where considered for CV estimation. A linear regression analysis of CV values during 80% MVC was performed. 2.6. Statistical analysis
5s
40 5s 5 min
20
5s
6 s on / 4 s off 5s 5 min
% MVC
CV ¼ d=s0
MVC 3s
5s
0 Before
Fatiguing exercise
After
Fig. 1. Experimental design. A set of five contractions at 20%, 40%, 60%, 80% and 100% MVC was performed in a balanced random order, before and after the fatiguing exercise. The contractions set was performed at three different muscle temperatures, i.e., thermo-neutral (TmN, 34.1 ± 0.2 °C), high (TmH, 42.5 ± 0.3 °C) and low (TmL, 22.8 ± 0.7 °C) Tm. The fatiguing exercise consisted of isometric contractions at 50% MVC (6 s on–4 s off) until fatigue.
The isometric contraction at 80% MVC was sustained for 20 s, while the other contractions were performed in 5 s. Similarly to Esposito et al. (1998), a fatiguing exercise of the elbow flexors at 50% MVC (repetitive cycles of 6-s on and 4-s rest) was then performed until target force could not be reached for three times in a row. Before and immediately after the fatiguing exercise, participants indicated the level of perceived effort using a 6–20 RPE Borg Scale (Borg, 1982). Immediately after the fatiguing exercise MVC was re-assessed. Then, a second set of isometric contractions at 20%, 40%, 60%, 80% and 100% MVC was performed. The 20 s contraction at 80% MVC was included in the study, both before and after the fatiguing protocol, to assess the combined effect of Tm manipulation and fatigue on the time course of EMG parameters during a sustained contraction. During sessions, Ts was continuously monitored by three laser thermometers (mod. 826-T2, Testo, Lenzkirch, Germany), which measured the temperature on the proximal, medial and distal third of the muscle belly. Before tests, laser thermometers were calibrated against a reference thermo-couple (mod. 950, Testo, Lenzkirch Germany). Throughout the entire experimental session, a tolerance of ±1 °C was allowed. When temperature exceeded the level of tolerance, packs were replaced. According to a previous study (de Ruiter et al., 1999), during which Tm was calculated from the measured Ts at rest and during isometric contractions, the following equation was utilized:
Statistical analysis was performed using a SigmaPlot 12 software (Systat Software Inc., San Josè, USA). A sample size of 16 participants was selected to ensure a statistical power >0.70. The normal distribution of the sampling was checked by a Kolgomorov–Smirnov test. A three-way (fatigue [pre and post]protocol [TmN, TmH and TmL]contraction intensity [20%, 40%, 60%, 80% and 100% MVC]) ANOVA for repeated measures was applied to determine possible differences in EMG and force variables during the 5-s contractions. A three-way (fatigue [pre and post] protocol [TmN, TmH and TmL]time) ANOVA for repeated measures was applied to determine possible differences in force and EMG variables during 80% MVC. A two-way [fatigueprotocol] ANOVA for repeated measures was applied to determine possible differences among MVC values and linear regression slopes and intercepts. The posthoc Holm–Sidak test was applied when necessary to establish the location of the differences. The level of significance was set at P < 0.05. Test–retest reliability for MVC, EMG RMS, MF and CV was calculated by the Intraclass Correlation Coefficient (ICC) and the Standard Error of Measurements (SEM). We considered an ICC > 0.90 as very high, between 0.70 and 0.89 as high and between 0.50 and 0.69 as moderate (Munro, 1997). Unless otherwise stated, results are expressed as mean ± standard error (SE). 3. Results Fig. 2 shows Tb and Tm values throughout the experimental session during the three tested conditions. Tb and Tm values before warm-up were not significantly different among the three conditions. No significant changes in Tb occurred throughout the session. Immediately after warm-up, Tm increased significantly from 32.1 ± 0.3 °C, 31.6 ± 0.3 °C and 31.3 ± 0.3 °C to 33.5 ± 0.4, 34.2 ± 0.3 °C and 33.8 ± 0.5 °C for TmN, TmH and TmL, respectively (P < 0.05). Thereafter, Tm did not change in TmN, while after warming or cooling procedures, it changed significantly to 42.5 ± 0.3 °C and to 22.8 ± 0.7 °C in TmH and in TmL, respectively. From here on, Tm remained constant under all experimental conditions. At the end of the session, Tm was 34.4 ± 0.4 °C, 42.7 ± 0.2 °C and 22.7 ± 0.4 °C in TmN, TmH and TmL, respectively (P < 0.05).
50
Tm ¼ 1:02 Ts þ 0:89
2.5. Data analysis EMG was analyzed in time domain: to avoid transient phenomena from rest to exertion and vice versa, the RMS of the signal was calculated from epochs of 2 s, corresponding to the central part of the force plateau reached during each 5-s contraction. After dividing the same 2 s windows in 8 epochs of 250 ms each, the CV was estimated in double differential modality according to the following equation:
up
40 Temperature (°C)
Body temperature (Tb) was estimated using an infrared tympanic thermistor (mod. 510, Braun, Type IRT 1020, Kronberg, Germany), which was validated with a mercury thermometer (Quiromed, Valencia, Spain).
warm MVC
contractions
fatiguing MVC
contractions
exercise
*,#
Tb N Tb H Tb L
30
*
TmN TmH TmL
20 *, #
10 Fig. 2. Body (open symbols) and muscle (closed symbols) temperatures measured during the several protocol phases in TmN (circle), TmH (square) and TmL (triangle) experimental conditions. ⁄P < 0.05 vs pre warm up value; #P < 0.05 vs TmN.
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500 400
MVC (N)
The duration of the fatiguing exercise (time to fatigue) was similar under all experimental conditions (408 ± 28 s, 402 ± 29 s and 421 ± 30 s, in TmN, TmH and TmL, respectively; P > 0.05). After the fatiguing exercise, RPE increased significantly from 9 ± 1; 9 ± 2 and 9 ± 1 to 16 ± 2; 16 ± 1 and 15 ± 3 in TmN, TmH and TmL, respectively (P < 0.05). No inter-groups differences were found.
*
*
*
300 200 100
3.1. Reliability
3.2. Maximum voluntary contractions (MVC)
0 before after
1.0
EMG RMS (mV)
ICC values for force, EMG RMS and CV ranged from Cronbach’s a between 0.96–0.98, 0.93–0.97 and 0.89–0.92, respectively. No significant differences among the three sessions were found at any contraction level. According to our categories (see methods), reliability was very high, with the only exception of CV, which was between high and very high. SEM for force, EMG RMS and CV ranged from 1.56 N to 2.20 N, 0.01 mV to 0.02 mV and 0.05 m s1 to 0.06 m s1, respectively.
0.8 0.6 0.4 0.2 0.0
3.3. Brief contractions The mean values of EMG RMS were normalized as a function of the value at 100% MVC determined during the same session (0.658 ± 0.075, 0.669 ± 0.073 and 0.727 ± 0.076 mV before the fatiguing exercise in TmN, TmH and TmL, respectively; P > 0.05; 0.718 ± 0.105, 0.704 ± 0.078 and 0.746 ± 0.082 mV after the fatiguing exercise in TmN, TmH and TmL, respectively; P > 0.05). EMG RMS and CV at different % MVC in the three experimental conditions are given in Fig. 4, for both before and after the fatiguing exercise. EMG RMS increased significantly with the increase in the level of contraction (P < 0.05) in all sessions, both before and after the fatiguing exercise. ANOVA did not disclose significant effects of fatigue and interactions among the main factors. CV increased significantly from 20% to 100% MVC in all sessions, (P < 0.05), with CV values in TmL significantly lower than in TmN and TmH at all contraction intensities (P < 0.05), both before and after the fatiguing exercise. A significant interaction among the main factors was found (P < 0.05). After the fatiguing exercise, CV
5
CV (m s -1)
MVC, EMG RMS and CV mean values in the three experimental conditions are shown in Fig. 3, before and after fatiguing exercise. No significant differences were found among conditions in MVC values, neither before nor after the fatiguing exercise. MVC decreased significantly after the fatiguing exercise in all conditions (by 22.1 ± 1.6%, 18.6 ± 1.3% and 24.7 ± 1.1% for TmN, TmH and TmL, respectively; P < 0.05). No significant interaction between the two main factors was found. ANOVA did not disclose any significant effect of Tm and fatigue on EMG RMS values determined during MVC (0.649 ± 0.078, 0.670 ± 0.074 and 0.717 ± 0.066 mV before the fatiguing exercise in TmN, TmH and TmL, respectively; P > 0.05; 0.779 ± 0.084, 0.714 ± 0.081 and 0.776 ± 0.079 mV after the fatiguing exercise in TmN, TmH and TmL, respectively; P > 0.05). Moreover, no significant interaction between the two main factors was retrieved. A significant effect of Tm and fatigue on CV was found by ANOVA, without any significant interaction between the main factors. Post hoc analysis revealed that before the fatiguing exercise CV was significantly lower by about 20% in TmL than in the other conditions (P < 0.05). Moreover, after the fatiguing exercise, a significant decrease in CV occurred in all conditions (by 7.5 ± 1.4%, 6.7 ± 1.7% and 27.1 ± 1.6% for TmN, TmH and TmL, respectively; P < 0.05), the drop in CV being significantly larger in TmL (P < 0.05).
*
* #
4 3
*,#
2 1
TmN
TmH
TmL
Fig. 3. Mean MVC, EMG RMS and CV values before (black bars) and after (white bars) the fatiguing exercise, in TmN, TmH and TmL conditions. ⁄P < 0.05 after vs before fatiguing exercise; #P < 0.05 among conditions.
values in TmL dropped to a larger extent than in the other conditions at all contraction intensities (P < 0.05). 3.4. Sustained contractions The mean values of EMG RMS, normalized to the initial value (0.602 ± 0.068, 0.552 ± 0.050 and 0.551 ± 0.049 mV before the fatiguing exercise in TmN, TmH and TmL, respectively; P > 0.05; 0.638 ± 0.083, 0.582 ± 0.069 and 0.578 ± 0.060 mV after the fatiguing exercise in TmN, TmH and TmL, respectively; P > 0.05), and of CV during the sustained 80% MVC contraction before (left-side panels) and after (right-side panels) the fatiguing exercise, are shown in Fig. 5. Before the fatiguing exercise, EMG RMS increased significantly in the first 12 s of contraction in all conditions (P < 0.05). After the fatiguing exercise, EMG RMS did not change throughout contraction. No significant inter-groups differences were found, both before and after the fatiguing exercise. CV decreased significantly with time in all conditions, both before and after the fatiguing exercise (P < 0.05), with CV mean values in TmL always significantly lower than in TmN and TmH (P < 0.05). After the fatiguing exercise, CV was significantly lower than before in all conditions, the drop in TmL being significantly larger than in TmN and TmH. From linear regression analysis of CV as a function of contraction time (Table 2), a significant decrease in slope was found in TmN and TmH after the fatiguing exer-
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CV (m s-1)
EMG RMS (%)
Before
After
100
TmN pre TmH pre
100
TmN post TmH post
80
TmL pre
80
TmL post
60
60
40
40
20
20
0
0
5
5
4
4
3
3
2
2
#
1
*
1
0
#
0 20
40
60
80
100
Force (% MVC)
20
40
60
80
100
Force (% MVC)
Fig. 4. Mean values of normalized EMG RMS as a function of 100% MVC and CV during 5 s contractions, before (left column) and after (right column) fatiguing exercise, in TmN (circle), TmH (triangle) and TmL (square) conditions. ⁄P < 0.05 after vs before fatiguing exercise; #P < 0.05 among conditions.
cise (P < 0.05). A significant interaction among main factors was found (P < 0.05). Intercept in TmL was significantly lower than in TmN and TmH both before and after the fatiguing exercise (P < 0.05).
1981)). TmL in the present study was lower (about 23 °C), thus potentially explaining the discrepancy with previous reports.
4. Discussion
In agreement with previous studies (Mitchell et al., 2008; Mito et al., 2007), MVC and EMG RMS were unaffected by Tm manipulation. On the contrary, CV in TmL was significantly lower than in the other two experimental conditions. This was also the case during the brief contractions at submaximal intensities and during the sustained contraction at 80% MVC. During the brief submaximal contractions, the linear increase in EMG RMS with intensity is well in agreement with previous reports (Mito et al., 2007; Petrofsky and Laymon, 2005). On the contrary, previous studies (de Ruiter and de Haan, 2000; de Ruiter et al., 1999; Petrofsky and Lind, 1980; Ranatunga et al., 1987) found that below a Tm of 22 °C, a reduction in EMG amplitude occurs (Petrofsky and Lind, 1980). This discrepancy could be explained by (i) the slightly different TmL utilized (23 °C in the present study), and (ii) the muscle with a larger volume than those previously investigated, such as the first dorsal interosseum (Ranatunga et al., 1987), finger flexors (Holewijn and Heus, 1992; Petrofsky and Lind, 1980) and adductor pollicis (de Ruiter and de Haan, 2000; de Ruiter et al., 1999) muscles. Although Ts was similar along muscle surface, the calculation of Tm might have underestimated the temperature of the muscle fibers positioned deeper or near the vessels. Also CV increased significantly with force, but as previously stated, CV was always lower in TmL than in TmN and TmH. As supported by previous investigations (Kimura et al., 2003; Merletti et al., 1984; Mito et al., 2007; Mucke and Heuer, 1989; Petrofsky and Lind, 1980), this finding is suggestive of an alteration of the sarcolemmal propagation properties with cooling. In particular, a longer opening time of Na+ channels and a slowing of the delayed K+ rectifying current at the cell membrane level have been indicated as the main mechanisms responsible for the reduction in the motor unit action potential propagation velocity
The effects of Tm manipulation on muscle electrical activation and isometric force production of a previously fatigued skeletal muscle have never been investigated. The main result of the present study was that after the fatiguing exercise CV always decreased to a further extent in TmL compared to the other two experimental conditions, regardless of contraction intensity or duration. This finding suggests that TmL and fatigue seem to have a combined and additional effect in decreasing sarcolemmal propagation velocity. However, this combined effect was not accompanied by Tm-induced changes in EMG RMS and maximum force production.
4.1. Preliminary considerations Given the similar Tb during the three experimental conditions, the changes in the investigated parameters retrieved in the present study can be reasonably attributed to Tm manipulation itself. The lack of differences among conditions in time to fatigue and in RPE values at the end of the fatiguing exercise is in line with previous reports (Blomstrand et al., 1985; Lannergren and Westerblad, 1988) and suggests that Tm, at least in the temperature range considered in the present study, had no significant effect on muscle fatigability and on the level of perceived exertion. Previous investigations reported a cooling-induced increase in time to fatigue in an electrically stimulated rat muscle (Segal et al., 1986] and in a voluntary activated plantar flexor muscle (Petrofsky and Lind, 1981). Both studies found cooling effects on time to fatigue only in a temperature range close to that reported to maximize muscle endurance (27–29 °C; (Edwards et al., 1972; Petrofsky and Lind,
4.2. Effect of Tm manipulation
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-1
CV (m s )
EMG RMS (%)
Before
After
160
TmN pre TmH pre
160
TmN post TmH post
140
TmL pre
140
TmL post
120
120
100
100
80
80
5
5
4
4
3
3
2
*
2
# 1
1
0
0 0
5
10
15
20
# 0
5
time (s)
10
15
20
time (s)
Fig. 5. Mean values of EMG RMS, normalised as a function of the initial value, and CV during 80% MVC (20 s), before (left column) and after (right column) fatiguing exercise, in TmN (circle), TmH (triangle) and TmL (square) conditions. ⁄P < 0.05 after vs before fatiguing exercise; #P < 0.05 among conditions.
Table 2 Linear regression analysis of CV as a function of contraction time during 80% MVC in thermo-neutral (TmN), high (TmH) and low (TmL) muscle temperature, before (left-side columns) and after (right-side columns) the fatiguing protocol. ⁄P < 0.05 vs before; #P < 0.05 vs other Tm. Before TmN TmH TmL
Slope (m s2) 0.08 ± 0.01 0.07 ± 0.01 0.06 ± 0.01
After Intercept (m s1) 3.7 ± 0.2 4.4 ± 0.3 3.5 ± 0.2#
R 0.93 ± 0.01 0.95 ± 0.02 0.95 ± 0.01
(Hicks and McComas, 1989; Hodgkin and Huxley, 1952; Merletti et al., 1984). During sustained contractions, the time course of EMG RMS was the same in all experimental conditions. This is in agreement with previous findings in TmN (Esposito et al., 1998; Krogh-Lund and Jorgensen, 1993; Linssen et al., 1993), TmH (Holewijn and Heus, 1992; Petrofsky and Laymon, 2005) and TmL (Holewijn and Heus, 1992; Petrofsky and Laymon, 2005). The increase in EMG RMS during sustained contractions of heavy intensity has been attributed to (i) changes in muscle fiber action potential and CV (De Luca, 1984); (ii) recruitment of additional motor units, so to keep the force output constant (Gamet and Maton, 1989; Linssen et al., 1993); and (iii) the synchronization of active motor units (KroghLund and Jorgensen, 1993). Throughout the sustained contraction, CV decreased with time in all experimental conditions. An alteration in the sarcolemmal action potential characteristics, due to changes in intramuscular metabolites, was alleged to explain the linear decay with time of this parameter (Arabadzhiev et al., 2010; Fitts, 1994; Merletti et al., 1992). However, CV was always significantly lower in TmL than in TmN and TmH, suggesting that muscle cooling has affected sarcolemmal propagation properties since the beginning of contraction, as witnessed also by the significantly lower intercept of the CV vs time relationship in TmL (see Table 2). Despite no previous studies reported direct CV changes due to Tm manipulation during sustained contractions of heavy intensity, a lower EMG mean frequency, a parameter strongly influenced by CV, was found
Slope (m s2) 0.10 ± 0.01⁄ 0.11 ± 0.01⁄ 0.07 ± 0.01
Intercept (m s1) 3.5 ± 0.2 4.4 ± 0.3 3.1 ± 0.3⁄,#
R 0.94 ± 0.03 0.95 ± 0.01 0.96 ± 0.01
in TmL (Petrofsky and Laymon, 2005), giving further evidence of a slowed motor unit action potential propagation with cooling. No comparisons with previous reports, though, can be made in TmH.
4.3. Combined effect of fatigue and Tm manipulation After the fatiguing exercise, MVC decreased similarly in all experimental conditions, suggesting that neither TmH nor TmL depressed to a further extent muscle force generating capacity. EMG RMS determined during MVC did not change significantly in all experimental conditions, indicating that central command was not affected by Tm manipulation or fatigue. Nevertheless, in spite of a similar muscle electrical activation, after the fatiguing exercise MVC was significantly lower, unveiling a decrease in neuromuscular efficiency, defined as the ratio of force output to the EMG amplitude (Milner-Brown et al., 1986). However, Tm manipulation did not play an additional role in this phenomenon, as already reported in the literature (Mitchell et al., 2008; Mito et al., 2007). EMG RMS was found to be similar under all experimental conditions also during brief and sustained contractions, both before and after the fatiguing exercise. Moreover, as previously reported (Esposito et al., 1998), after fatigue EMG RMS showed a tendency to an increase (P = 0.09), possibly due to the previously cited and still persisting mechanisms involved in increasing the power of the signal with fatigue, i.e., changes in the muscle fiber action potentials, decreases in CV, and synchronization/grouping of active MU.
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Under all the three Tm conditions, CV showed a significant reduction with fatigue, which might be explained by the previously described alteration of sarcolemmal propagation properties due to changes in intramuscular metabolites (altered intra/extracellular K+ concentration and increased intracellular H+ concentration) occurring with fatigue (Arabadzhiev et al., 2010; Fitts, 1994; Merletti et al., 1992). Noteworthy, the CV drop in TmL was larger than in the other conditions, and this was also the case during both brief and sustained contractions. Moreover, CV values in TmL at the beginning of the sustained contraction after the fatiguing exercise were not different from the last CV value before the fatiguing exercise. This was not the case in TmN and TmH. Collectively, these findings are suggestive of a combined and additional effect of TmL and fatigue on the sarcolemmal propagation properties. The larger CV drop in TmL, though, was not accompanied by a broader reduction in MVC. As an explanation, it might be speculated that the reduction in Ca2+ efflux from the sarcoplasmic reticulum in TmL (Caputo, 1972a,b) might have been counteracted by the increase in Ca2+ sensitivity of the thin-filament regulatory system with cooling (de Ruiter et al., 1999; Godt and Lindley, 1982; MacIntosh, 2010; Xu et al., 2003). Another possible mechanism could be a synergistic compensation to elbow flexion of the brachioradialis muscle, which was only marginally affected by cooling but contributed to the force output. Conversely, CV was measured on the belly of the biceps brachii muscle, which was directly affected by Tm manipulation procedures. Further studies, though, are needed to provide support to these hypotheses. Interestingly, whereas CV time course during sustained contraction had a similar slope in all Tm conditions before the fatiguing exercise, after fatigue the slope increased significantly in TmN and TmH, but not in TmL. Given the already lower initial CV values of TmL after fatigue, the effect of cooling on sarcolemmal propagation properties was limited and CV could not be reduced below a certain value.
5. Conclusion During both brief and sustained contractions, CV values were constantly lower in TmL than in the other Tm. After fatigue, muscle cooling decreased CV to a further extent compared to the other conditions, suggesting a combined and additional effect of cooling and fatigue in altering the sarcolemmal propagation properties. However, Tm manipulation did not affect time to fatigue and force generating capacity, neither before nor after the fatiguing exercise. A change in Tm, at least in the range of temperatures investigated in the present study, did not show detrimental effects on force output, in spite of altered sarcolemmal propagation properties. The present findings may have practical implications also for sport trainers and/or physiotherapists about muscle performance after Tm manipulation (for example after diathermy or local cryotheraphy) under both fresh and fatigued conditions.
Conflicts of interest None declared.
Acknowledgements The authors wish to thank all the participants involved in the study, for their patience and committed involvement. The study was supported by the University of Milan (PUR grant #12-15059281-42).
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Emiliano Cè was born in Italy in 1976. He received the degree in Sport Sciences from the University of Milan (Italy) in 2002. He achieved his PhD in Morphological Sciences (2007) at the University of Milan (Italy). He received the BSc in Osteopathy from the University of Wales (Wales, UK) in 2010. Present position: He is Assistant Professor at the Department of Sport, Nutrition and Health Sciences of the University of Milan (Italy). His research interests are addressed to the study of muscle biological signals, in particular force, surface electromyogram (EMG) and mechanomyogram (MMG), in muscle activity during voluntary or stimulated contractions in different physiological models (fatigue, training, temperature, etc.). Currently his scientific activity is focused on the properties of the muscle-tendon unit mechanical model and the possibility to monitor the motor unit activation strategy by the analysis of force, EMG and MMG signals. He is a member of the Italian Society of Sport Sciences (SISMES) and of the Interuniversity Institute of Myology (IIM).
Susanna Rampichini graduated in Physical Education from Istituto Superiore di Educazione Fisica di Milano, Italy in 1998 and in Biomedical Engineering from Politecnico di Milano (Italy) in 2003. She actually works at the laboratory of Exercise physiology of the Department of Sport, Nutrition and Health Sciences of the University of Milan (Italy) as an assistant of research. She is interested in the neuromuscular control and in biological signal processing. She is a member of the Italian Society of Sport Sciences (SISMES).
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Luca Agnello was born in 1983. He graduated in Sport Science at the University of Milan, Italy, in 2007. He received his PhD in Biochemistry, Physiology and Pathology of the Muscle at the Department of Basic and Applied Medical Sciences, University of Chieti, in 2011.
Eloisa Limonta was born in Milan in 1979. She graduated in Exercise Sciences at the University of Milan in 2002. She received her PhD in Sport and Physical Activity in 2007 at University of Milan and she is an Assistant Professor at the Department of Sport, Nutrition and Health Sciences since 2005. Her research field is in Exercise Physiology and Training in Sports, with special interests in electrical and mechanical aspects of muscle contraction and in physiological responses to different strength training modalities.
Arsenio Veicsteinas entered in the Institute of Physiology, School of Medicine, University of Milan to initiate his thesis in 1968 under the supervision of Rodolfo Margaria and Paolo Cerretelli. Then in 1971 received the position of Assistant Professor with Rodolfo Margaria. In 1980 he moved to the newly founded University of Brescia as Associate Professor, where he developed the Laboratory of Exercise Physiology. In 2001 returned to the Faculty of Exercise Sciences of the University of Milan as Professor of Physiology. As Visiting Professor spent several time in the Physiology Department, Max Planck Institute for Experimental Medicine, Goettingen, Germany (under the supervision of Johannes Piiper), and in the Dept. of Physiology, Yale University (John Stitt and Ethan Nadel), and SUNY at Buffalo, U.S. (Donald Rennie and David Pendergast). The reserch fields of interest cover the Physiology of Muscle Exercise (metabolism, respiratory and cardiocirculatory adjustments to exercise), the control of Muscle Contraction (electromyogram and mechanomyogram), the study of the Biological Effects of electromagnetic fields on animals and cells, and more recently the molecular mechanism involved in exercise and in cardioprotection.
Fabio Esposito was born in Vedano al Lambro (Milan, Italy) in 1963. He received his medical degree from the University of Milan in 1990 and the specialization in Sports Medicine from the University of Brescia in 1994. He joined the faculty at University of Milan in 2003 and is currently Associate Professor of Physiology. His research field is Exercise Physiology, with special interests in muscle electro-mechanical behavior, pulmonary gas exchange, physiological responses to exercise training and exercise-induced cardioprotection. He is a member of the Italian Physiological Society (SIF), of the Italian Society of Sport Sciences (SISMES) and of the American College of Sport Medicine (ACSM).
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Journal of Electromyography and Kinesiology journal homepage: www.elsevier.com/locate/jelekin
Combined effects of fatigue and temperature manipulation on skeletal muscle electrical and mechanical characteristics during isometric contraction Emiliano Cè a,b,⇑, Susanna Rampichini a, Luca Agnello b, Eloisa Limonta a, Arsenio Veicsteinas a,b, Fabio Esposito a,c a b c
Department of Sport, Nutrition and Health Sciences, University of Milan, Via G. Colombo 71, 20133 Milan, Italy Center of Sport Medicine, Don Gnocchi Foundation, Via Capecelatro 66, 20148 Milan, Italy Centro Sicurezza nelle Attività Sportive e Corretti Stili di Vita (SAS), University of Milan, Via G. Colombo 71, 20133 Milan, Italy
a r t i c l e
i n f o
Article history: Received 19 December 2011 Received in revised form 19 January 2012 Accepted 19 January 2012
Keywords: EMG Isometric contraction Muscle temperature Neuromuscular efficiency Conduction velocity
a b s t r a c t Peripheral fatigue and muscle cooling induce similar effects on sarcolemmal propagation properties. The aim of the study was to assess the combined effects of muscle temperature (Tm) manipulation and fatigue on skeletal muscle electrical and mechanical characteristics during isometric contraction. After maximum voluntary contraction (MVC) assessment, 16 participants performed brief and sustained isometric tasks of different intensities in low (TmL), high (TmH) and neutral (TmN) temperature conditions, before and after a fatiguing exercise (6 s on/4 s off at 50% MVC, to the point of fatigue). During contraction, the surface electromyogram (EMG) and force were recorded from the biceps brachii muscle. The root mean square (RMS) and conduction velocity (CV) were calculated off-line. After the fatiguing exercise: (i) MVC decreased similarly in all Tm conditions (P < 0.05), while EMG RMS did not change; and (ii) CV decreased to a further extent in TmL compared to TmN and TmH in all brief and sustained contractions (P < 0.05). The larger CV drop in TmL after fatigue suggests that TmL and fatigue have a combined and additional effect on sarcolemmal propagation properties. Despite these changes, force generating capacity was not affected by Tm manipulation. A compensatory mechanism has been proposed to explain this phenomenon. Ó 2012 Elsevier Ltd. All rights reserved.
1. Introduction Temperature is an important determinant of skeletal muscle contractile and metabolic properties (Gray et al., 2006; Rall and Woledge, 1990; Ranatunga, 1998). Several investigations have dealt with the role of local temperature on skeletal muscle performance, yet its effects on neuromuscular activation still remain poorly understood. While some studies did not retrieve any significant alteration in neuromuscular function due to changes in muscle temperature (Tm) (Mitchell et al., 2008; Mito et al., 2007), the manipulation of Tm has been found to modify significantly neuromuscular activation during both voluntary (Cheung and Sleivert, 2004; de Ruiter et al., 1999; Dewhurst et al., 2010; Holewijn and Heus, 1992; Petrofsky and Laymon, 2005; Petrofsky and Lind, 1980; Racinais and Oksa, 2010; Ranatunga et al., 1987) and electrically-evoked contractions (Bigland-Ritchie et al., 1992; de Ruiter et al., 1999; Hopf and Maurer, 1990; Racinais and Oksa, 2010; Ranatunga et al., ⇑ Corresponding author. Address: Department of Sport, Nutrition and Health Sciences, Division of Human Physiology, University of Milan, Via G. Colombo 71, 20133 Milan, Italy. Tel.: +39 02 5031 4644; fax: +39 02 5031 4630. E-mail address: [email protected] (E. Cè). 1050-6411/$ - see front matter Ó 2012 Elsevier Ltd. All rights reserved. doi:10.1016/j.jelekin.2012.01.012
1987). In particular, despite no changes have been reported with high Tm (TmH) (Cheung and Sleivert, 2004; Dewhurst et al., 2010; Holewijn and Heus, 1992; Petrofsky and Laymon, 2005), a significant decrease in force generating capacity (Bigland-Ritchie et al., 1992; Cheung and Sleivert, 2004; de Ruiter et al., 1999; Holewijn and Heus, 1992; Hopf and Maurer, 1990; Petrofsky and Laymon, 2005; Ranatunga et al., 1987), in surface electromyogram (EMG) frequency content (Dewhurst et al., 2010; Holewijn and Heus, 1992; Petrofsky and Lind, 1980; Ranatunga et al., 1987) and in mean fiber conduction velocity (CV) (Merletti et al., 1984; Mucke and Heuer, 1989) have been previously described with low Tm (TmL). In TmL, indeed, a change in the action potential architecture and a decrease in its propagation velocity occur (Hicks and McComas, 1989; Hodgkin and Huxley, 1952) together with a reduction in Ca2+ release from the sarcoplasmic reticulum (Caputo, 1972a,b). This leads to a decline in adenosine triphosphate availability and cross-bridge cycling rate (Godt and Lindley, 1982). Also muscle fatigue, which is generally defined as a temporary loss in force generating capacity due to previous muscle contractions (Fitts, 1994), shows similar effects on the neuromuscular activation to those induced by TmL. Fatigue, indeed, decreases
E. Cè et al. / Journal of Electromyography and Kinesiology 22 (2012) 348–355
the motor unit action potential amplitude, its propagation across the sarcolemma, the Ca2+ efflux from sarcoplasmic reticulum and the cross-bridge cycling rate (Allen et al., 1989, 1995; Fitts, 2008). Surface EMG detection by linear electrode arrays is a non-invasive and reliable tool to provide information on muscle electrical activation (Merletti et al., 2003). The alterations in sarcolemmal propagation properties, action potential duration and muscle activation strategies can be evaluated indirectly by EMG time and frequency domain analysis and CV calculation (Farina et al., 2005; Gray et al., 2006; Merletti et al., 2003). While the separate effects of muscle fatigue and TmL alone have been already investigated during isometric contraction (Merletti et al., 1984, 2003; Mucke and Heuer, 1989), to the best of our knowledge, no studies have examined the effect of cooling on the electrical activation and force generating capacity of a previously fatigued muscle. TmL and fatigue, indeed, may have a combined and additional effect on the sarcolemmal propagation properties and force production. Thus, the aim of the present study was to assess the effects of Tm manipulation on the neuromuscular activation characteristics and maximum voluntary contraction (MVC) of a previously fatigued muscle. We hypothesize that, given the partly shared consequences of cooling and fatigue on muscle characteristics, the changes in the fatigued muscle would be more pronounced in TmL than in TmH or in thermo-neutral condition (TmN). To this purpose, we assessed the maximum force production of the elbow flexors and the electrical activation of the biceps brachii muscle at different contraction intensities and durations, before and after a fatiguing exercise. 2. Methods 2.1. Participants After a full explanation of the purpose of the study and experimental procedures, 16 male participants gave their written informed consent to be enrolled in the study. Their physical and anthropometric characteristics are given in Table 1. Participants were all clinically healthy, with no previous history of upper limbs, shoulder joint and muscle injuries. At the time of the study, none of them was involved in any specific training program. The study was approved by the local University Ethics Committee and performed in accordance with the principles of the 1975 Declaration of Helsinki. 2.2. Experimental design Before the experimental sessions, participants were allowed to familiarize with the ergometer and to test their ability to reach and maintain different contraction levels. Then, participants reported to the laboratory six times on different days, with at least 72 h of rest in between, during which they were invited to avoid fatiguing efforts involving the muscles of the upper limbs. The first three sessions were devoted to assess test–retest reliability of force and EMG parameters in TmN. In the other three sessions, the effects of TmN, TmH and TmL were evaluated in random order. All experiments were carried out in a laboratory at a constant temperature of 22 ± 1 °C and relative humidity of 50 ± 5%.
Table 1 Physical and anthropometric characteristics of the participants (n = 16). Data are expressed as mean ± SD. Age (yrs) Body mass (kg) Stature (m) Arm circumference (cm) Biceps brachii skin fold (mm)
23 ± 6 72.5 ± 5.8 1.73 ± 0.04 31.3 ± 2.4 5.1 ± 1.9
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2.3. Reliability assessment As previously described, the first three sessions were devoted to assess the reliability of the measurements. The maximum voluntary contraction (MVC) of each subject was the highest value of three maximum efforts of the elbow flexor muscles of the dominant arm. Then, contractions at 20%, 40%, 60%, 80% and 100% MVC of 5 s each were carried out in a random order. During contractions, the EMG signal was detected from the biceps brachii muscle. From the two central seconds of each contraction, the mean value of force, EMG root mean square (RMS) and CV were calculated. Before the first session, a map with some skin identification points (moles, angiomas and scars) and the positioning of the EMG probe was drawn on a transparency, to allow repeated measurements from the same muscle area during all tests. Each participant was positioned on an ergometer with the hand kept in a position midway between pronation and supination at an elbow angle of 115°. The whole detection apparatus had a resonant frequency >200 Hz. The skin area under the electrodes was carefully cleaned with ethyl alcohol and gently abraded with fine sand paper. A conductive gel was also applied to achieve an inter-electrode impedance below 2000 X. Surface EMG signal was detected by a 16-channel linear array, which included 16 silver bar electrodes (diameter 1 mm, length 5 mm, inter-electrode distance 10 mm) for differential EMG detection. The array was placed along muscle fibers direction, with the EMG electrodes positioned perpendicularly to the major axes of the fibers, in accordance with the European recommendations about surface EMG (Hermens et al., 2000). The force output was detected by a calibrated load cell (mod SM-1000 N, Interface, Crowthorne, UK), operating linearly between 0 and 1000 N. EMG and force signals were amplified and filtered (mod. ASE16, LiSin, Turin, Italy; gain 1000 and 50 for EMG and force, respectively; bandwidth of 10–500 Hz and 2– 20 Hz for EMG and force, respectively), converted to digital data by a 12-bit acquisition board (mod. DAQcard-6024E, National Instruments, Austin, USA; sample rate of 2048 Hz) and stored on a personal computer for further analysis. Contraction intensity was maintained within ±3% of the requested force target with the aid of a visual feedback (both target and force output shown on a computer screen). 2.4. Temperature effects assessment The assessment of the effects of temperature manipulation on neuromuscular activation characteristics has been performed, in a random order, during the other three experimental sessions. A schematic representation of the experimental procedures during the last three sessions is given in Fig. 1. After a warm-up routine (two series of six contractions at 60% MVC with 6 s on and 4 s off), warming or cooling maneuvers started about 30 min before contractions. In both procedures, two packs (mod. Thermogel, Artsana, Milan, Italy) were applied to the lateral surface of the investigated muscle by elastic straps. To avoid a direct application on the skin, packs were covered by two antiallergen sleeves. In TmL session, packs were kept in a refrigerator at 4 °C for 10 h. Once applied, packs reduced the temperature of the skin (Ts) over the biceps brachii from 32 ± 1 °C to 20 ± 2 °C in 30 min. In TmH session, packs were placed in a water bath (mod. TW8, Julabo, Seelbach, Germany) at 60 °C, and a raise in Ts from 31 ± 2 °C to 40 ± 2 °C was obtained in 30 min. Sessions in TmN condition were conducted with a Ts of 31 ± 2 °C. After positioning of the EMG probe in the same place as during reliability tests by the use of a transparency, the experimental protocol started with MVC assessment. Participants were then required to perform, in randomized order, five isometric contractions at 20%, 40%, 60%, 80% and 100% MVC, each interspersed by a 5 min rest.
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100
MVC 3s
5s
20 s
20 s
80 5s
60
5s
where d is the inter-electrode distance and s0 is the time delay between the two double differential signals (Farina et al., 2005). Only the epochs with a correlation coefficient >0.70 where considered for CV estimation. A linear regression analysis of CV values during 80% MVC was performed. 2.6. Statistical analysis
5s
40 5s 5 min
20
5s
6 s on / 4 s off 5s 5 min
% MVC
CV ¼ d=s0
MVC 3s
5s
0 Before
Fatiguing exercise
After
Fig. 1. Experimental design. A set of five contractions at 20%, 40%, 60%, 80% and 100% MVC was performed in a balanced random order, before and after the fatiguing exercise. The contractions set was performed at three different muscle temperatures, i.e., thermo-neutral (TmN, 34.1 ± 0.2 °C), high (TmH, 42.5 ± 0.3 °C) and low (TmL, 22.8 ± 0.7 °C) Tm. The fatiguing exercise consisted of isometric contractions at 50% MVC (6 s on–4 s off) until fatigue.
The isometric contraction at 80% MVC was sustained for 20 s, while the other contractions were performed in 5 s. Similarly to Esposito et al. (1998), a fatiguing exercise of the elbow flexors at 50% MVC (repetitive cycles of 6-s on and 4-s rest) was then performed until target force could not be reached for three times in a row. Before and immediately after the fatiguing exercise, participants indicated the level of perceived effort using a 6–20 RPE Borg Scale (Borg, 1982). Immediately after the fatiguing exercise MVC was re-assessed. Then, a second set of isometric contractions at 20%, 40%, 60%, 80% and 100% MVC was performed. The 20 s contraction at 80% MVC was included in the study, both before and after the fatiguing protocol, to assess the combined effect of Tm manipulation and fatigue on the time course of EMG parameters during a sustained contraction. During sessions, Ts was continuously monitored by three laser thermometers (mod. 826-T2, Testo, Lenzkirch, Germany), which measured the temperature on the proximal, medial and distal third of the muscle belly. Before tests, laser thermometers were calibrated against a reference thermo-couple (mod. 950, Testo, Lenzkirch Germany). Throughout the entire experimental session, a tolerance of ±1 °C was allowed. When temperature exceeded the level of tolerance, packs were replaced. According to a previous study (de Ruiter et al., 1999), during which Tm was calculated from the measured Ts at rest and during isometric contractions, the following equation was utilized:
Statistical analysis was performed using a SigmaPlot 12 software (Systat Software Inc., San Josè, USA). A sample size of 16 participants was selected to ensure a statistical power >0.70. The normal distribution of the sampling was checked by a Kolgomorov–Smirnov test. A three-way (fatigue [pre and post]protocol [TmN, TmH and TmL]contraction intensity [20%, 40%, 60%, 80% and 100% MVC]) ANOVA for repeated measures was applied to determine possible differences in EMG and force variables during the 5-s contractions. A three-way (fatigue [pre and post] protocol [TmN, TmH and TmL]time) ANOVA for repeated measures was applied to determine possible differences in force and EMG variables during 80% MVC. A two-way [fatigueprotocol] ANOVA for repeated measures was applied to determine possible differences among MVC values and linear regression slopes and intercepts. The posthoc Holm–Sidak test was applied when necessary to establish the location of the differences. The level of significance was set at P < 0.05. Test–retest reliability for MVC, EMG RMS, MF and CV was calculated by the Intraclass Correlation Coefficient (ICC) and the Standard Error of Measurements (SEM). We considered an ICC > 0.90 as very high, between 0.70 and 0.89 as high and between 0.50 and 0.69 as moderate (Munro, 1997). Unless otherwise stated, results are expressed as mean ± standard error (SE). 3. Results Fig. 2 shows Tb and Tm values throughout the experimental session during the three tested conditions. Tb and Tm values before warm-up were not significantly different among the three conditions. No significant changes in Tb occurred throughout the session. Immediately after warm-up, Tm increased significantly from 32.1 ± 0.3 °C, 31.6 ± 0.3 °C and 31.3 ± 0.3 °C to 33.5 ± 0.4, 34.2 ± 0.3 °C and 33.8 ± 0.5 °C for TmN, TmH and TmL, respectively (P < 0.05). Thereafter, Tm did not change in TmN, while after warming or cooling procedures, it changed significantly to 42.5 ± 0.3 °C and to 22.8 ± 0.7 °C in TmH and in TmL, respectively. From here on, Tm remained constant under all experimental conditions. At the end of the session, Tm was 34.4 ± 0.4 °C, 42.7 ± 0.2 °C and 22.7 ± 0.4 °C in TmN, TmH and TmL, respectively (P < 0.05).
50
Tm ¼ 1:02 Ts þ 0:89
2.5. Data analysis EMG was analyzed in time domain: to avoid transient phenomena from rest to exertion and vice versa, the RMS of the signal was calculated from epochs of 2 s, corresponding to the central part of the force plateau reached during each 5-s contraction. After dividing the same 2 s windows in 8 epochs of 250 ms each, the CV was estimated in double differential modality according to the following equation:
up
40 Temperature (°C)
Body temperature (Tb) was estimated using an infrared tympanic thermistor (mod. 510, Braun, Type IRT 1020, Kronberg, Germany), which was validated with a mercury thermometer (Quiromed, Valencia, Spain).
warm MVC
contractions
fatiguing MVC
contractions
exercise
*,#
Tb N Tb H Tb L
30
*
TmN TmH TmL
20 *, #
10 Fig. 2. Body (open symbols) and muscle (closed symbols) temperatures measured during the several protocol phases in TmN (circle), TmH (square) and TmL (triangle) experimental conditions. ⁄P < 0.05 vs pre warm up value; #P < 0.05 vs TmN.
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500 400
MVC (N)
The duration of the fatiguing exercise (time to fatigue) was similar under all experimental conditions (408 ± 28 s, 402 ± 29 s and 421 ± 30 s, in TmN, TmH and TmL, respectively; P > 0.05). After the fatiguing exercise, RPE increased significantly from 9 ± 1; 9 ± 2 and 9 ± 1 to 16 ± 2; 16 ± 1 and 15 ± 3 in TmN, TmH and TmL, respectively (P < 0.05). No inter-groups differences were found.
*
*
*
300 200 100
3.1. Reliability
3.2. Maximum voluntary contractions (MVC)
0 before after
1.0
EMG RMS (mV)
ICC values for force, EMG RMS and CV ranged from Cronbach’s a between 0.96–0.98, 0.93–0.97 and 0.89–0.92, respectively. No significant differences among the three sessions were found at any contraction level. According to our categories (see methods), reliability was very high, with the only exception of CV, which was between high and very high. SEM for force, EMG RMS and CV ranged from 1.56 N to 2.20 N, 0.01 mV to 0.02 mV and 0.05 m s1 to 0.06 m s1, respectively.
0.8 0.6 0.4 0.2 0.0
3.3. Brief contractions The mean values of EMG RMS were normalized as a function of the value at 100% MVC determined during the same session (0.658 ± 0.075, 0.669 ± 0.073 and 0.727 ± 0.076 mV before the fatiguing exercise in TmN, TmH and TmL, respectively; P > 0.05; 0.718 ± 0.105, 0.704 ± 0.078 and 0.746 ± 0.082 mV after the fatiguing exercise in TmN, TmH and TmL, respectively; P > 0.05). EMG RMS and CV at different % MVC in the three experimental conditions are given in Fig. 4, for both before and after the fatiguing exercise. EMG RMS increased significantly with the increase in the level of contraction (P < 0.05) in all sessions, both before and after the fatiguing exercise. ANOVA did not disclose significant effects of fatigue and interactions among the main factors. CV increased significantly from 20% to 100% MVC in all sessions, (P < 0.05), with CV values in TmL significantly lower than in TmN and TmH at all contraction intensities (P < 0.05), both before and after the fatiguing exercise. A significant interaction among the main factors was found (P < 0.05). After the fatiguing exercise, CV
5
CV (m s -1)
MVC, EMG RMS and CV mean values in the three experimental conditions are shown in Fig. 3, before and after fatiguing exercise. No significant differences were found among conditions in MVC values, neither before nor after the fatiguing exercise. MVC decreased significantly after the fatiguing exercise in all conditions (by 22.1 ± 1.6%, 18.6 ± 1.3% and 24.7 ± 1.1% for TmN, TmH and TmL, respectively; P < 0.05). No significant interaction between the two main factors was found. ANOVA did not disclose any significant effect of Tm and fatigue on EMG RMS values determined during MVC (0.649 ± 0.078, 0.670 ± 0.074 and 0.717 ± 0.066 mV before the fatiguing exercise in TmN, TmH and TmL, respectively; P > 0.05; 0.779 ± 0.084, 0.714 ± 0.081 and 0.776 ± 0.079 mV after the fatiguing exercise in TmN, TmH and TmL, respectively; P > 0.05). Moreover, no significant interaction between the two main factors was retrieved. A significant effect of Tm and fatigue on CV was found by ANOVA, without any significant interaction between the main factors. Post hoc analysis revealed that before the fatiguing exercise CV was significantly lower by about 20% in TmL than in the other conditions (P < 0.05). Moreover, after the fatiguing exercise, a significant decrease in CV occurred in all conditions (by 7.5 ± 1.4%, 6.7 ± 1.7% and 27.1 ± 1.6% for TmN, TmH and TmL, respectively; P < 0.05), the drop in CV being significantly larger in TmL (P < 0.05).
*
* #
4 3
*,#
2 1
TmN
TmH
TmL
Fig. 3. Mean MVC, EMG RMS and CV values before (black bars) and after (white bars) the fatiguing exercise, in TmN, TmH and TmL conditions. ⁄P < 0.05 after vs before fatiguing exercise; #P < 0.05 among conditions.
values in TmL dropped to a larger extent than in the other conditions at all contraction intensities (P < 0.05). 3.4. Sustained contractions The mean values of EMG RMS, normalized to the initial value (0.602 ± 0.068, 0.552 ± 0.050 and 0.551 ± 0.049 mV before the fatiguing exercise in TmN, TmH and TmL, respectively; P > 0.05; 0.638 ± 0.083, 0.582 ± 0.069 and 0.578 ± 0.060 mV after the fatiguing exercise in TmN, TmH and TmL, respectively; P > 0.05), and of CV during the sustained 80% MVC contraction before (left-side panels) and after (right-side panels) the fatiguing exercise, are shown in Fig. 5. Before the fatiguing exercise, EMG RMS increased significantly in the first 12 s of contraction in all conditions (P < 0.05). After the fatiguing exercise, EMG RMS did not change throughout contraction. No significant inter-groups differences were found, both before and after the fatiguing exercise. CV decreased significantly with time in all conditions, both before and after the fatiguing exercise (P < 0.05), with CV mean values in TmL always significantly lower than in TmN and TmH (P < 0.05). After the fatiguing exercise, CV was significantly lower than before in all conditions, the drop in TmL being significantly larger than in TmN and TmH. From linear regression analysis of CV as a function of contraction time (Table 2), a significant decrease in slope was found in TmN and TmH after the fatiguing exer-
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CV (m s-1)
EMG RMS (%)
Before
After
100
TmN pre TmH pre
100
TmN post TmH post
80
TmL pre
80
TmL post
60
60
40
40
20
20
0
0
5
5
4
4
3
3
2
2
#
1
*
1
0
#
0 20
40
60
80
100
Force (% MVC)
20
40
60
80
100
Force (% MVC)
Fig. 4. Mean values of normalized EMG RMS as a function of 100% MVC and CV during 5 s contractions, before (left column) and after (right column) fatiguing exercise, in TmN (circle), TmH (triangle) and TmL (square) conditions. ⁄P < 0.05 after vs before fatiguing exercise; #P < 0.05 among conditions.
cise (P < 0.05). A significant interaction among main factors was found (P < 0.05). Intercept in TmL was significantly lower than in TmN and TmH both before and after the fatiguing exercise (P < 0.05).
1981)). TmL in the present study was lower (about 23 °C), thus potentially explaining the discrepancy with previous reports.
4. Discussion
In agreement with previous studies (Mitchell et al., 2008; Mito et al., 2007), MVC and EMG RMS were unaffected by Tm manipulation. On the contrary, CV in TmL was significantly lower than in the other two experimental conditions. This was also the case during the brief contractions at submaximal intensities and during the sustained contraction at 80% MVC. During the brief submaximal contractions, the linear increase in EMG RMS with intensity is well in agreement with previous reports (Mito et al., 2007; Petrofsky and Laymon, 2005). On the contrary, previous studies (de Ruiter and de Haan, 2000; de Ruiter et al., 1999; Petrofsky and Lind, 1980; Ranatunga et al., 1987) found that below a Tm of 22 °C, a reduction in EMG amplitude occurs (Petrofsky and Lind, 1980). This discrepancy could be explained by (i) the slightly different TmL utilized (23 °C in the present study), and (ii) the muscle with a larger volume than those previously investigated, such as the first dorsal interosseum (Ranatunga et al., 1987), finger flexors (Holewijn and Heus, 1992; Petrofsky and Lind, 1980) and adductor pollicis (de Ruiter and de Haan, 2000; de Ruiter et al., 1999) muscles. Although Ts was similar along muscle surface, the calculation of Tm might have underestimated the temperature of the muscle fibers positioned deeper or near the vessels. Also CV increased significantly with force, but as previously stated, CV was always lower in TmL than in TmN and TmH. As supported by previous investigations (Kimura et al., 2003; Merletti et al., 1984; Mito et al., 2007; Mucke and Heuer, 1989; Petrofsky and Lind, 1980), this finding is suggestive of an alteration of the sarcolemmal propagation properties with cooling. In particular, a longer opening time of Na+ channels and a slowing of the delayed K+ rectifying current at the cell membrane level have been indicated as the main mechanisms responsible for the reduction in the motor unit action potential propagation velocity
The effects of Tm manipulation on muscle electrical activation and isometric force production of a previously fatigued skeletal muscle have never been investigated. The main result of the present study was that after the fatiguing exercise CV always decreased to a further extent in TmL compared to the other two experimental conditions, regardless of contraction intensity or duration. This finding suggests that TmL and fatigue seem to have a combined and additional effect in decreasing sarcolemmal propagation velocity. However, this combined effect was not accompanied by Tm-induced changes in EMG RMS and maximum force production.
4.1. Preliminary considerations Given the similar Tb during the three experimental conditions, the changes in the investigated parameters retrieved in the present study can be reasonably attributed to Tm manipulation itself. The lack of differences among conditions in time to fatigue and in RPE values at the end of the fatiguing exercise is in line with previous reports (Blomstrand et al., 1985; Lannergren and Westerblad, 1988) and suggests that Tm, at least in the temperature range considered in the present study, had no significant effect on muscle fatigability and on the level of perceived exertion. Previous investigations reported a cooling-induced increase in time to fatigue in an electrically stimulated rat muscle (Segal et al., 1986] and in a voluntary activated plantar flexor muscle (Petrofsky and Lind, 1981). Both studies found cooling effects on time to fatigue only in a temperature range close to that reported to maximize muscle endurance (27–29 °C; (Edwards et al., 1972; Petrofsky and Lind,
4.2. Effect of Tm manipulation
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-1
CV (m s )
EMG RMS (%)
Before
After
160
TmN pre TmH pre
160
TmN post TmH post
140
TmL pre
140
TmL post
120
120
100
100
80
80
5
5
4
4
3
3
2
*
2
# 1
1
0
0 0
5
10
15
20
# 0
5
time (s)
10
15
20
time (s)
Fig. 5. Mean values of EMG RMS, normalised as a function of the initial value, and CV during 80% MVC (20 s), before (left column) and after (right column) fatiguing exercise, in TmN (circle), TmH (triangle) and TmL (square) conditions. ⁄P < 0.05 after vs before fatiguing exercise; #P < 0.05 among conditions.
Table 2 Linear regression analysis of CV as a function of contraction time during 80% MVC in thermo-neutral (TmN), high (TmH) and low (TmL) muscle temperature, before (left-side columns) and after (right-side columns) the fatiguing protocol. ⁄P < 0.05 vs before; #P < 0.05 vs other Tm. Before TmN TmH TmL
Slope (m s2) 0.08 ± 0.01 0.07 ± 0.01 0.06 ± 0.01
After Intercept (m s1) 3.7 ± 0.2 4.4 ± 0.3 3.5 ± 0.2#
R 0.93 ± 0.01 0.95 ± 0.02 0.95 ± 0.01
(Hicks and McComas, 1989; Hodgkin and Huxley, 1952; Merletti et al., 1984). During sustained contractions, the time course of EMG RMS was the same in all experimental conditions. This is in agreement with previous findings in TmN (Esposito et al., 1998; Krogh-Lund and Jorgensen, 1993; Linssen et al., 1993), TmH (Holewijn and Heus, 1992; Petrofsky and Laymon, 2005) and TmL (Holewijn and Heus, 1992; Petrofsky and Laymon, 2005). The increase in EMG RMS during sustained contractions of heavy intensity has been attributed to (i) changes in muscle fiber action potential and CV (De Luca, 1984); (ii) recruitment of additional motor units, so to keep the force output constant (Gamet and Maton, 1989; Linssen et al., 1993); and (iii) the synchronization of active motor units (KroghLund and Jorgensen, 1993). Throughout the sustained contraction, CV decreased with time in all experimental conditions. An alteration in the sarcolemmal action potential characteristics, due to changes in intramuscular metabolites, was alleged to explain the linear decay with time of this parameter (Arabadzhiev et al., 2010; Fitts, 1994; Merletti et al., 1992). However, CV was always significantly lower in TmL than in TmN and TmH, suggesting that muscle cooling has affected sarcolemmal propagation properties since the beginning of contraction, as witnessed also by the significantly lower intercept of the CV vs time relationship in TmL (see Table 2). Despite no previous studies reported direct CV changes due to Tm manipulation during sustained contractions of heavy intensity, a lower EMG mean frequency, a parameter strongly influenced by CV, was found
Slope (m s2) 0.10 ± 0.01⁄ 0.11 ± 0.01⁄ 0.07 ± 0.01
Intercept (m s1) 3.5 ± 0.2 4.4 ± 0.3 3.1 ± 0.3⁄,#
R 0.94 ± 0.03 0.95 ± 0.01 0.96 ± 0.01
in TmL (Petrofsky and Laymon, 2005), giving further evidence of a slowed motor unit action potential propagation with cooling. No comparisons with previous reports, though, can be made in TmH.
4.3. Combined effect of fatigue and Tm manipulation After the fatiguing exercise, MVC decreased similarly in all experimental conditions, suggesting that neither TmH nor TmL depressed to a further extent muscle force generating capacity. EMG RMS determined during MVC did not change significantly in all experimental conditions, indicating that central command was not affected by Tm manipulation or fatigue. Nevertheless, in spite of a similar muscle electrical activation, after the fatiguing exercise MVC was significantly lower, unveiling a decrease in neuromuscular efficiency, defined as the ratio of force output to the EMG amplitude (Milner-Brown et al., 1986). However, Tm manipulation did not play an additional role in this phenomenon, as already reported in the literature (Mitchell et al., 2008; Mito et al., 2007). EMG RMS was found to be similar under all experimental conditions also during brief and sustained contractions, both before and after the fatiguing exercise. Moreover, as previously reported (Esposito et al., 1998), after fatigue EMG RMS showed a tendency to an increase (P = 0.09), possibly due to the previously cited and still persisting mechanisms involved in increasing the power of the signal with fatigue, i.e., changes in the muscle fiber action potentials, decreases in CV, and synchronization/grouping of active MU.
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Under all the three Tm conditions, CV showed a significant reduction with fatigue, which might be explained by the previously described alteration of sarcolemmal propagation properties due to changes in intramuscular metabolites (altered intra/extracellular K+ concentration and increased intracellular H+ concentration) occurring with fatigue (Arabadzhiev et al., 2010; Fitts, 1994; Merletti et al., 1992). Noteworthy, the CV drop in TmL was larger than in the other conditions, and this was also the case during both brief and sustained contractions. Moreover, CV values in TmL at the beginning of the sustained contraction after the fatiguing exercise were not different from the last CV value before the fatiguing exercise. This was not the case in TmN and TmH. Collectively, these findings are suggestive of a combined and additional effect of TmL and fatigue on the sarcolemmal propagation properties. The larger CV drop in TmL, though, was not accompanied by a broader reduction in MVC. As an explanation, it might be speculated that the reduction in Ca2+ efflux from the sarcoplasmic reticulum in TmL (Caputo, 1972a,b) might have been counteracted by the increase in Ca2+ sensitivity of the thin-filament regulatory system with cooling (de Ruiter et al., 1999; Godt and Lindley, 1982; MacIntosh, 2010; Xu et al., 2003). Another possible mechanism could be a synergistic compensation to elbow flexion of the brachioradialis muscle, which was only marginally affected by cooling but contributed to the force output. Conversely, CV was measured on the belly of the biceps brachii muscle, which was directly affected by Tm manipulation procedures. Further studies, though, are needed to provide support to these hypotheses. Interestingly, whereas CV time course during sustained contraction had a similar slope in all Tm conditions before the fatiguing exercise, after fatigue the slope increased significantly in TmN and TmH, but not in TmL. Given the already lower initial CV values of TmL after fatigue, the effect of cooling on sarcolemmal propagation properties was limited and CV could not be reduced below a certain value.
5. Conclusion During both brief and sustained contractions, CV values were constantly lower in TmL than in the other Tm. After fatigue, muscle cooling decreased CV to a further extent compared to the other conditions, suggesting a combined and additional effect of cooling and fatigue in altering the sarcolemmal propagation properties. However, Tm manipulation did not affect time to fatigue and force generating capacity, neither before nor after the fatiguing exercise. A change in Tm, at least in the range of temperatures investigated in the present study, did not show detrimental effects on force output, in spite of altered sarcolemmal propagation properties. The present findings may have practical implications also for sport trainers and/or physiotherapists about muscle performance after Tm manipulation (for example after diathermy or local cryotheraphy) under both fresh and fatigued conditions.
Conflicts of interest None declared.
Acknowledgements The authors wish to thank all the participants involved in the study, for their patience and committed involvement. The study was supported by the University of Milan (PUR grant #12-15059281-42).
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Emiliano Cè was born in Italy in 1976. He received the degree in Sport Sciences from the University of Milan (Italy) in 2002. He achieved his PhD in Morphological Sciences (2007) at the University of Milan (Italy). He received the BSc in Osteopathy from the University of Wales (Wales, UK) in 2010. Present position: He is Assistant Professor at the Department of Sport, Nutrition and Health Sciences of the University of Milan (Italy). His research interests are addressed to the study of muscle biological signals, in particular force, surface electromyogram (EMG) and mechanomyogram (MMG), in muscle activity during voluntary or stimulated contractions in different physiological models (fatigue, training, temperature, etc.). Currently his scientific activity is focused on the properties of the muscle-tendon unit mechanical model and the possibility to monitor the motor unit activation strategy by the analysis of force, EMG and MMG signals. He is a member of the Italian Society of Sport Sciences (SISMES) and of the Interuniversity Institute of Myology (IIM).
Susanna Rampichini graduated in Physical Education from Istituto Superiore di Educazione Fisica di Milano, Italy in 1998 and in Biomedical Engineering from Politecnico di Milano (Italy) in 2003. She actually works at the laboratory of Exercise physiology of the Department of Sport, Nutrition and Health Sciences of the University of Milan (Italy) as an assistant of research. She is interested in the neuromuscular control and in biological signal processing. She is a member of the Italian Society of Sport Sciences (SISMES).
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Luca Agnello was born in 1983. He graduated in Sport Science at the University of Milan, Italy, in 2007. He received his PhD in Biochemistry, Physiology and Pathology of the Muscle at the Department of Basic and Applied Medical Sciences, University of Chieti, in 2011.
Eloisa Limonta was born in Milan in 1979. She graduated in Exercise Sciences at the University of Milan in 2002. She received her PhD in Sport and Physical Activity in 2007 at University of Milan and she is an Assistant Professor at the Department of Sport, Nutrition and Health Sciences since 2005. Her research field is in Exercise Physiology and Training in Sports, with special interests in electrical and mechanical aspects of muscle contraction and in physiological responses to different strength training modalities.
Arsenio Veicsteinas entered in the Institute of Physiology, School of Medicine, University of Milan to initiate his thesis in 1968 under the supervision of Rodolfo Margaria and Paolo Cerretelli. Then in 1971 received the position of Assistant Professor with Rodolfo Margaria. In 1980 he moved to the newly founded University of Brescia as Associate Professor, where he developed the Laboratory of Exercise Physiology. In 2001 returned to the Faculty of Exercise Sciences of the University of Milan as Professor of Physiology. As Visiting Professor spent several time in the Physiology Department, Max Planck Institute for Experimental Medicine, Goettingen, Germany (under the supervision of Johannes Piiper), and in the Dept. of Physiology, Yale University (John Stitt and Ethan Nadel), and SUNY at Buffalo, U.S. (Donald Rennie and David Pendergast). The reserch fields of interest cover the Physiology of Muscle Exercise (metabolism, respiratory and cardiocirculatory adjustments to exercise), the control of Muscle Contraction (electromyogram and mechanomyogram), the study of the Biological Effects of electromagnetic fields on animals and cells, and more recently the molecular mechanism involved in exercise and in cardioprotection.
Fabio Esposito was born in Vedano al Lambro (Milan, Italy) in 1963. He received his medical degree from the University of Milan in 1990 and the specialization in Sports Medicine from the University of Brescia in 1994. He joined the faculty at University of Milan in 2003 and is currently Associate Professor of Physiology. His research field is Exercise Physiology, with special interests in muscle electro-mechanical behavior, pulmonary gas exchange, physiological responses to exercise training and exercise-induced cardioprotection. He is a member of the Italian Physiological Society (SIF), of the Italian Society of Sport Sciences (SISMES) and of the American College of Sport Medicine (ACSM).