Photomedicine and Laser Surgery Volume 31, Number 12, 2013 ª Mary Ann Liebert, Inc. Pp. 586–594 DOI: 10.1089/pho.2012.3388

Effects of Low-Level Laser Therapy on Biceps Braquialis Muscle Fatigue in Young Women Renan Hideki Higashi, PT,1 Renata Luri Toma, MSc,1 Helga Tatiana Tucci, PhD,2 Cristiane Rodrigues Pedroni, PhD,3 Pryscilla Dieguez Ferreira, PT,1 Gabriel Sobrinho Baldini,1 Mariana Chaves Aveiro, PhD,2 Audrey Borghi-Silva, PhD,4 Anamaria Siriani de Oliveira, PhD,5 and Ana Claudia Muniz Renno, PhD1

Abstract

Objective: This study aims to investigate the effects of low-level laser therapy (LLLT) on biceps brachi muscular fatigue in 20 young females. Background data: Exhausting physical activity leads to muscular fatigue, which could decrease muscular strength, and may cause impairment in motor control and muscle pain. Several biochemical and biophysical resources have been studied in an attempt to accelerate the recovery of muscle fatigue. Among these, LLLT is emphasized. Methods: Twenty subjects were randomized in one laser group and one placebo group in two sessions of a crossover design experimental procedure; the second session taking place within 7 days of the first. In the first session, subjects underwent a collection of surface electromyographic (SEMG) data of the biceps brachii muscle, followed by active or placebo LLLT at the same muscle, followed then by another EMG sample of biceps brachii. Blood samples were collected five times during the experimental procedure. Second session procedures were identical to the first, with exception of LLLT, which was the opposite of the first session. The fatigue protocol consisted of 60 sec of elbow flexion-extension movement performed with 75% of one maximum repetition. Blood lactate, EMG fatigue, and the number of elbow flexion-extension repetitions during the fatigue protocol were used to evaluate the effects of laser therapy (808 nm wavelength, 100 mW output power, power density of 35.7 W/cm2, 70 sec each point and 7 J/point on eight points). Results: No statistical differences were found for eletromyographic fatigue and blood lactate values between groups. Mean numbers of elbow flexion-extension repetitions were 22.6 – 7.58 after placebo, and 25.1 – 9.89 after active LLLT group, but these differences were not statistically significant ( p = 0.342). Conclusions: LLLT had limited effects on delaying muscle fatigue in a young female sample, although a tendency was observed in the active laser group toward showing lower electromyography fatigue of biceps brachii muscle. No intergroup differences were found in the number of muscle contractions and lactate concentration. Introduction

R

epeated or exhausting muscle contractions lead to a decline in muscle performance known as ‘‘muscle fatigue,’’ which is defined as a complex and multifaceted process involving several physiological and biomechanical elements.1 Fatigue is related to the intensity of physical exercise, type of recruited fiber type, level of physical activity, age, and gender.2,3 Several mechanisms are involved in the development of fatigue, such as lack of metabolic substrates, changes in blood flow, and alterations in muscle excitation-

contraction coupling.4 Those effects may be related to the reduction of muscle strength,5 which influences the execution of movements and training performance, and also could predispose to a variety of musculoskeletal disorders.6 In this context, several studies have focused on methods that can be used to prevent or reduce muscular fatigue effects, especially in athletes.2,6,7 Among them, low-level laser therapy (LLLT) has emerged as an important resource that could interact with biological tissues, producing several physiological and/or therapeutics effects, including the enhancement of muscle performance.7–12 LLLT is absorbed by

1

Department of Biosciences, Federal University of Sa˜o Paulo, Campus Baixada Santista, Santos, SP, Brazil. Department of Human Movement Sciences, Federal University of Sa˜o Paulo, Campus Baixada Santista, Santos, SP, Brazil. 3 Department of Physical Therapy, Estadual University Paulista Ju´lio de Mesquita Filho. Campus Marı´lia, Marı´lia, SP, Brazil. 4 Department of Physical Therapy, Federal University of Sa˜o Carlos, Sa˜o Carlos, SP, Brazil. 5 Department of Biomechanics, Medicine and Rehabilitation of The Locomotor System, Faculdade de Medicina de Ribeira˜o Preto, Sa˜o Paulo University, Ribeira˜o Preto, SP, Brazil. 2

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EFFECTS OF LLLT ON MUSCLE FATIGUE chromophers and cell photoreceptors, causing modifications in cellular processes, such as excitation in the components of the mitochondrial respiratory chain.13,14 Therefore, stimulation of the respiratory chain alters the cytoplasm and mitochondria potential redox, accelerating electron exchange and consequently increasing adenosine triphosphate (ATP) production. Moreover, LLLT could increase cellular metabolism, leading to an increase in DNA, RNA, proteins, and enzymes synthesis.15–17 Studies have also demonstrated the generation of giant mitochondria through fusion of adjacent lower mitochondria, which leads to greater levels of cell respiration, and, consequently, to more energy (ATP).18,19 The effects of LLLT on bioenergetics favor the predominance of aerobic metabolism, which contributes to the increase in the availability of energy. Recent research has demonstrated that laser therapy applied before exercise significantly attenuated the serum lactate, creatine kinase (CK), and produced a faster muscle recovery after exercise sessions.6,7,10,12,20 Moreover, there is strong evidence that LLLT leads to a greater reduction in fatigue, which is relevant to rehabilitation and sports medicine.11 To date, there is no known research that studies the possible influence that LLLT could have in the biceps brachi muscle performance associating surface eletromyography, blood lactate concentration, and the number of elbow flexion-extensions in fatigue experimental procedures in a young female population. Moreover, to date there have been no known studies that performed this analysis using a crossover design, aiming to have results that could bring knowledge to the field of exercise practice studies on the possible effects on muscle fatigue recovery, if LLLT takes place before or after an exercise. Only one study was found in the literature that had a similar design, but the study was conducted on an elderly population.21 However, research has demonstrated that LLLT is considered more effective in situations in which the tissue redox potential is altered, as in the musculoskeletal system of elderly people.21,22 Moreover, it has been demonstrated that LLLT has different influences in muscular tissues related to age.10 Therefore, the results of this study could bring important findings to the field of rehabilitation and training. Based on the positive effects of LLLT on muscle metabolism, it was hypothesized in this study that LLLT would be effective in reducing electromyographic fatigue of biceps brachii, increasing the number of repetitions during elbow flexion-extension movement, with concomitantly decrease of blood lactate levels. The aim of this study was to investigate the effects of LLLT (808 nm, 250 J/cm2, 100 mW) in the biceps brachi muscular performance in young females. Therefore, to test our hypothesis, subjects underwent an experimental procedure of two session biceps brachii muscular fatigue protocol, receiving previously active or placebo LLLT. The number of elbow flexion-extension repetitions, blood lactate, and surface electromyography were evaluated. Methods Subjects Twenty healthy females participated in this study [mean age (SD) of 21.9 (1.1) years, height of 1.62 (0.05) m and body mass 58.7 (8.2) kg]. Anthropometric data are shown in Table 1. During physical evaluation, three subjects did not

587 Table 1. Means and SDs of the Anthropometric Characteristics and 1 MR Variables

Means

SD

Age (years) Body mass (kg) Height (m) 1 MR for elbow (kg)

21.9 58.7 1.62 4.2

– 1.1 – 8.2 – 0.05 – 0.5

MR, maximum repetition.

complete the second session, and one subject tested positive for shoulder impairments. Therefore, 20 subjects who met inclusions criteria were recruited to participate in the study. All subjects were informed of study purpose and procedures, and signed an informed consent form before their participation. Procedures were approved by the Ethics Committee of the Federal University of Sa˜o Paulo (Approval Number 182/10#). Twenty-four subjects were evaluated. Inclusion criteria were to be clinically healthy and female, between 18 and 25 years of age, with body mass index between 20 and 25, and classified as an active person. A person considered as ‘‘active’’ should perform physical activity with a frequency of at least five times a week, totaling a minimum of 150 min/week, according to criteria established by the International Physical Activity Questionnaire – Short Version (IPAQ). Subjects were excluded if they had any previous musculoskeletal injury or reported pain in the dominant upper extremity and shoulder girdle; or tested positive in two orthopedic tests for shoulder, elbow, and wrist injuries. Design This study was designed as a crossover, randomized, triple-blinded, placebo-controlled trial. Subjects were crossed over in a two session experimental procedure, performed 7 days apart, to compare outcomes after placebo and after active LLLT application for each participant. Thus, all subjects were subjected to the same procedures, but in a different order, according to randomization. The subject’s allocation was blinded to subjects and researchers who were responsible, by laser application and data analysis. Subjects were randomly allocated in two groups by simple drawing of lots, which determined whether they should first receive active LLLT or placebo LLLT. Group A: Subjects who received active laser during the first session and placebo laser at the second session (n = 20) Group B: Subjects who received placebo laser during the first session and at active laser the second session (n = 20) Randomization procedure was performed through a computer program that created a random table of numbers, in which each code number corresponded to a group A or B. Then, subjects were allocated according to the corresponding number of their evaluation forms. The subject’s code number was known only to researcher 1, who was responsible for preparing the laser probe as placebo or active LLLT. A second researcher (2), who was blind to subject allocation code number, was responsible for applying LLLT to the subjects. Researcher 1 was instructed not to communicate the type of

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LLLT application to either subject or researcher 2. To ensure the complete blinding of the researcher 2, that person was instructed not to remain inside the laboratory while researcher 1 prepared the laser probe and adjustment device. In addition, an adhesive tape was used to cover the specifications written in the laser probe. A third researcher, responsible for data analysis, also was blind about subject’s group.

of this research was to analyze the possible influence that LLLT could have in reducing electromygraphic fatigue of biceps brachii using a biomechanical tool. To consider that an alteration in the surface electromyographic (SEMG) signal was caused by the LLLT applied in the muscle, it was necessary to guarantee that those parameters were enough to cause it. Otherwise, correlations could not be established. Blood sample collection

Fatigue protocol Fatigue protocol followed the one designed by Leal Junior et al.,6 which evaluates the ability of a subject to perform during 60 sec as much of an elbow flexion-extension movement as possible with 75% of weight of 1 maximum repetition (MR). This fatigue protocol was chosen because we wanted to analyze the influence of LLLT on electromyographic fatigue. Therefore, it was necessary to choose a fatigue protocol previously described as adequate to cause biceps brachii fatigue. Otherwise, if the fatigue protocol used was not considered appropriate, it would not have been possible to suggest that signals indicated a fatigued muscle. Additionally, it would not have been possible to suggest that modifications in the electromyographic signal could be caused by the use of LLLT. The fatigue protocol was performed with the subject seated in a chair without an armrest, and with height adjustment that allowed hip, knee, and ankle joints to be positioned at 90 degrees of flexion. The weight of the fatigue protocol was individualized for each participant, and all subjects were instructed to perform elbow flexion-extension movements starting from elbow full extension to elbow full flexion as many times as possible during 1 min. The number of repetitions was counted by the same evaluator in all subjects, who was blind to the randomization. Laser photostimulation Subjects received both active and placebo LLLT in different sessions, according to randomization. Laser application was performed before the fatigue protocol. Laser (DMC Sa˜o Carlos, SP, Brazil) was punctually applied at eight points, properly distributed in the biceps brachii of the dominant side. The laser irradiation parameters and dose applied in the active laser protocol are demonstrated in Table 2. These laser parameters were chosen based on the results of a previous study that investigated the effects of LLLT in muscle fatigue. This choice was important, because the aim Table 2. Laser Parameters Wavelength Laser frequency Optical output Spot diameter Spot size Power density Energy Energy density Treatment time Number of points Total energy delivered Application mode probe

808 nm (infrared) Continuous output 100 mW 0.6 mm 0.0028 cm2 35.7 W/cm2 7 J each point 250 J/cm2 70 sec at each point 8 56 J Stationary in skin contact mode

Blood samples were collected in the second and third sessions of this study, and in each session, four samples were obtained at the following times: at rest, immediately after the fatigue protocol, and 3 and 6 min after the fatigue protocol (as previously described). Blood samples were collected from the earlobe using a disposable lancet. The first drop of blood was discarded to avoid contamination and then, 25 lL of arterial blood were collected using a calibrated and heparinized glass capillary. Samples were placed in tubes containing 50 lL of sodium fluoride at 1% to prevent the continuation of glycolysis, and stored in a freezer at - 10l for further analysis. Eletromyography sampling SEMG signals were sampled using equipment with a 12 bit analog-to-digital (A/D) with converter board with a 4 kHz frequency, and band-pass filtered at 0.01–1.5 kHz (DataHomins Ltda, Uberlaˆndia, Minas Gerais, Brazil). For capturing biceps brachii mioelectrical surface signal, the equipment used was a surface differential electrode with two bars of Ag-AgCl, with 10 mm interelectrode distance, gain of 20, input impedance of 10 GO, and a common mode rejection ratio of 130 dB (EMG System Ltda, Sa˜o Jose´ dos Campos, Sa˜o Paulo, Brazil). The electrode was placed one third from the line between the medial acromion and the fossa cubit. A circular electrode (3 cm2) attached at the sternal notch was used as a reference electrode. The skin at the electrodes’ sites was shaved and cleaned with alcohol before the electrodes were attached, to reduce skin impedance and to achieve good fixation. Electrodes were fixed using adhesive tape. Electromyography signals were sampled in the second and third sessions, as described in the experimental procedures. Experimental procedures Experimental procedures consisted of 1 day for physical evaluation and two sessions of data collection. On the first day, the value of 1 MR for elbow flexion-extension exercise was obtained. Before 1 MR procedure, all subjects were familiarized with the exercise, being instructed to perform one single total flexion from full elbow extension and ending at the initial position. Then, 1 MR test was performed with the subject sitting in a chair without an armrest and with adjustable height, which allowed the subject to be seated with hips, knees, and ankle joints in 90 degrees of flexion. Free weights were used to determine the weight that corresponded to the maximum load lift by the subject in only one repetition. Up to five attempts of determining 1 MR for a subject were considered on 1 day, and if weight was not determined, the test was rescheduled to avoiding muscle fatigue influence. All procedures were based on the American College of Sports Medicine Guidelines.23 The second

EFFECTS OF LLLT ON MUSCLE FATIGUE session was performed 48 h after 1 MR weight determination. The first experimental session was performed 48 h after 1 MR determination. The blood lactate over time, and SEMG signal in both groups. were collected in this session. Data collecting was composed of the following steps, in the order in which they are described: (1) rest blood sample was collected; (2) in sequence, biceps brachii SEMG was sampled during 5 sec of maximal voluntary isometric contraction (MVIC); (3) after 1 min, biceps brachii SEMG was sampled during 30 sec of MVIC; (4) in sequence, active or placebo laser application was done according to the randomization; (5) immediately after, biceps brachii fatigue protocol was performed during 60 sec with work load of 75% of 1 MR; (6) blood sample was collected immediately after fatigue protocol; (7) blood sample was collected 3 min after fatigue protocol; (8) blood sample was collected after 6 min of fatigue protocol; and (9) 30 sec of MVIC EMG was sampled. The sessions are illustrated in Fig. 1. After 1 week, subjects performed the second session, which consisted of the same procedures as the first session, with only laser therapy changed according to previous randomization. The dominant arm was tested for all subjects, and the fatigue protocol was performed unilaterally with 75% of 1 MR as previously described.23 Subjects performed MVIC with the arm in a neutral position, 90 degrees of elbow flexion, and with the forearm in supination. Proper attention was given to obtaining standardization in the subject’s positioning in the fatigue protocol performance. Data were collected at the same time of the day to control the circadian rhythm. Data analysis SEMG sampling and digital signal processing were performed using Myosystem Br-1 software. Raw SEMG data were digitally filtered at a frequency bandwidth of 10– 500 Hz and the median frequency (MDF) of the signal was used for analysis. The MDF obtained from 5 sec MCIV was used for subsequent normalization of EMG data obtained in both 30 sec MCIV contractions. The 30 sec MCIVs sampled before and after the fatigue protocol were used to analyze biceps brachii muscular fatigue. Therefore, each 30 sec MVIC was divided into three windows. The first window corresponded to the value obtained from the beginning of the sampled signal (0–3 sec), the second to the middle of the sampled signal (15–18 sec), and the last to the end of the sampled signal (27–30 sec). Each one of those windows was normalized by dividing the MDF from each one by the MDF

FIG. 1. Flow chart timeline of the study.

589 obtained from the 5 sec MCIV. Normalized values obtained from the three windows of 30 sec MCIV were used to calculate the slope of the curve for both the laser and placebo groups. Therefore, for each group, two slopes were calculated, one obtained from 30 sec MVIC sampled before fatigue protocol (slope 1) and other obtained from 30 sec MVIC sampled after fatigue protocol (slope 2). Slope 1 was compared with slope 2 to obtain results from electromyographic fatigue of biceps brachii muscle pre- and post-LLLT application. Slope comparison was done for active and placebo groups. Therefore, the slope coefficient was the summary measure of MDF. Blood samples were analyzed using the Yellow Springs Instrument—YSI 1500 Sports Lactate Analyzer (Yellow Springs, OH), which was calibrated according to the manufacturer’s instructions before each analysis. Procedures followed methods performed by Simo˜es et al.24 Statistical analysis Statistical analyses were performed using Statistica software (StatSoft Inc., Tulsa, OK). To evaluate the influence of order of laser application (active or placebo) and to determine the slope of the curve, a model of analysis of variance (ANOVA) with repeated measures crossover experiments was used. For the variable number of repetition, lactate blood level and SEMG measurements, nonparametric tests were performed, because they did not present a normal distribution verified by Shapiro–Wilks test. The intragroup analysis was conducted using Friedman ANOVA and Wilcoxon nonparametric tests. Intergroup analysis was performed using the Mann–Whitney U nonparametric test. When the main effect was significant, pairwise comparisons were performed using Bonferroni adjustment for multiple comparisons. The level of significance used for all comparisons was 5% ( p £ 0.05). Results Anthropometric data description and values obtained from 1 MR test are described in Table 1. The mean number of elbow flexion-extension repetitions performed in the fatigue protocol was 22.6 – 7.58, when subjects previously received placebo LLLT, and 25.1 – 9.89 when subjects previously received active LLLT before fatigue protocol (Fig. 2). However,

FIG. 2. Number of repetitions performed in the fatigue protocol.

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HIGASHI ET AL. active LLLT ( p = 0.968) (Fig. 4). Therefore, for this experimental procedure, LLLT has no statistical significance on the EMG fatigue of biceps brachii muscle, even though the placebo group had a lower value when compared with the active group. Additionally, results showed that no statistical difference was found among groups when values of slope (obtained from the three windows of normalized MDF over time) were compared ( p = 0.133). However, it was possible to observe a decrease in slope values over time in both groups that were lower in the placebo group if compared with the active laser group (Fig. 5). Therefore, results suggest that in spite of no statistical significance having been observed, it is possible to propose the influence of LLLT in reducing EMG fatigue. Discussion FIG. 3. Blood lactate levels (*p < 0.001).

significant differences were not found when the number of repetitions was compared between the placebo and active laser groups ( p = 0.342). Figure 3 shows blood lactate results over time in both groups. Results showed that basal lactate evaluation, for both experimental conditions, was significantly lower compared to the post-exercise lactate measurements. In the active laser group, a significantly lower lactate measurement was found 6 min after the exercise ( p = 0.0035). However, the placebo and active groups did not present significant intergroup differences for lactate levels measured immediately after fatigue protocol 3 and 6 min after (0.23, 0.19, 0.11, p values, respectively). Results demonstrated that there was no significant difference in the slope values obtained from normalized MDF between the two experimental conditions, that is, placebo or

The present study investigated the effects of LLLT in biceps brachii induced muscle fatigue in young women. It was hypothesized that LLLT would have an interaction with muscular tissue, which may provide more energy supply, improving the muscle performance and delaying the EMG fatigue. In this study, the order of laser application has not influenced the subject’s performance during fatigue protocol, which justifies the crossover design of this study. The main findings demonstrated that the number of repetitions was not different in placebo and active groups. In addition, results also showed no difference in the EMG fatigue and lactate blood removal between groups. There are four suggestive results that could indicate that active LLLT influenced muscle performance improvement in this research, although no significance was found. The first indication of this improvement is the fact that the active laser group showed greater slope values for SEMG analysis, indicating a tendency of lower muscular fatigue after laser irradiation. The second indicative is the improvement of 11.1% in the number of elbow flexion-extensions in the active group

FIG. 4. Mean profile for the variable ‘‘slope.’’

EFFECTS OF LLLT ON MUSCLE FATIGUE

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FIG. 5. Median frequency (MDF) over time.

compared with the placebo group. The third indication is that lactate levels in the active group immediately after 3 and 6 min post-fatigue protocol had a tendency toward presenting lower values than the placebo group. Finally, it is important to point out that the lack of significant effect of LLLT in EMG results may be related to the difference in the number of repetitions and workload in the fatigue protocol among groups. As the laser group’s number of repetitions and workload was greater if compared with the placebo group, post-fatigue protocol baseline for EMG also could be different between groups. Therefore, it may negatively have affected the 30 sec MVIC EMG signal sampled after fatigue protocol for the laser group. The effects of LLLT on delaying fatigue onset and increase of muscle performance have been demonstrated by some authors.7–12 However, the results found in the literature are still controversial, and parameters are not defined. In a similar study investigating the effects of LLLT cluster probe with five laser diodes (810 nm; 200 mW; 30 sec of irradiation; 60 J of total in muscle fatigue) in volleyball players, it was observed that LLLT produced a significant increase in the number of muscle repetitions, whereas no effect in blood lactate concentration was demonstrated.7 Nonetheless, De Marchi et al.8 observed a decrease in lactate concentration in untrained men irradiated with 810 nm. 200 mW, 30 J in each site, and demonstrated the benefits of LLLT irradiation before progressive-intensity running exercise. Baroni et al.9 observed that LEDT irradiation (34 red diodes of 660 nm; 10mW; 35 infrared diodes of 850 nm; 30mW) before knee extensor eccentric exercise in healthy men did not demonstrate an increase in exercise performance, although it was effective in terms of attenuating the decrease in muscle force. Toma et al.21 showed that LLLT irradiation (808 nm wavelength; 7 J/point; totaling 56 J) increased the number of knee flexion-extension repetitions in a fatigue protocol in elderly people, and did not demonstrate significant difference in EMG fatigue. In a study evaluating the effects during strength training and LLLT protocol using an infrared laser 808 nm with six diodes of 60 mW each and a total energy of 50.4 J, the group irradiated presented an increase in muscle performance in an isokinetic dynamometry test; however, it

did not demonstrate effects in the thigh perimeter.25 Another similar study observed an endurance training protocol in women involving cycle ergometer exercise with load applied to the ventilatory threshold (VT) and LLLT irradiation 808 nm, with six 60 mW diodes, 0.6 J/diode, and a total energy applied of 18 J, suggests that LLLT leads to a greater reduction in fatigue than an endurance training program without LLLT.11 Therefore, it is observed that the disparity between results is related to differences in the experimental conditions, such as the use of different fatigue protocols, different populations, different methods of evaluation, and the wide range of laser devices and parameters used by authors, such as power, number of irradiated points, and energy per point.2,6–8 Adding to this, there is a lack of studies evaluating the acute effects in non-athlete women. Furthermore, the parameters used in this study are reproducible in clinical practice because a standard LLLT device was used as opposed to a cluster or LEDT. The costs of the devices used in this study are greater than for the ones used by other authors, and furthermore, the devices here employed are not widely found in clinical practice. Research about the effects of LLLT on muscle fatigue is an important area of investigation. In this context, blood analysis has been widely used as a reliable tool in studies reported in the literature.2,6–8 The positive effects of LLLT on circulation have been evidenced by some authors.7,8,20,26,27 It was reported that infrared laser irradiation led to an increase in arterial blood flow in rats, and increased the superficial blood flow in foot soles of healthy individuals. These changes in blood flow may be related to positive results such as healing improvement and pain reduction.28 In the present study, the absence of statistical difference in the number of elbow flexion-extension repetitions and in lactate levels in intergroup comparisons did not corroborate the results found in the literature.2,6,7,20,21 A possible justification for these different results may be the existence of a curve-dose response related to the parameters of irradiation.29,30 In this context, the hypothesis is that the laser energy offered to the muscle tissue during the experimental procedure was not efficient to produce an improvement in the muscle performance. Finally, a justification about our

592 results does not corroborate those found by Toma et al., which relates to the different effects that LLLT could have in the muscular tissue of the elderly and youth.21 Studies demonstrate that the biological action of LLLT reaches an optimum effect in situations in which the redox potential of tissues are altered, as in the presence of injuries or in the musculoskeletal system of elderly people.10,31 The present study also analyzed the possible effects of LLLT in the EMG fatigue of biceps brachii, associating the results with those obtained from blood lactate level and muscle performance. This related information would help the comprehension of muscular and metabolic changes caused by active laser irradiation. In this study, infrared wavelength and mean energy per point of 7 J was chosen based on the efficacy of these parameters observed in previous studies.7–9,21 We hypothesized that if such doses were considered effective by other authors for removal of blood lactate and improving muscular performance, the same parameters could also be effective for reducing EMG fatigue. As previously described in the laser therapy session, those parameters were chosen because our aim was to analyze the effects of laser theraphy on the electromyography signal, and not to test a specific laser therapy parameter. Therefore, we needed to choose a a dose previously considered adequate to reduce biceps brachii fatigue. Otherwise, if significant effects had been found in EMG signal, it would not have been possible to suggest that it was caused by LLLT effects. Results of electromyography fatigue evaluation showed lower values of slope coefficient after fatigue protocol in the placebo than in the active laser group. Although this difference was not significant, it could indicate a tendency that fatigue occurred earlier when volunteers were irradiated with placebo compared with the active laser. Toma et al. also found this tendency on electromyography fatigue delay in the quadriceps femoris muscle of elderly women.21 These findings may be related to laser effects on blood flow that leads to an increase in microcirculation, allowing metabolite clearance.22 Considering the results of this study, it seems that LLLT had limited effects on delaying muscle fatigue in a young female sample, although a tendency toward lower EMG fatigue of biceps brachii muscle was observed in the laser group. Some hypotheses can be raised attempting to explain the present findings. Perhaps the parameters used were not sufficient to produce structural changes in mitochondria, which would directly increase the ATP synthesis and accelerate cell metabolism.32 Additionally, the laser energy offered to the tissue may not be able to increase the amounts of alactic energy (phosphocreatine) during muscle contraction and, consequently, the conversion of pyruvate to lactate in the mitochondrial matrix of muscle cells, which would result in a lower lactate accumulation and increased energy availability during exercise.25 Moreover, the characterization of the population studied may have contributed to these results. It is well known that the biological action of LLLT reaches an optimum effect in situations in which the redox potential of the tissues is altered, as in the presence of injuries or in elderly people.10,31 Some considerations of our findings may be limited by the experimental model, which only measured the immediate effects of LLLT on muscle performance. Although laser parameters were similar to the ones used in previous research, there were no significant differences in the number of repe-

HIGASHI ET AL. titions and in the reduction of electromyography fatigue. Some possible explanations for the lack of difference would be the reduced sample, the fatigue protocol, and also the laser therapy parameters. Furthermore, despite SEMG fatigue having been used to characterize peripheral muscle during isometric contractions,33–39 the method proposed to analyze electromyography fatigue using isometric contraction may not have been the most adequate for this type of experimental procedure, which could have been unfavorable for the analysis of muscle performance. However, the present study followed the standards of a pioneer study in this field21 which established standards for reporting EMG data according to the International Society of Electrophysiology and Kinesiology.40 The procedure was designed equally for all volunteers in order to avoid bias in the experiment’s aim of evaluating the possibility of having more muscular work in the laser group. Finally, laser parameters were similar to those used in previous studies that obtained a positive effect of LLLT in reducing muscular fatigue. However, these parameters may not have been effective for delaying electromyography fatigue in a sample of young women. We believe that the dosage used was not sufficient to produce significant differences between the groups, showing that the question of dosimetry should be further investigated. Conclusions In this study, LLLT had limited effects on delaying muscle fatigue in a young female sample, although a tendency toward showing lower electromyography fatigue of biceps brachii muscle was observed in the active laser group. Furthermore, no intergroup differences were found in the number of muscle contractions and in lactate concentration. Further studies are necessary to elucidate the interaction between LLLT and the myoeletrical responses in young women. Acknowledgments The authors thank Mariane Stanev from the English Department, Florida International University, and the research subjects. Author Disclosure Statement No competing financial interests exist. References 1. Ivey, F.M., Roth, S.M., Ferrell, R.E., et al. (2000). Effects of age, gender, and myostatin genotype on the hypertrophic response to heavy resistance strength training. J. Gerontol. A. Biol. Sci. Med. Sci. 55, 641–648. 2. Leal Junior, E.C., Lopes-Martins, R.B., Dalan, F., et al. (2008). Effect of 655-nm low level laser therapy (LLLT) on exerciseinduced skeletal muscle fatigue in humans. Photomed. Laser Surg. 26, 419–424. 3. Fitts, R.H. (1993). Cellular mechanisms of muscular fatigue. Physiol. Rev. 74, 49–94. 4. Enoka, R.M., and Stuart, D.G. (1992). Neurobiology of muscle fatigue. J. Appl. Physiol. 72, 1631–1648. 5. Rahnama, N., Reilly, T., Lees, A., Graham–Smith, P. (2003). Muscle fatigue induced by exercise simulating the work rate of competitive soccer. J. Sports Sci. 21, 933–942.

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HIGASHI ET AL. Address correspondence to: Ana Claudia Muniz Renno Department of Biosciences Federal University of Sa˜o Paulo Campus Baixada Santista Av. Ana Costa, 95 11060-001, Santos, SP Brazil E-mail: [email protected]