Posttetanic Potentiation in Knee Extensors After High-Frequency Submaximal Percutaneous Electrical Stimulation

J Sport Rehabil. 2005;14:248-257. 2005 Human Kinetics, Inc. Posttetanic Potentiation in Knee Extensors After High-Frequency Submaximal Percutaneous Electrical Stimulation Bernardo Requena, Jaan Ereline, Helena Gapeyeva, and Mati Pääsuke Context: The understanding of posttetanic potentiation (PTP) in human muscles induced by percutaneous electrical stimulation (PES) is important for effective application of electrical stimulation in rehabilitation. Objective: To examine the effect of 7-second high-frequency (100-Hz) submaximal (25% of maximal voluntary contraction force) direct PES on contractile characteristics of the knee-extensor (KE) muscles. Design: Single-group repeated measures. Setting: Kinesiology laboratory. Subjects: 13 healthy men age 18 27 years. Measurement: Peak force (PF), maximal rates of force development (RFD) and relaxation (RR) of supramaximal twitch, and PF of doublet and 10-Hz tetanic contractions before and after direct tetanic PES. Results: A significant potentiation of twitch, doublet, and 10-Hz tetanic-contraction PF has been observed at 1 5 minutes posttetanic. Twitch RFD and RR were markedly potentiated throughout the 10-minute posttetanic period. Conclusions: A brief high-frequency submaximal tetanic PES induces PTP in KE muscles associated with small increase at 1 5 minutes. Key Words: knee extensors, electrical stimulation, contractile properties, posttetanic potentiation Neuromuscular electrical stimulation has often been used to prevent loss of muscle function or to restore muscle function after injuries and as a strengthening modality in healthy subjects and highly trained athletes. 1,2 It is commonly known that indirect or direct percutaneous electrical stimulation (PES) affects the contractile properties of skeletal muscles. 3-5 This induced activation might evoke muscle fatigue but might also result in increased muscle-force production (potentiation). 6 Potentiation induced by electrical stimulation can be defined as staircase or posttetanic potentiation (PTP). 3 Staircase potentiation occurs with low-frequency stimulation of the muscle, during which the force gradually increases, whereas PTP is the increase in muscle forces after repetitive tetanic stimulation. 3,4,7 In addition to potentiation caused by electrically evoked contractions, maximal voluntary contraction (MVC) also creates potentiation that is defined as postactivation potentiation. 8-10 Muscles are Requena is with the Dept of Physical Education and Sport, University of Granada, Granada, Spain. Ereline, Gapeyeva, and Pääsuke are with the Institute of Exercise Biology and Physiotherapy, University of Tartu, Tartu 51014 Estonia. 248

Posttetanic Potentiation After Electrostimulation 249 often activated through the use of electrical stimulation in rehabilitation. For such applications it is essential to know how muscle-contractile properties change during activation and how previous stimulation influences muscle-force production. PTP is greatest immediately after the tetanic electrical stimulation and then decays rapidly but is still evident for approximately 5 minutes. 4,5 It is often associated with increased peak force, maximal rates of force development, and relaxation of supramaximal isometric twitch, 5,11 as well as shortening of twitch-contraction and half-relaxation times. 5,12,13 Thus, preceding activation history influences both muscle-force generation and the time course of muscle contraction. The principal mechanism of PTP is commonly believed to be phosphorylation of myosinregulatory light chains during tetanic contraction, which renders actin myosin more sensitive to Ca 2+ in subsequent contraction. 11,14-16 PTP has been shown in a variety of human muscles including small hand muscles, 17 elbow flexors, 18 knee extensors, 4,7 and ankle dorsiflexors. 5 Muscles with the shortest twitch-contraction and half-relaxation times and highest proportion of fast-twitch fibers show the greatest PTP. 5,19 The magnitude of PTP is influenced by the methods and conditions under which it is evoked. PTP is affected by the intensity, frequency, and duration of the conditioning tetanic stimulation and by the total number of pulses. 7,11 A brief indirect supramaximal PES at high frequency causes the greatest immediate PTP. 20 Supramaximal indirect or direct PES could, however, potentially induce muscle injury, pain, or discomfort. 21 Submaximal direct PES has often been used in physical therapy to prevent the atrophy and strength loss associated with athletic injuries. 1 In recent years, electromyostimulation training with brief high-frequency submaximal direct tetanic PES has been used by athletes in the context of training programs to develop physical performance. 22 The phenomenon of PTP in different human skeletal muscles after submaximal high- or low-frequency direct PES has been previously investigated. 5,7,12,18 These studies are difficult to compare, however, because they have used different muscle groups and electrical-stimulation protocols. The development of PTP in human muscles after brief high-frequency submaximal direct PES is not fully understood. The aim of the present study was to examine the development of PTP in kneeextensor muscles after a brief (7-second) high-frequency (100-Hz) submaximal (25% MVC) direct PES. A similar PES has often been used in rehabilitation and electromyostimulation training programs. We monitored PTP by measuring the changes in supramaximal isometric-twitch, doublet (induced by paired stimuli with an interval of 10 milliseconds), and 10-Hz tetanic-contraction (with duration of 1 second) peak force and twitch maximal rates of force development and relaxation immediately and for several minutes after direct tetanic PES. Methods Subjects Thirteen healthy men (mean age 21.6 ± 0.8 years, height 180.7 ± 2.1 cm, body mass 73.9 ± 2.5 kg) volunteered to participate in the present study. They were physically active students with no history of neuromuscular disorders. After a routine medical examination, an informed written consent to participate was obtained from

250 Requena et al each. During the 14 days before the study, the subjects were familiarized with the experimental setup. The study was approved by the university ethics committee for human studies. Measurement On reporting to the laboratory, each subject sat resting for approximately 30 minutes before commencing the experiment to minimize any potentiation effect from walking to the laboratory. During the experiment the subject sat on the custom-made dynamometer with the knee and hip angles equal to 90 and 110, respectively. 23 The body position of the subject was secured by Velcro belts placed over the chest, hip, and thigh. The unilateral knee-extension isometric force of the dominant leg was recorded by a standard strain-gauge transducer (1778 DST-02, Russia) mounted inside a metal frame, which was placed around the distal part of the ankle above the malleoli using a Velcro belt. The subject s dominant leg was determined based on kicking preference. The electrical signals from the strain-gauge transducer were digitally low-passed (5 Hz), and the resulting curve was then differentiated to obtain the maximal rates of force development (df/dt) and relaxation ( df/dt) using a personal computer. The digitized signals were stored on a hard disk for further analysis by custom-written software. To assess the contractile characteristics of knee-extensor muscles, electrically evoked isometric twitch, doublet, and 10-Hz tetanic contractions were elicited by supramaximal percutaneous nerve stimulation. Before the stimulating electrodes were applied, electrode gel was applied to the contact surface, and the underlying skin was prepared by shaving and cleaning with isopropyl alcohol. Two 2-mm-thick, self-adhesive stimulating electrodes (Medicompex SA, Ecublens, Switzerland) were used the cathode (5 5 cm) placed on the skin over the femoral nerve in the inguinal crease and the anode (5 10 cm) placed over the midportion of the thigh. The electrical stimuli were rectangular voltage pulses of 1-millisecond duration applied at supramaximal intensity (130 150 V) delivered from an isolated voltage stimulator (Medicor MG-440, Hungary). To determine the supramaximal stimulation intensity, the voltage of the rectangular electrical pulse was progressively increased to obtain a plateau in the twitch force, that is, when twitch force failed to increase despite additional increases in stimulation intensity. The same stimulation intensity (~20% greater than that needed for maximal twitch response) was further used for twitches, doublets (with interstimulus interval of 10 milliseconds), and 10- Hz tetanic contractions (with duration of 1 second) evoked before the conditioning tetanic stimulation and during the recovery period. The following characteristics of supramaximal isometric twitch were calculated: peak force, the highest value of isometric force production; maximal rate of force development, the first derivate of the development of force (+df/dt); and maximal rate of relaxation as the first derivate of the decline of force ( df/dt). Peak force for supramaximal doublet and 10-Hz tetanic contractions was calculated as the highest value of isometric force production during doublet and unfused tetanus, respectively. Two supramaximal single twitches, doublets, and 10-Hz tetanic contractions were provoked in relaxed knee-extensor muscles with 3-second intervals between stimulations before and after a conditioning 7-second submaximal highfrequency direct PES.

Posttetanic Potentiation After Electrostimulation 251 Five minutes after the pretetanic testing of contractile characteristics had been established, MVC force of the knee-extensor muscles was measured. Each subject was asked to exert maximum voluntary isometric knee extension against the belt of the strain-gauge transducer as forcefully as possible for approximately 3 seconds. Three maximal attempts were recorded, and the best result was taken for further analysis. A rest period of 2 minutes was allowed between attempts. Two minutes after MVC-force testing, direct tetanic PES voltage for target level of force at 25% MVC of the knee-extensor muscles was determined and controlled by 2 separated stimulations of 2-second duration. A portable batterypowered stimulator (Compex Sport 400, Medicompex SA, Ecublens, Switzerland) was used. Three 2-mm-thick, self-adhesive electrodes were placed over the thigh. The positive electrodes (5 5 cm), which had membrane-depolarizing properties, were placed on the motor-point area of the vastus lateralis and vastus medialis muscles and near the proximal insertion of each muscle. The negative electrode (5 10 cm) was placed over the proximal portion of the thigh between stimulating electrodes to measure muscle-contractile properties. Rectangular voltage pulses of 0.4-millisecond duration at the frequency of 100 Hz were used. The stimulation voltage was calculated for each subject before the testing, according to individual force response. After 15 minutes of rest, the submaximal tetanic contraction (approximately 25% MVC) of the knee-extensor muscles of 7-second duration was evoked by direct PES. After the end of direct tetanic PES, the subject remained seated without moving the legs for a recovery period of 10 minutes. The posttetanic supramaximal isometric twitch, doublet, and 10-Hz tetanic contractions were evoked at 2 seconds and 1, 3, 5 and 10 minutes. The decrease in force during 7-second direct tetanic PES was determined. Statistical Analysis Data are means and standard errors of the mean (± SEM). A repeated-measures analysis of variance (ANOVA) was used to test whether the tetanic PES changed the contractile characteristics at the various time points after stimulation. To determine whether there were significant differences from pretetanic values, ANOVA was performed on the measures expressed in absolute units (eg, N, N/s). To determine significant differences between various contractile characteristics, ANOVA was conducted on the posttetanic values normalized as a percentage of the pretetanic value (pretetanic value = 100%). When significant main effect or interactions were found, a Tukey post hoc procedure was used to test differences among mean values. A level of P <.05 was selected to indicate statistical significance. Results The mean pretetanic values of twitch, doublet, and 10-Hz tetanic-contraction peak force were 63.8 ± 3.5, 113.8 ± 22.4, and 96.9 ± 6.9 N, respectively. The mean values of twitch maximal rates of force development and relaxation were 526.1 ± 2.7 and 265.5 ± 18.7 N/s, respectively. Tetanic force decreased by 21% (P <.05) during a 7-second direct PES. Figure 1 shows the mean relative potentiation of supramaximal isometric twitch, doublet, and 10-Hz tetanic-contraction peak force after direct submaximal

252 Requena et al Figure 1 Changes in isometric-twitch, doublet, and 10-Hz tetanic-contraction peak force (PF) of the knee-extensor muscles after a brief direct tetanic submaximal percutaneous electrical stimulation (PES). Data are means ± SEMs presented as a percentage of pretetanic value. *Significantly different (P <.05) from pretetanic value. #Significantly different (P <.05) from doublet contraction value at designated time point. tetanic PES. Immediately after the end of applied direct tetanic PES, the supramaximal 10-Hz tetanic-contraction peak force was significantly potentiated (16%, P <.05), whereas twitch and doublet contraction peak force did not potentiate significantly (12% and 7%, respectively, P >.05). The potentiation of supramaximal 10-Hz tetanic-contraction peak force significantly exceeded (P <.05) potentiation of the doublet contraction peak force. A significant (P <.05) potentiation of twitch, doublet, and 10-Hz tetanic-contraction peak force has been observed at 1, 3, and 5 minutes posttetanic. The greatest potentiation of the supramaximal 10-Hz tetanic-contraction peak force has been observed at 1 minute, and potentiation of twitch and doublet contraction peak force, at 3 minutes posttetanic (20%, 17%, and 13%, respectively). Twitch peak force was significantly (P <.05) potentiated at 10 minutes posttetanic, whereas doublet and 10-Hz tetanic-contraction peak force did not potentiate significantly (P >.05). There were no significant differences (P >.05) in potentiation of twitch, doublet, and 10-Hz tetanic-contraction peak force at 1, 3, 5 and 10 minutes posttetanic. Figure 2 shows the mean relative potentiation of supramaximal isometric twitch-contraction maximal rates of force development and relaxation after direct submaximal tetanic PES. Twitch maximal rates of force development and relaxation were significantly potentiated (P <.05) immediately after direct tetanic PES (29% and 26%, respectively). The potentiation was significant (P <.05) throughout the 10-minute posttetanic period, and the greatest potentiations of twitch maximal rate

Posttetanic Potentiation After Electrostimulation 253 Figure 2 Changes in isometric-twitch maximal rates of force development (RFD) and relaxation (RR) of the knee-extensor muscles after a brief direct tetanic submaximal percutaneous electrical stimulation. Data are means ± SEMs presented as a percentage of pretetanic value. *Significantly different (P <.05) from pretetanic value. of force development (38%) and maximal rate of relaxation (32%) were observed at 3 and 5 minutes posttetanic, respectively. There were no significant differences (P >.05) between relative potentiation of twitch maximal rate of force development and maximal rate of relaxation throughout the 10-minute posttetanic period. Comments The present study indicated that PTP in knee-extensor muscles after a 7-second submaximal tetanic PES at 100 Hz was associated with a significant increase in supramaximal twitch, doublet, and 10-Hz tetanic-contraction peak force at 1, 3, and 5 minutes, whereas immediately after tetanic PES twitch and doublet contraction the peak force did not potentiate significantly. The potentiation of the twitch maximal rates of force development and relaxation was significant throughout the 10-minute posttetanic period, with small increases at 3 and 5 minutes. The twitch potentiation we observed in knee-extensor muscles with submaximal direct PES, however, was less than the potentiation induced by MVCs in knee-extensor muscles 8,9,24 and by supramaximal indirect PES in dorsiflexor muscles. 5 The present results indicated that the decay in PTP from the immediate posttetanic value was not a simple exponent function, as sometimes observed. 4,18,19 In this study, PTP after percutaneous direct submaximal tetanic stimulation showed small increases at 1, 3, and 5 minutes followed by a small decrease at 10 minutes. O Leary et al 5

254 Requena et al observed that in dorsiflexor muscles PTP after a 7-second indirect supramaximal tetanic stimulation at the frequency of 100 Hz declined over the first minute but then showed a small increase at 2 minutes before it decreased again. The potentiation of twitch peak force and maximal rates of force development and relaxation were maximal immediately after supramaximal tetanic stimulation. This triphasic pattern of decay after MVCs has been shown by several investigators. 12,25,26 It has been suggested that an increase in twitch potentiation is caused particularly by fatigue; as fatigue wanes, the level of potentiation increases before falling away. 26 Fatigue might have been a factor in the present study, because tetanic force decreased by 21% (P <.05) during the 7-second direct submaximal PES. In a study by O Leary et al, 5 tetanic force declined by 15% during a 7-second supramaximal indirect PES in dorsiflexor muscles. Direct PES evokes action potentials in intramuscular nerve branches generating force directly by activation of motor axons. It is well known that during direct PES the current is applied extracellularly to the nerve endings with preferential activation of the larger fast-twitch (type II) muscle fibers. These fast-twitch fibers have larger axons with much lower electrical resistance for a given externally applied electrical current. Fast-twitch muscle fibers show greater potentiation 8 but are more susceptible to fatigue. The fatigability of preferentially activated fast-twitch fibers is one possible explanation for the marked decline in submaximal tetanic force during a 7-second direct PES at 100 Hz observed in the present study. The asynchronous and orderly (slow to fast) recruitment of motor units that occurs during voluntary activation is absent during direct PES. This lack of asynchrony and orderly recruitment contributes to the increased fatigability observed with electrical stimulation when compared with voluntary contraction. Darques et al 27 indicated that tetanic-force failure during electrostimulation at the frequency of 100 Hz results in an impaired propagation of muscle-action potentials with no metabolic changes. No significant changes have been shown, however, in M-wave amplitude during a 7-second supramaximal indirect PES at 100 Hz in dorsiflexors. 5 Our results indicated that immediately after the end of direct PES the potentiation of supramaximal 10-Hz tetanic-contraction peak force markedly exceeded the potentiation of doublet-contraction peak force, whereas no significant differences have been observed in relative potentiation of these characteristics at 1, 3, 5, and 10 minutes posttetanic. Doublets were evoked with an interstimulus interval of 10 milliseconds, that is, with the stimulation frequency of 100 Hz. These facts suggest that the PTP assessed by low-frequency supramaximal indirect activation immediately after direct submaximal tetanic PES is less affected by fatigue than is PTP assessed by high-frequency activation. The mechanism of PTP involves excitation contraction coupling and/or myosin actin interaction, rather than amplified excitation of muscle, that is, enlarged muscle-action potential. 4,11 Potentiation is caused by phosphorylation of the regulatory light chains of myosin, a Ca 2+ -dependent process. 14,15 It has been shown, however, that the muscle compound action potential (M wave) increased sharply at 2 minutes after high-frequency tetanic stimulation and then subsided. 5 The M-wave amplitude might also enlarge after low-frequency tetanic stimulation or brief MVCs. 8,28,29 The mechanism of M-wave potentiation results from the stimulation of the sarcolemma s Na + -K + -pumping mechanism. 29 A large muscle-action potential might increase Ca 2+ release from the sarcoplasmic reticulum, thereby increasing force.

Posttetanic Potentiation After Electrostimulation 255 The present study indicated that after tetanic stimulation a relative potentiation of twitch maximal rates of force development and relaxation was greater than the potentiation of twitch peak force (with peak values of 38%, 32%, and 17%, respectively). This is in agreement with O Leary et al, 5 who suggested a greater potentiation of twitch maximal rates of force development and relaxation (75% and 71%, respectively) in dorsiflexor muscles after a 7-second supramaximal indirect PES at 100 Hz as compared with potentiation of twitch peak force (50%). Thus, the twitch maximal rates of force development and relaxation are more sensitive indicators of PTP than twitch peak force is. The rate of force development has rarely been used as an indicator of muscle-contraction speed and probably depends largely on the rate of formation of cross-bridges between myosin and actin during contraction. 30 The rate of relaxation is an indicator of muscle-relaxation speed and depends on the rate of reattachment of cross-bridges during the relaxation process. 26,31 Our results showed that these intracellular processes are highly affected by PTP after a brief high-frequency submaximal direct PES. Neuromuscular electrical stimulation is often used by physical therapists to improve muscle performance. The present results might have important clinical relevance when using brief trains of electric stimulation to strengthen muscles and helping patients perform functional movements in rehabilitation. These results contribute to our understanding of the relationship between the activation pattern of muscles and the force produced. In conclusion, this study indicated that a brief high-frequency direct submaximal tetanic PES induces posttetanic potentiation in knee-extensor muscles with a small increase at 1 5 minutes followed by a small decrease at 10 minutes. The supramaximal twitch maximal rates of force development and relaxation seem to be more sensitive indicators of posttetanic potentiation than twitch peak force is. Acknowledgments This study was partly supported by Estonian Science Foundation Grant No. 5504. References 1. Hainaut K, Duchateau J. Neuromuscular electrical stimulation and voluntary exercise. Sports Med. 1992;14:100-113. 2. Maffiuletti NA, Pensini M, Martin A. Activation of human plantar flexor muscles increases after electromyostimulation training. J Appl Physiol. 2002;92:1383-1391. 3. Krarup C. Enhancement and diminution of mechanical tension evoked by staircase and by tetanus in rat muscle. J Physiol, Lond. 1981;311:355-372. 4. Houston ME, Grange RW. Myosin phosphorylation, twitch potentiation, and fatigue in human skeletal muscle. Can J Physiol Pharmacol. 1990;68:908-913. 5. O Leary DD, Hope K, Sale DG. Posttetanic potentiation of human dorsiflexors. J Appl Physiol. 1997;83:2131-2138. 6. Rassier DE, MacIntosh BR. Coexistence of potentiation and fatigue in skeletal muscle. Braz J Med Biol Res. 2000;33:499-508. 7. Binder-Macleod SA, Dean JC, Ding J. Electrical stimulation factors in potentiation of human quadriceps femoris. Muscle Nerve. 2002;25:271-279.

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