THE COURSE OF MOTOR UNIT TWITCH IN DEPENDENCE ON MUSCLE STRETCHING FORCE. STUDIES ON MEDIAL GASTROCNEMIUS MUSCLE OF THE RAT

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1 ACTA NEUROBIOL. EXP. 1990, 50: THE COURSE OF MOTOR UNIT TWITCH IN DEPENDENCE ON MUSCLE STRETCHING FORCE. STUDIES ON MEDIAL GASTROCNEMIUS MUSCLE OF THE RAT Kazimierz GROTTEL, Jan CELICHOWSKI and Nat,alia ANISSIMOVA Department of Neurobiology, Academy of Physical Education 10 Droga Debinska St., Poznali, Poland and Laboratory of Motor Physiology, Pavlov Institute of Physiology, Leningrad, USSR Key words: motor unit, muscle stretching, twitch force, contraction time, half-relaxation time Abstract. Studies were performed on 46 units of medial gastrocnemius muscles in 23 rats. In the examined population, three types of motor units were distinguished: fast fatigable (FF), fast resistant (FR) and slow (S) units. The studied muscle was stretched with force gradually increasing from 0.5 to 10 g. Most sf FF units and some of FR units reached the highest twitch force when the muscle was stretched with 4 to 7.5 g. The remaining fast units and alniost all slow units reached the highest twitch force at 10 g stretching of the muscle. In some fast units, twitch force even decreased slightly at stretching of the muscle with 10 g force. On the other hand, contraction tiine and half-relaxation time were getting prolonged in the whole process of stretching the muscle. Most pronounced changes in contraction tiine and half-relaxation time look place in S type units while the least pronounced ones - in FF type units. The prolongation of half-relaxation times in motor units of all three types was more pronounced than the prolongation of contraction times. INTRODUCTION Recording the twitch force of the muscle or motor unit in isometric conditions requires that the muscle be stretched before the test. The stretching of the muscle may be expressed by an increase in muscle

2 length or by the force used for the stretching. Depending upon the dcgree of stretching, the course of the contraction is different. In studies on motor units performed up to the present day the twitch force has been measured in variable conditions. Usually during the experiments muscles have been maintained in a length optimal for the contraction of the whole muscle (8, 17-19, 21, 22, 29). In these studies, the optimal length has been defined as a length of the muscle at which the highest muscle twitch force is registered. In other studies the optimal length of the muscle has been determined for contraction of its individual motor units (2, 10, 23, 24). In some experiments muscle stretching with constant, defined force has been applied, permitting to record the highest twitch force of most of its motor units (3, 4, 5, 20, 31). In the majority of units, the maximal twitch force has been reached at the length of the muscle distinct from the length which has been optimal for the contraction of the whole muscle (27, 28, 30, 33). In conditions optimal for the contraction of the whole muscle, usually a somewhat lowered twitch force has been registered for its motor units (30). Till now, the interrelationship has been examined mainly between muscle length and twitch force and, ocassionally, contraction time as well. The relation has been examined inducing a single cmtraction or maximal tetanus, either in studies on the whole muscles (6, 7, 9, 25, 32) or in studies on motor units (1, 27, 28, 30, 33). In these papers the moltor unit twitch force was related mainly to muscle length (10, 25-28, 33). It should be mentioned that in these studies the motor units have been divided into fast and slow ones only. In the presented study we have examined the interrelationships between twitch force, contraction time and half-relaxation time in the motor units of medial gastrocnemius muscle in the rat on one hand and the force used for stretching the muscle on the other. So far the relations have not been studied in detail and, as far as rat motor units are considered, they have not been examined at all. In accordance to results obtained in earlier publications (Celichowski, in preparation; 13), we have applied a division of the motor units into three types (FF, FR and S). The measurements have been conducted at stretching the muscle using force up to 10 g, since at such stretching of the studied muscle most of its motor units have reached the maximal twitch force, as shown by preliminary observations. MATERIAL AND METHODS The experiments were performed on 23 adult rats of Wistar strain, weighing on the average, g, in pentobarbital anesthesia. Details

3 related to the administering of anesthesia, surgery, regulatilon of the body temperature, immobilization of extremities and isolation of individual motor units were described in earlier publications (Celichowski, in preparation; 15). Muscle fullction was taken as reflecting the function of a single motlor unit if both twitch force and EMG record were consistent with "all or none" reaction type. Methods used to register EMG and the electrodes used were also described earlier (14, 15). Twitch force of the motor units was measured using inductive force transducer in isometric conditions. After finding out that an individual unit was stimulated (at stretching the muscle with 10 g force), sequential twitch records were obtained, stretching the muscle with increasing force, according to the sequence g. Subsequently, the sag test was performed (stimulation of the motor unit with a sequence of stimuli at 40 Hz frequency and 500 ms duration), which permitted to divide the units into fast and slow ones. Finally, the unit was subjected to fatigue test (stimulation of the unit with 330 ms lasting bursts of stimuli at 40 Hz frequency and repeated every one second, for a total of 4 min). Results of the fatigue test allowed to calculate the fatigue index and to divide fast motor units into resistant to fatigue, (FR) and nonresistant ones (FF) (4, 13, Celichowski, in preparation). The way in which twitch force, contraction time, half-relaxation time were measured and fatigue index calculated were described in detail in other papers (12, 13). In the presented study, we have calculated the means of investigated properties and their standard deviations. The significance of differences between the means was calculated using t-student test. RESULTS For three animals the relation was investigated between the muscle length increment and the force with which the muscle was stretched. This relationship is exemplified in Fig. 1. Out of 46 motor units studied, 23 were classified as FF type, 12 units as FR type and 11 as S type. The mean values of twitch force, contraction time, half-relaxation time and of fatigue index are given in Table I. Examples of records of motor unit twitches for all three types of units, obtained during the increasing stretching of the muscle, are presented in Fig. 2. Increasing force of stretching the muscle was paralleled by increasing motor unit twitch force and increasing duration of its course (both contraction time and half-relaxation time).

4 nm Fig. 1. Relationships between muscle stretching force (MS) and its length (L) increment. The measurements were collected durlng stretching the mu- -.: scle with increasing force. Muscle I O A ?,J stretching force equal to 1 g was used 0 to stretch the muscle when its origi- MS nal length (0) was measured. Twitch force The relationships between the twitch force of motor units (expressed in percentage) and muscle stretching is presented in Fig. 3A and B separately for different types of units. Increasing stretching of the muscle induced an increase in twitch force to its highest value (i.e. 100 /o) in most of investigated motor units. Mean values and thcir standard deviations of principal properties of studied motor units of three types. Testing was performed when the muscle was stretched with 10 g force. TwF, twitch force; CT, contraction time; HRT, half- -relaxation time; FI, fatigue index. The same abbreviations were also used in Fig Motor ur~it type TwF CT 13 RT FI (9) (111s) (ms) In the extreme case among investigated FF units (marginal curve delimiting FF type unit area at the left hand side, Fig. 34, already at stretching the muscle with 4 g force, the twitch force of the unit reached its maximum. In other units - this maximum was reached at stretching the muscle with 5 or 7.5 g force. Further stretching of the muscle (with force from 7.5 up to 10 g) in experiments on the part of FF type units

5 IFF FR S Fig. 2. Twitch courses in FF, FR and S type motor units recorded during stretching the muscle with increasing force. Sequential records for each unit are placed in separate columns. Numbers to the left relate to records in all three columns and indicate the force used to stretch the muscle at the time of recording. Fig. 3. A, course of changes of twitch force (TwF) in motor units of various type, recorded during stretching the muscle with increased force (MS). The highest recorded twitch force was taken as 100 /o TwF. Each point represents the mean value and its standard deviation. FF type units are marked in filled circles, FR units in open circles, S in crosses. B, areas between marginal (on the axis marked by dotted line) curves of dependence of twitch force upon muscle stretching force, separately for each of the distinguished types of motor unlts.

6 induced a decrease in their twitch force by 2 to 8% (n = lo), or the units maintained the maximal twitch force (n = 3). In the remaining 10 FF type motor units the maximal twitch force was recorded at stretching the muscle with 10 g force. In FR group, the marginal motor unit reached its maximal twitch forte also at 4 g stretching of the muscle, in 4 other cases, - at 5 or 7.5 g stretching. At,still higher stretching of the muscle, i.e. at stretching with 10 g force, the twitch force of one of the units decreased by 4"o. In the remaining 7 units of FR type the highest twitch force was reached when the muscle was stretched with 10 g force. The twitch force of slow motor units reached usually its maximal value at the strongest stretching, applied i.e. 10 g (Fig. 3A and B)..In only two of the units the highest twitch,force values were reached already at stretchi~g the muscle with 7.5 g force. On the other hand, still stronger stretching of the muscle (with 10 g force) did not induce a decrease in twitch force in these two units. Differences in twitch force values between motor units of various types at'stretching the muscle with force ranging from 0.5 t;o 10 g (Fig. 3A) proved insignificant. In 5 muscles in which more than 1 motor unit was examined (2-6), the course of curves of dependence between twitch force and muscle stretching was analysed for motor units of various types. Usually also in a single muscle twitch the force of fast motor units reached higher relative values (in percentage) in sequential points of increasing stretching than did slow units (Fig. 4). However, no relations were detected between'the values of twitch force (expressed in grams) of units and the 100 % Cli 3 50 b Fig. 4. Course of changes in twich force in 6 individual motor units of the same muscle. The curves were drawn similarly to those in Fig. 3. Numbers in the Figure denote motor unit twitch force in grams (tested at g muscle stretching force). Symbols indicating types of motor units are MS 9 identical to those of Fig. 3A.

7 Fig. 5. Relative changes in contraction times (CT) of mot~r units in each of three types (mean f SD) in function C % ti of tnuscle stretching. Symbols identical to those of Fig. 3 A. The longest 50 contraction time was accepted as 100% (always at 10 g stretching force). MS 9 course of twitch force changes during the stretching of the muscle (Fig. 4). ~ontractio?time The relationship between the cointraction time of motor units of various type and the stretching of the muscle is presented in Fi,g. 5. In the course of the entire process of stretching the muscle contraction time was becoming prolonged in all types of units. Contraction time in FF units showed, on the average, the least pronounced alterations when the muscle was stretched with force increasing from 0.5 to 10 g, while contraction time of slow units showed greatest alterations. In all units, an increase in contraction time was greater when the muscle stretching force increased from 0.5 to 5 g than in the remaining portion of the experiment. Significant differences (p < 0.05) were detected when contraction times expressed in percentages were compared between FF, and S type units at stretching the muscle with the force up to 5 g. I Relation between half-relaxation time and muscle stretching is prcsented in Fig. 6. Increasing stretching of the muscle induced also a prolongation of half-relaxation times in units of any type. The greatest increase in the half-relaxation time upon stretching the muscle within the studied range took place in S type motor ulnits, the smallest - in FF type units. Similarly to contraction time, half-relaxation time was getting prolonged, particularly in the earlier part of the experiment, i.e. upon stretching the muscle with force increasing up to 5 g, in all three

8 Fig. 6. Relative changes in half-relaxation times (HRT, mean i SD) in motor units of three types. For symbol explanation see Fig The long est half-relaxation time was taken as hf!3 Q 1000/o (always at 10 g stretching force). types of units. Significant differences (p < 0.05) were noted between relative values of half-relaxation times between FF and S type units, upon stretching the muscle with force increasing up to 2 g. Comparisons were made also between relative values (in percentages) of contraction times on one hand and half-relaxation times on the other, separately for each of all three studied types of motor units. Values of the two compared properties in FF units differed significantly from each other upon stretching the muscle with force to 5 g, in FR units - with force up to 3 g, in S units - with force up t~ 7.5 g. DISCUSSION Multiple studies on the twitch of entire muscle and motor units indicate that the twitch force registered during stretching the muscle (during increase in its length) increases originally and, after reaching a certain value, gradually decreases (6, 9, 26-28, 33). The studies were usually performed by progressive stretching the muscle each time by the same unit of length (9, 27, 33). In this study we have examined changes in the twitch force of motor units, defining the force with which, the muscle was stretched. This way of testing provides more dependable control of a degree of muscle stretching examining motor units. This is particularly important when the muscle is stretched with gradually increasing force. At the beginning, stretching the muscle with a unit of force has resulted in a relatively marked increase in its length, while further stretching has lead to a still lower increase in muscle length per increase in the unit of force (Fig. I), (7, 33). The discussed results relate to records made during stretching the muscle with force increasing from

9 0.5 to 10 g. The upper limit of testing, equal to 10 g, reflects the force of stretching the muscle at which the twitch force of most motor units reaches its highest value. In such muscle load, the twitch force of some fast units begins even to decrease. However, the decrease is slight and amounts up to 8O/d. At stretching the muscle with 10 g force, at the average, a higher twitch force is reached in motor units of any type, as compared to stretching the muscle with 7.5 g force. The difference is also inconsiderable, amounting from 3O/o to 7010 for FF and S type units, respectively. Thus, stretching the medial gastrocnemius muscle of the rat with 10 g force may be thought optimal for testing the units of the muscle. In the presented experiments we have noted that among the distinguished three types of motor units, the twitch force of FF type motor units is the earliest to reach maximal values and the twitch force of S units is the last one to reach maximal values during a gradual increase in stretching force up to 10 g. Similar results hqve been obtained by other authors (1, 33). The results indicate that FF type motor units are adjusted to work in conditions of lower extending of the muscle (at its shorter length) than FR and S type units. This fact may have a functional importance. During normal movement, the slow motor units are first recruited into contraction (16). The muscle is then extended, (i.e. it has its greatest length). In the extended muscle slow motor units work in optimal length colnditions. In the course of contractioln the slow motor units start the shortening of the muscle and then, with some delay (upon certain shortening of the muscle), fast motor units become involved. However, the latter do not reach the maximal values of their force until the belly of the muscle becomes shortened. Stephens et al. (33) have related the differeince between fast and slow motor units in their optimal muscle length to differences in absolute values of twitch force between fast and slow units. In the experiments presented in this study much higher twitch forces have been obtained for FF type units than for the remaining ones. On the other hand, twitch forces expressed in percentages for various extent of muscle stretching have not, differed from each other so significantly. The absence of statistical significance of the differences may reflect a low number of examined units and, in part, individual differences between motor units of even the same type within a given muscle. The latter factor may reflect, e.g. the position of muscle fibers of the studied units within the cross-section of the muscle, length of the fibres or other properties. In all motor units, contraction time and half-relaxation time have become prolonged upon stretching the muscle. This has taken place even if the twitch force of some units was becoming constant or has

10 been decreasing at the end of stretching. These observations confirm the results of twitch studies on muscles and motor units by other authors (6, 26, 32-34). In FF units, contraction time and half-relaxation time became only slightly, while in S units, markedly prolonged. Changes in half-relaxation times in units of all three types have been more evident than corresponding changes in contraction times. The latter observation has also corroborated the results of other authors on colntsactions of muscles and motor units (6, 26, 33). As evident from respective literature, the muscles of extremities remain in the length close to the optimal one for contraction during their normal activity (1, 7, 9, 26). Changes in muscle stretching during changes in angles of joints at which the muscle acts lead probably not only to changes in the twitch force of motor units but probably induce changes in contraction and relaxation times as well. The latter changes, in turn, may affect the course of unfused tetani (34). Motor units usually work in such unfused tetani (11) and the degree to which the tetanus is fused (and, in effect tetanic tension) depends, apart from the frequency of motoaeurone firing, upon cointraction and relaxation times. Thus, the twitch force of the muscle during its normal activity should be considered to represent not only the net result of a number of functioning motor units and frequencies of their contractions. Undoubtedly, the original stretching of the muscle, dependent upon angular position of the extremity, may also affect the original twitch force. This investigation was partly supported by Project RP V.3.4. REFERENCES 1. BAGUST, J., KNOTT, S., LEWIS, D. M., LUCK, J. C. and WESTERMAN, R. A Isometric contractions of motor units in a fast twitch muscle of the cat. J. Physiol. 231: BAGUST, J., LEWIS, D. M. and LUCK, J. C Post-tetanic effects in motor units of fast and slow twitch muscle of the cat. J. Physiol. 237: BURKE, R. E Motor units types of cat triceps surae muscle. J. Physiol. 193: BURKE, R. E., LEVINE, D. M., TSAIRIS, P. and ZAJAC, I?. E Physiological types and histochemical profiles in motor units of the cat gastrocnemius. J. Physiol. 234: CLAMAN, H. P. and ROBINSON, A. J A comparison of elecromyographic and mechanical fatlgue properties In motor units of the cat hindlimb. Brain Res. 327: CLOSE, R. I Dynamic properties of fast and slow skeletal muscles of the rat during development. J. Physiol. 173:

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