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The normal motor unit in man
Journal of the Neurological Sciences, 27 (1976) 291-301 © Elsevier Scientific Publishing Company, Amsterdam - Printed in The Netherlands
291
THE NORMAL MOTOR UNIT IN MAN A Single Fibre E M G Multielectrode Investigation
E. ST.~LBERG, M. S. SCHWARTZ, BARBARA THIELE and H. H. SCHILLER Department of Clinical Neurophysiology (E.S., M.S.S., H.H.S.), University Hospital, Uppsala (Sweden), and Department of Clinical Neurophysiology (B.T.), Klinikum Steglitz, Berlin (WestGermany)
(Received 18 July, 1975)
SUMMARY The motor unit in the normal E D C and biceps brachii muscle was studied with a multielectrode technique permitting recording from 44 sites 300 # m apart. It was found that the muscle fibres in man are scattered with no evidence of grouping, similar to the findings in glycogen depletion experiments in animals.
INTRODUCTION The distribution of muscle fibres in a single motor unit in man has been a source of controversy for many years. The concept of subunit arrangement in man was advanced by Buchthal, Guld and Rosenfalck (1957a), following multilead physiological investigations but was at variance with physiological investigations in the rat diaphragm (Krnjevic and Miledi 1958) and gastrocnemius muscles (Norris and Irwin 1961). The subunit was questioned in single fibre investigations in mart (Ekstedt 1964) and the subunit concept is no longer he!d (Buchthal and Rosenfalck 1973). In the previous neurophysiological multielectrode studies in man (Buchthal, Guld and Rosenfalck 1967b) 12 lead-off surfaces 1.5 m m long were used with a spacing of 0.5 mm to measure the motor unit territory. The distribution of individual fibres in the m o t o r unit was not studied as this was prevented by the relatively wide interelectrode distances. For volume conduction studies another type of multielectrode with 12 lead-off surfaces six of which were 50 # m in diameter and six 100/~m, with
This investigation was supported by the Swedish Medical Research Council (Grant No. 135). Hans H. Schiller was supported by a Swiss Grant (Stiftung fiir Biologisch-Medizinische Stipendien).
292 an interelectrode distance of 0.15-0.3 mm was used. Since this electrode had a total length of just 2.5 mm it was too short for motor unit mapping. We have developed a multielectrode with characteristics which make it possible to map the distribution of single muscle fibres of motor units in man. METHODS
Since the average diameters of the motor unit territory in the extensor digitorum communis and biceps brachii muscles in man are reported to be about 5 mm (Buchthal, Erminio and Rosenfalck 1959), the length of the present electrode was constructed to be long enough to cover even the large motor units. In the side of a steel cannula 0.6 mm in diameter, 14 platinum electrode surfaces 25/~m in diameter were arranged over a total distance of 12 ram. The interelectrode distances of electrodes 1 to 6 and 10 to 14 were approximately 1.2 mm and the interelectrode distances for the central electrodes 6 to 10 were approximately 0.3 mm (Fig. 1). Thus, the total distance between electrode 6 and 10 was constructed to be similar to the larger interelectrode distances. The recordings were made simultaneously from 2 of the electrodes against a cutaneous indifferent electrode. Two amplifiers with an input impedance greater thart 200 Mohm and frequency limits of 500-30000 Hz (12 db) were used. The action potentials were displayed on a Tektronix oscilloscope (D! 5) and recorded on analog tape (Akai X330) and later displayed on Medelec UV film for further analysis. Delay lines (5 msec, Medelec, Disa) were used.
Recordingprocedure The multielectrode was inserted perpendicular to the fibres. In order to ascertain that the electrode was inside the muscle, recording was made from electrodes 1 and 14. When some activity was obtained from both on voluntary activity the recording began. The investigations were made at slight voluntary activity. Recording was first made from electrode 6, which was the first "reference electrode". The multielectrode was adjusted for obtaining a single fibre action potential of high amplitude (maximum) and short rise time. This action potential was displayed on the first beam of the oscilloscope and triggered the sweep. The electrode position was maintained and recordings were made successively from the other electrode surfaces by means of automatic or manual electronic switching after 10 or 20 discharges or" the triggering potential. The activity from the other electrodes was displayed on the second beam. The presence of action potentials time-locked to the triggering one, i.e., activity from fibres belonging to the same motor unit, was noted for each of the electrodes. Delay lines (5 msec) were used for both of the recording channels to identify action potentials occurring before the triggering one. In order to be accepted, the action potentials had to fulfil the criteria for a single muscle fibre action potential (Ekstedt 1964), reach an amplitude greater than 200/~V, have a fast rise time (maxmin time less than 300 /~sec) and be seen on consecutive discharges. With these criteria only single fibre action potentials relatively close (approximately 270 #m)
293
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Fig. 1. The multielectrode, 90° rotated and en face, with 14 platinum 25 pm recording surfaces. The uptake radius for each electrode (on average 270/~m) is indicated for the left multielectrode. The recorded corridor (C) after the 4 multielectrode advancements of 300 pm each is 13.2 × 0.27 mm. The action potentials recorded at one position are shown to the right. Two of the action potentials (O m) are each recorded from two electrodes (8-9, circles and 9-10 squares) but the responses are only counted when they appear with maximal amplitude (8 and 9). A third action potential (*) fulfilling the criteria for acceptance is seen over electrode 7.
to the active r e c o r d i n g electrodes were counted. Muscle fibres close to one electrode were usually r e c o r d e d with a n a m p l i t u d e less t h a n 200 /~V at the next r e c o r d i n g p o s i t i o n , 3 0 0 / ~ m away. H o w e v e r , a muscle fibre situated between 2 r e c o r d i n g p o sitions, 300 /~m a p a r t , m a y fulfil the criteria for acceptance over b o t h electrodes. H e r e tests were m a d e by r e c o r d i n g f r o m b o t h surfaces s i m u l t a n e o u s l y a n d the time r e l a t i o n between the p o t e n t i a l s was studied. I f the second a c t i o n p o t e n t i a l a p p e a r e d at a n o t h e r p o s i t i o n o n the oscilloscope sweep this i n d i c a t e d r e c o r d i n g f r o m a n o t h e r fibre. W h e n the triggering p o t e n t i a l a n d the response o n the other channel o c c u r r e d at the same oscilloscope sweep p o s i t i o n the jitter between t h e m was studied. T h e presence o f a j i t t e r i n d i c a t e d recordings f r o m 2 fibres (Sthlberg, E k s t e d t a n d Brom a n 1970). I f there was no j i t t e r at successive discharges the recordings were o b t a i n e d f r o m the same muscle fibre (Fig. 2). W h e n the same a c t i o n p o t e n t i a l was r e c o r d e d
294
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Fig. 2. Simultaneously recorded action potentials at one multielectrode position. Reference action potential over electrode 8 triggers the oscilloscope sweep. Over electrode 9 two responses are obtained, one on the same sweep position as the triggering one. This is the volume-conducted action potential from the muscle fibre generating the triggering action potential as there is no jitter between them and is not counted as an extra response. From electrode 9 an additional action potential is recorded which jitters relative to the triggering action potential indicating another muscle fibre belonging to the same motor unit. over 2 or more recording sites, 300/~m apart, it was only counted once, at that electrode where the amplitude was highest. W h e n 2 or more action potentials from fibres within the same m o t o r unit interfered, difficulties sometimes arose in determining the numbers o f individual spike components. There is always, however, a small time variability between action potentials from muscle fibres o f the same m o t o r unit at consecutive discharges (St~ilberg et al. 1970) and therefore the individual spike c o m p o n e n t s even in a c o m p o u n d action potential can usually be identified. After scanning all the 13 other electrodes, some fibres belonging to the same m o t o r unit within the uptake corridor would still not be recorded due to the long interelectrode distances (1.2 mm) o f electrodes 1-6 and 10-14. Therefore the multielectrode was advanced 300 # m until the "reference action potential" was recorded over electrode 7 with the same shape as it had over electrode 6. This was facilitated by having recordings from both the new and the former triggering electrode displayed, when advancing the multielectrode. The remainder o f the electrodes were then rescanned. The multielectrode was then successively m o v e d in 300-/tin steps such that the "reference action potential" was recorded from electrodes 8, 9 and 10 so that all fibres along the entire recording corridor could be identified (Fig. 3). W h e n the "reference action potential" had been advanced f r o m electrode 6 to 10 the same responses should be seen when the other electrodes were scanned. If, e.g., responses were initially obtained over electrodes 5 and 6 the same responses should then be recorded over electrodes 6 and 10 (Fig. 3). This constancy validated the electrode movements and was a general finding t h r o u g h o u t the investigations. I f the response profile changed after a full advancement, that m o t o r unit was discarded. In cases when no responses were obtained aside f r o m the "reference action potential", characteristics such as shape and firing pattern were employed to identify this action
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Fig. 3. The principles for the multielectrode movements allowing recording from 44 sites, 300/~m apart. Only the electrode surfaces are indicated. The electrode arrays for each step are drawn below each other although the 4 advancements are along the multielectrode axis (+-). Top row: initial position with "reference action potential" over electrode 6. Responses are obtained over electrodes 5, 6, 8 and 9. The electrode is then advanced 300 pm so that electrode 7 becomes the reference electrode and responses are recorded over electrodes 9 and 10. At the third advancement (with ref. electrode 9) a new response is obtained over electrode 5 and the responses initially over electrodes 8 and 9 are not within the uptake area of any electrode. At the fourth advancement (with ref. electrode 10) the action potentials initially recorded over electrodes 5 and 6 are now over electrodes 6 and 10, respectively. The distribution of the motor unit's muscle fibres in the corridor is indicated at the bottom.
potential with successive electrode movements. It was m u c h easier to follow the "reference action potential" when it was complex. The investigation o f an individual m o t o r unit usually t o o k 5-10 min. The recordings were usually reanalysed from the analogue tape with the above-described technique.
The uptake area of the electrode The total recording area o f this electrode is somewhat complex. In volume conduction studies it was f o u n d that the distance between the recording electrode and a muscle fibre generating an action potential just fulfilling the amplitude criterion o f 200/zV, having a rise time less than 300 #sec and having a constant shape at consecutive discharges was on average 270 # m (range o f 44 tested fibres was 150-350 #m). The uptake area at one multielectrode position therefore consists o f 14 semicircles each with a radius o f 270 #m. Since the 5 central electrodes have an intercentre distance o f 300 # m their uptake areas overlap. W h e n the electrode had been advanced successively over the four 300-/zm steps the recorded " c o r r i d o r " consisted o f 44 semicircles with a radius o f 270/~m with an intercentre distance o f 300/~m, corresponding to a total length o f 13.2 m m (44 x 3 0 0 / z m = 13.2). There was a slight " d e a d space" in the corridor o f a b o u t 15 ~o due to the semicircular uptake area o f each recording surface (Fig. 1).
Material Nine subjects aged 19-70 years without neurological disorders were investigated. F o u r to 6 recordings at different sites were made f r o m all subjects in extensor digitorum c o m m u n i s (EDC) muscle and in 7 o f them in the biceps brachii muscle. A total o f 53 m o t o r unit studies were carried out in the E D C and 48 in the biceps brachii muscle.
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A typical recording from one electrode position is shown in Fig. 1. In Fig. 4 responses over the central electrodes are shown for a complete study. The distribution of fibres for the investigated individual recorded corridors in the 2 muscles is seen in Figs. 5 and 6. The average number of fibres over the reference electrode was 1.4 -4- 0.2 in EDC and 1.3 ± 0.2 in the biceps brachii muscle. For the other electrodes the fibre density over the electrode where potentials were recorded was 1.2 in EDC and 1.2 in the biceps brachii muscle. There were no definite differences relative to age in the material. In the individual corridor 1 to 9 fibres were seen both in the EDC and in the biceps brachii muscle. Nine fibres were seen in 3 of the 101 recordings from the 2 muscles. The range of the mean values of the number of fibres in the recorded corridors was 2.5 to 4.9 (mean 3.9; SD 0.8) in the EDC and 1.9 to 5.5 (mean 3.7; SD 1.3) in the biceps brachii muscle. In the EDC there were 13 recordings with fibres only over the reference electrode ( 2 4 ~ ) and in biceps brachii also 13 recordings showed no additional fibres (27 ~). When the "reference action potential" was a single fibre action potential there were, for EDC on average 2.1 fibres outside the reference e}ectrode, for double spikes on average 2.9 additional fibres and for 3 reference action potentials another 3.4 fibres were recorded. The corresponding figures for biceps were t.9, 3.5 and 5 (1 case), respectively. Even though the number of additional fibres was increased in the
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corridor when the "reference action potential" was complex the fibre density over active electrodes was not similarly increased. In none o f the 101 corridors studied were there more than 3 fibres over any electrode. The longest distance between 2 fibres recorded in a single motor unit study was 6 m m (3 recordings) in E D C and 6-7 m m (5 recordings) and 9 m m (1 recording) in biceps brachii muscle. In about a third o f the trials the entire procedure could not be completed due to the loss o f the "reference action potential".
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Motor unit corridors in normal biceps brachii muscles.
DISCUSSION
The distribution of muscle fibres within the individual motor units has not previously been demonstrated in vivo in man. In previous physiological multielectrode studies of the motor unit territory in man (Buchthal et al. 1957b; Buchthal et al. 1959), the size of the individual recording surface and their relatively wide spacings did not facilitate the study of the distribution of single fibres. This problem has been overcome with the present technique with selective recordings from small electrode surfaces representing 44 sites, 300/~m apart. The validity of the results of the present method depends principally on: (1) The electrode direction. The electrode must be positioned perpendicular to the fibre direction.
299
(2) The electrode position within the muscle. All the electrode surfaces must be positioned within the muscle. In order to help position the multielectrode over the motor unit cross-section, the reference electrodes (6-10) were constructed to be at the centre of the electrode array. I f the 5 close electrodes had been at the end of the array there would be greater risk for the remaining electrodes to be positioned outside the motor unit under study. The maximal distance between electrode surfaces recording action potentials from fibres in the same motor unit is some indication of the territory, but this particular parameter was not studied in this investigation. (3) Accuracy of electrode movement. As the electrode has to be advanced 4 steps of 300 # m each during the investigation, there is a risk to lose the "reference action potential" of the m o t o r unit under study. Also, some muscle fibre damage or fibre dislocation can occur. A check of the correct electrode movement was the constancy of the response profile when the triggering electrode had been advanced from 6 to 10, i.e., the multielectrode was moved one long interelectrode distance (1.2 mm) (Fig. 4). The fibre distribution gave an impression of centering with the highest fibre density around the centre of the multielectrode. Two methodological factors are primarily responsible. Firstly, one of the central electrodes (number 6) was used as a reference electrode recording activity from at least 1 fibre of the motor unit. Secondly, as this reference electrode was always inside the motor unit, the chance of being within the territory of the same m o t o r unit is highest for the closely adjoining electrodes. I f the multielectrode was positioned in the middle of the motor unit many electrodes would be within the m o t o r unit territory but if the multielectrode were placed more peripherally, only a few of the more central electrodes could theoretically record activity from that motor unit. The number o f action potentials over the reference electrode tended to characterize the m o t o r unit under study. When only a single response was obtained as the "reference action potential" less additional fibres were recorded in the corridor than when multiple fibres were recorded over the reference electrode. This was particularly evident in the biceps brachii muscle. This probably indicates a higher fibre concentration in some m o t o r units than in others. Whether these motor units have on average a higher total number of fibres cannot be determined. It is known that m o t o r units are of different size with a positive correlation to activation threshold and motor neurone size (Wuerker, McPhedran and Henneman 1965; Burke, Levine, Zajac, Tsaris and Engel 1971). In the present investigation, motor units with low threshold were studied to limit disturbances from other motor units. However, other m o t o r units were often recorded during the course of a single investigation, some with a higher innervation rate, indicating a lower activation threshold than the one under study (St~lberg and Thiele, 1973; Hannertz 1974). There were also some differences between the muscle fibre distribution in motor units from the 2 muscles. In biceps brachii the motor unit tended to be more dispersed. There was a slightly lower fibre density over the reference electrode in biceps brachii than in E D C although there was a similar number of fibres in the corridor
300 for the 2 muscles and a larger maximal distance between fibres recorded from motor" units in biceps brachii than in EDC. The present study has demonstrated that in the normal EDC and biceps muscles the muscle fibres in a single motor unit are scattered similarly to those shown in experimental animals (Edstr6m and Kugelberg 1968; Brandstater and Lambert 1969; Doyle and Mayer 1969) with glycogen depletion experiments. A single motor unit's distribution covers about 17 ~ of the cross-sectioned area in the rat tibialis anterior muscle (Kugelberg 1973). In such studies it was found in rats that the fibres of ~t single motor unit are usually isolated, with 76 ~o of the fibres not adjacent to another fibre of the same motor unit and only rarely 3 fibres together (Brandstater and Lambert 1969). These figures are roughly comparable to the fibre density values obtained with single fibre E M G (St~dberg and Thiele 1975) where only a single fibre is recorded in about 60-65 )/o of random electrode placements. In the present investigation there were never more than 3 fibres over 1 recording electrode in the approximately 300 active electrodes in 101 motor units studied. With conventional histochemistry the principal fibre types can be identified but a motor unit's distribution cannot be made. A mosaic of fibre type distribution in the cross section area of the muscle in man (Engel 1966; Jennekens, Tomlinson and Walton 1971) also indicated scattering although the individual motor units were not identified. In a single case with myokymia, glycogen-depleted fibres of the one or more active motor units were randomly scattered (Williamsson and Brooke 1972). The presented multielectrode investigations have shown the distribution of motor unit fibres in the normal EDC and biceps brachii. The fibres were dispersed in a manner similar to that shown in experimental glycogen depletion studies. In the following paper the changes in motor units with reinnervation are presented (Schwartz, St~dberg, Schiller and Thiele 1976). ACKNOWLEDGEMENTS We thank G6sta Lov6n who made the electrode and Kerstin Mellqvist who assisted during the experiments and made the drawings.
REFERENCES Brandstater, M. E. and E. H. Lambert (1969) A histochemical study of the spatial arrangement of muscle fibers in single motor units within rat tibialis anterior muscle, Bull. Amer. Ass. EMG Electrodiagn., 82: 15-16. Buchthal, F. and P. Rosenfalck (1973) On the structure of motor units. In: J. E. Desmedt (Ed.), New Developments in Electromyography and Clinical Neurophysiology, Vol. 1, Karger, Basel, pp. 71-85. Buchthal, F., F. Erminio and P. Rosenfalck (1959) Motor unit territory in different human muscles, Acta physioL scand., 45: 72-87. Buchthal, F., C. Gutd and P. Ro~nfalck (1957a) Volume conduction of the spike of the motor unit potential investigated with a new type of multielectrode, .4cta physiol, scand., 38: 331-354. Buchthal, F., C. Guld and P. Rosenfalck (1957b) Multielectrode study of the territory of a motor unit, Acta physioL scand., 39: 83-104.
301 Burke, R. E., D. N. Levine, F. E. Zajac, P. Tsaris and W. K. Engel (1971) Mammalian motor units Physiological-histochemical correlation in three types in rat gastrocnemius, Science, 174: 709-712. Doyle, A. M. and R. F. Mayer (1969) Studies of the motor units in the cat, Bull. Sch. Med. Maryland, 54: 11-17. Edstr6m, L. and E. Kugelberg (1968) Histochemical composition, distribution of fibres and fatiguability of single motor units, J. NeuroL Neurosurg. Psychiat., 31 : 424-433. Ekstedt, J. (1964) Human single fibre action potentials, Actaphysiol. scand., 61 : Suppl. 226, pp. 1-96. Engel, W. H. (1966) Histochemistry of neuromuscular diseases - - Significance of muscle fibre types. In : Neuromuscular Diseases, VoL 2 (Proceedings of the 8th International Congress of Neurology, Vienna), pp. 67-101. Hannertz, J. (1974) Discharge properties of motor units in relation to recruitment order in voluntary contraction, Acta physiol, scand., 91: 374-384. Jennekens, F. G. I., B. E. Tomlinson and J. N. Walton (1971) Data on the distribution of fibre types in five h u m a n limb muscles, J. neurol. Sci., 14" 245-257. Krnjevic, K. and R. Miledi (1958) Motor units in the rat diaphragm, J. Physiol. (Lond.), 140: 427439. Kugelberg, E. (1973) Properties of the rat hind-limb motor units. In: J. E. Desmedt (Ed.), New Developments in Electromyography and Clinical Neurophysiology, Vol. 1, Karger, Basel, pp. 213. Norris, F. H. and R. L. Irwin (1961) Motor unit area in rat muscle, Amer. J. Physiol., 200: 944-946. Schwartz, M. S., E. St~,lberg, H. H. Schiller and B. Thiele (1976) The reinnervated motor unit in man - - A single fibre E M G multielectrode investigation, J. neurol. Sci., 27: 303-312. Sthlberg, E. and B. Thiele (1973) Discharge pattern of motor neurones in humans. In: J. E. Desmedt (Ed.). New Developments in Electromyography and Clinical Neurophysiology, VoL 3, Karger, Basel, pp. 234-241. St~dberg, E. and B. Thiele (1975) Motor unit fibre density in the extensor digitorum communis muscles - - Single fibre electromyographic study in normal subjects at different ages, J. Neurol. Neurosurg. Psychiat., 38: 874-880. Sthlberg, E., J. Ekstedt and A. Broman (1970) The electromyographic jitter in normal human muscles, Electroenceph. clin. Neurophysiol., 31: 429438. Williamsson, E. and M. H. Brooke (1972) Myokymia and the motor unit, Acta physioL scand., 26: 11-16. Wuerker, E. B., A. M. McPhedran and E. Henneman (1965) Properties of motor units in a heterogenous pale muscle (m. gastrocnemius) of the cat, J. Neurophysiol., 28 : 85-99. -
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291
THE NORMAL MOTOR UNIT IN MAN A Single Fibre E M G Multielectrode Investigation
E. ST.~LBERG, M. S. SCHWARTZ, BARBARA THIELE and H. H. SCHILLER Department of Clinical Neurophysiology (E.S., M.S.S., H.H.S.), University Hospital, Uppsala (Sweden), and Department of Clinical Neurophysiology (B.T.), Klinikum Steglitz, Berlin (WestGermany)
(Received 18 July, 1975)
SUMMARY The motor unit in the normal E D C and biceps brachii muscle was studied with a multielectrode technique permitting recording from 44 sites 300 # m apart. It was found that the muscle fibres in man are scattered with no evidence of grouping, similar to the findings in glycogen depletion experiments in animals.
INTRODUCTION The distribution of muscle fibres in a single motor unit in man has been a source of controversy for many years. The concept of subunit arrangement in man was advanced by Buchthal, Guld and Rosenfalck (1957a), following multilead physiological investigations but was at variance with physiological investigations in the rat diaphragm (Krnjevic and Miledi 1958) and gastrocnemius muscles (Norris and Irwin 1961). The subunit was questioned in single fibre investigations in mart (Ekstedt 1964) and the subunit concept is no longer he!d (Buchthal and Rosenfalck 1973). In the previous neurophysiological multielectrode studies in man (Buchthal, Guld and Rosenfalck 1967b) 12 lead-off surfaces 1.5 m m long were used with a spacing of 0.5 mm to measure the motor unit territory. The distribution of individual fibres in the m o t o r unit was not studied as this was prevented by the relatively wide interelectrode distances. For volume conduction studies another type of multielectrode with 12 lead-off surfaces six of which were 50 # m in diameter and six 100/~m, with
This investigation was supported by the Swedish Medical Research Council (Grant No. 135). Hans H. Schiller was supported by a Swiss Grant (Stiftung fiir Biologisch-Medizinische Stipendien).
292 an interelectrode distance of 0.15-0.3 mm was used. Since this electrode had a total length of just 2.5 mm it was too short for motor unit mapping. We have developed a multielectrode with characteristics which make it possible to map the distribution of single muscle fibres of motor units in man. METHODS
Since the average diameters of the motor unit territory in the extensor digitorum communis and biceps brachii muscles in man are reported to be about 5 mm (Buchthal, Erminio and Rosenfalck 1959), the length of the present electrode was constructed to be long enough to cover even the large motor units. In the side of a steel cannula 0.6 mm in diameter, 14 platinum electrode surfaces 25/~m in diameter were arranged over a total distance of 12 ram. The interelectrode distances of electrodes 1 to 6 and 10 to 14 were approximately 1.2 mm and the interelectrode distances for the central electrodes 6 to 10 were approximately 0.3 mm (Fig. 1). Thus, the total distance between electrode 6 and 10 was constructed to be similar to the larger interelectrode distances. The recordings were made simultaneously from 2 of the electrodes against a cutaneous indifferent electrode. Two amplifiers with an input impedance greater thart 200 Mohm and frequency limits of 500-30000 Hz (12 db) were used. The action potentials were displayed on a Tektronix oscilloscope (D! 5) and recorded on analog tape (Akai X330) and later displayed on Medelec UV film for further analysis. Delay lines (5 msec, Medelec, Disa) were used.
Recordingprocedure The multielectrode was inserted perpendicular to the fibres. In order to ascertain that the electrode was inside the muscle, recording was made from electrodes 1 and 14. When some activity was obtained from both on voluntary activity the recording began. The investigations were made at slight voluntary activity. Recording was first made from electrode 6, which was the first "reference electrode". The multielectrode was adjusted for obtaining a single fibre action potential of high amplitude (maximum) and short rise time. This action potential was displayed on the first beam of the oscilloscope and triggered the sweep. The electrode position was maintained and recordings were made successively from the other electrode surfaces by means of automatic or manual electronic switching after 10 or 20 discharges or" the triggering potential. The activity from the other electrodes was displayed on the second beam. The presence of action potentials time-locked to the triggering one, i.e., activity from fibres belonging to the same motor unit, was noted for each of the electrodes. Delay lines (5 msec) were used for both of the recording channels to identify action potentials occurring before the triggering one. In order to be accepted, the action potentials had to fulfil the criteria for a single muscle fibre action potential (Ekstedt 1964), reach an amplitude greater than 200/~V, have a fast rise time (maxmin time less than 300 /~sec) and be seen on consecutive discharges. With these criteria only single fibre action potentials relatively close (approximately 270 #m)
293
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Fig. 1. The multielectrode, 90° rotated and en face, with 14 platinum 25 pm recording surfaces. The uptake radius for each electrode (on average 270/~m) is indicated for the left multielectrode. The recorded corridor (C) after the 4 multielectrode advancements of 300 pm each is 13.2 × 0.27 mm. The action potentials recorded at one position are shown to the right. Two of the action potentials (O m) are each recorded from two electrodes (8-9, circles and 9-10 squares) but the responses are only counted when they appear with maximal amplitude (8 and 9). A third action potential (*) fulfilling the criteria for acceptance is seen over electrode 7.
to the active r e c o r d i n g electrodes were counted. Muscle fibres close to one electrode were usually r e c o r d e d with a n a m p l i t u d e less t h a n 200 /~V at the next r e c o r d i n g p o s i t i o n , 3 0 0 / ~ m away. H o w e v e r , a muscle fibre situated between 2 r e c o r d i n g p o sitions, 300 /~m a p a r t , m a y fulfil the criteria for acceptance over b o t h electrodes. H e r e tests were m a d e by r e c o r d i n g f r o m b o t h surfaces s i m u l t a n e o u s l y a n d the time r e l a t i o n between the p o t e n t i a l s was studied. I f the second a c t i o n p o t e n t i a l a p p e a r e d at a n o t h e r p o s i t i o n o n the oscilloscope sweep this i n d i c a t e d r e c o r d i n g f r o m a n o t h e r fibre. W h e n the triggering p o t e n t i a l a n d the response o n the other channel o c c u r r e d at the same oscilloscope sweep p o s i t i o n the jitter between t h e m was studied. T h e presence o f a j i t t e r i n d i c a t e d recordings f r o m 2 fibres (Sthlberg, E k s t e d t a n d Brom a n 1970). I f there was no j i t t e r at successive discharges the recordings were o b t a i n e d f r o m the same muscle fibre (Fig. 2). W h e n the same a c t i o n p o t e n t i a l was r e c o r d e d
294
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Fig. 2. Simultaneously recorded action potentials at one multielectrode position. Reference action potential over electrode 8 triggers the oscilloscope sweep. Over electrode 9 two responses are obtained, one on the same sweep position as the triggering one. This is the volume-conducted action potential from the muscle fibre generating the triggering action potential as there is no jitter between them and is not counted as an extra response. From electrode 9 an additional action potential is recorded which jitters relative to the triggering action potential indicating another muscle fibre belonging to the same motor unit. over 2 or more recording sites, 300/~m apart, it was only counted once, at that electrode where the amplitude was highest. W h e n 2 or more action potentials from fibres within the same m o t o r unit interfered, difficulties sometimes arose in determining the numbers o f individual spike components. There is always, however, a small time variability between action potentials from muscle fibres o f the same m o t o r unit at consecutive discharges (St~ilberg et al. 1970) and therefore the individual spike c o m p o n e n t s even in a c o m p o u n d action potential can usually be identified. After scanning all the 13 other electrodes, some fibres belonging to the same m o t o r unit within the uptake corridor would still not be recorded due to the long interelectrode distances (1.2 mm) o f electrodes 1-6 and 10-14. Therefore the multielectrode was advanced 300 # m until the "reference action potential" was recorded over electrode 7 with the same shape as it had over electrode 6. This was facilitated by having recordings from both the new and the former triggering electrode displayed, when advancing the multielectrode. The remainder o f the electrodes were then rescanned. The multielectrode was then successively m o v e d in 300-/tin steps such that the "reference action potential" was recorded from electrodes 8, 9 and 10 so that all fibres along the entire recording corridor could be identified (Fig. 3). W h e n the "reference action potential" had been advanced f r o m electrode 6 to 10 the same responses should be seen when the other electrodes were scanned. If, e.g., responses were initially obtained over electrodes 5 and 6 the same responses should then be recorded over electrodes 6 and 10 (Fig. 3). This constancy validated the electrode movements and was a general finding t h r o u g h o u t the investigations. I f the response profile changed after a full advancement, that m o t o r unit was discarded. In cases when no responses were obtained aside f r o m the "reference action potential", characteristics such as shape and firing pattern were employed to identify this action
295 1 JII
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potential with successive electrode movements. It was m u c h easier to follow the "reference action potential" when it was complex. The investigation o f an individual m o t o r unit usually t o o k 5-10 min. The recordings were usually reanalysed from the analogue tape with the above-described technique.
The uptake area of the electrode The total recording area o f this electrode is somewhat complex. In volume conduction studies it was f o u n d that the distance between the recording electrode and a muscle fibre generating an action potential just fulfilling the amplitude criterion o f 200/zV, having a rise time less than 300 #sec and having a constant shape at consecutive discharges was on average 270 # m (range o f 44 tested fibres was 150-350 #m). The uptake area at one multielectrode position therefore consists o f 14 semicircles each with a radius o f 270 #m. Since the 5 central electrodes have an intercentre distance o f 300 # m their uptake areas overlap. W h e n the electrode had been advanced successively over the four 300-/zm steps the recorded " c o r r i d o r " consisted o f 44 semicircles with a radius o f 270/~m with an intercentre distance o f 300/~m, corresponding to a total length o f 13.2 m m (44 x 3 0 0 / z m = 13.2). There was a slight " d e a d space" in the corridor o f a b o u t 15 ~o due to the semicircular uptake area o f each recording surface (Fig. 1).
Material Nine subjects aged 19-70 years without neurological disorders were investigated. F o u r to 6 recordings at different sites were made f r o m all subjects in extensor digitorum c o m m u n i s (EDC) muscle and in 7 o f them in the biceps brachii muscle. A total o f 53 m o t o r unit studies were carried out in the E D C and 48 in the biceps brachii muscle.
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A typical recording from one electrode position is shown in Fig. 1. In Fig. 4 responses over the central electrodes are shown for a complete study. The distribution of fibres for the investigated individual recorded corridors in the 2 muscles is seen in Figs. 5 and 6. The average number of fibres over the reference electrode was 1.4 -4- 0.2 in EDC and 1.3 ± 0.2 in the biceps brachii muscle. For the other electrodes the fibre density over the electrode where potentials were recorded was 1.2 in EDC and 1.2 in the biceps brachii muscle. There were no definite differences relative to age in the material. In the individual corridor 1 to 9 fibres were seen both in the EDC and in the biceps brachii muscle. Nine fibres were seen in 3 of the 101 recordings from the 2 muscles. The range of the mean values of the number of fibres in the recorded corridors was 2.5 to 4.9 (mean 3.9; SD 0.8) in the EDC and 1.9 to 5.5 (mean 3.7; SD 1.3) in the biceps brachii muscle. In the EDC there were 13 recordings with fibres only over the reference electrode ( 2 4 ~ ) and in biceps brachii also 13 recordings showed no additional fibres (27 ~). When the "reference action potential" was a single fibre action potential there were, for EDC on average 2.1 fibres outside the reference e}ectrode, for double spikes on average 2.9 additional fibres and for 3 reference action potentials another 3.4 fibres were recorded. The corresponding figures for biceps were t.9, 3.5 and 5 (1 case), respectively. Even though the number of additional fibres was increased in the
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corridor when the "reference action potential" was complex the fibre density over active electrodes was not similarly increased. In none o f the 101 corridors studied were there more than 3 fibres over any electrode. The longest distance between 2 fibres recorded in a single motor unit study was 6 m m (3 recordings) in E D C and 6-7 m m (5 recordings) and 9 m m (1 recording) in biceps brachii muscle. In about a third o f the trials the entire procedure could not be completed due to the loss o f the "reference action potential".
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DISCUSSION
The distribution of muscle fibres within the individual motor units has not previously been demonstrated in vivo in man. In previous physiological multielectrode studies of the motor unit territory in man (Buchthal et al. 1957b; Buchthal et al. 1959), the size of the individual recording surface and their relatively wide spacings did not facilitate the study of the distribution of single fibres. This problem has been overcome with the present technique with selective recordings from small electrode surfaces representing 44 sites, 300/~m apart. The validity of the results of the present method depends principally on: (1) The electrode direction. The electrode must be positioned perpendicular to the fibre direction.
299
(2) The electrode position within the muscle. All the electrode surfaces must be positioned within the muscle. In order to help position the multielectrode over the motor unit cross-section, the reference electrodes (6-10) were constructed to be at the centre of the electrode array. I f the 5 close electrodes had been at the end of the array there would be greater risk for the remaining electrodes to be positioned outside the motor unit under study. The maximal distance between electrode surfaces recording action potentials from fibres in the same motor unit is some indication of the territory, but this particular parameter was not studied in this investigation. (3) Accuracy of electrode movement. As the electrode has to be advanced 4 steps of 300 # m each during the investigation, there is a risk to lose the "reference action potential" of the m o t o r unit under study. Also, some muscle fibre damage or fibre dislocation can occur. A check of the correct electrode movement was the constancy of the response profile when the triggering electrode had been advanced from 6 to 10, i.e., the multielectrode was moved one long interelectrode distance (1.2 mm) (Fig. 4). The fibre distribution gave an impression of centering with the highest fibre density around the centre of the multielectrode. Two methodological factors are primarily responsible. Firstly, one of the central electrodes (number 6) was used as a reference electrode recording activity from at least 1 fibre of the motor unit. Secondly, as this reference electrode was always inside the motor unit, the chance of being within the territory of the same m o t o r unit is highest for the closely adjoining electrodes. I f the multielectrode was positioned in the middle of the motor unit many electrodes would be within the m o t o r unit territory but if the multielectrode were placed more peripherally, only a few of the more central electrodes could theoretically record activity from that motor unit. The number o f action potentials over the reference electrode tended to characterize the m o t o r unit under study. When only a single response was obtained as the "reference action potential" less additional fibres were recorded in the corridor than when multiple fibres were recorded over the reference electrode. This was particularly evident in the biceps brachii muscle. This probably indicates a higher fibre concentration in some m o t o r units than in others. Whether these motor units have on average a higher total number of fibres cannot be determined. It is known that m o t o r units are of different size with a positive correlation to activation threshold and motor neurone size (Wuerker, McPhedran and Henneman 1965; Burke, Levine, Zajac, Tsaris and Engel 1971). In the present investigation, motor units with low threshold were studied to limit disturbances from other motor units. However, other m o t o r units were often recorded during the course of a single investigation, some with a higher innervation rate, indicating a lower activation threshold than the one under study (St~lberg and Thiele, 1973; Hannertz 1974). There were also some differences between the muscle fibre distribution in motor units from the 2 muscles. In biceps brachii the motor unit tended to be more dispersed. There was a slightly lower fibre density over the reference electrode in biceps brachii than in E D C although there was a similar number of fibres in the corridor
300 for the 2 muscles and a larger maximal distance between fibres recorded from motor" units in biceps brachii than in EDC. The present study has demonstrated that in the normal EDC and biceps muscles the muscle fibres in a single motor unit are scattered similarly to those shown in experimental animals (Edstr6m and Kugelberg 1968; Brandstater and Lambert 1969; Doyle and Mayer 1969) with glycogen depletion experiments. A single motor unit's distribution covers about 17 ~ of the cross-sectioned area in the rat tibialis anterior muscle (Kugelberg 1973). In such studies it was found in rats that the fibres of ~t single motor unit are usually isolated, with 76 ~o of the fibres not adjacent to another fibre of the same motor unit and only rarely 3 fibres together (Brandstater and Lambert 1969). These figures are roughly comparable to the fibre density values obtained with single fibre E M G (St~dberg and Thiele 1975) where only a single fibre is recorded in about 60-65 )/o of random electrode placements. In the present investigation there were never more than 3 fibres over 1 recording electrode in the approximately 300 active electrodes in 101 motor units studied. With conventional histochemistry the principal fibre types can be identified but a motor unit's distribution cannot be made. A mosaic of fibre type distribution in the cross section area of the muscle in man (Engel 1966; Jennekens, Tomlinson and Walton 1971) also indicated scattering although the individual motor units were not identified. In a single case with myokymia, glycogen-depleted fibres of the one or more active motor units were randomly scattered (Williamsson and Brooke 1972). The presented multielectrode investigations have shown the distribution of motor unit fibres in the normal EDC and biceps brachii. The fibres were dispersed in a manner similar to that shown in experimental glycogen depletion studies. In the following paper the changes in motor units with reinnervation are presented (Schwartz, St~dberg, Schiller and Thiele 1976). ACKNOWLEDGEMENTS We thank G6sta Lov6n who made the electrode and Kerstin Mellqvist who assisted during the experiments and made the drawings.
REFERENCES Brandstater, M. E. and E. H. Lambert (1969) A histochemical study of the spatial arrangement of muscle fibers in single motor units within rat tibialis anterior muscle, Bull. Amer. Ass. EMG Electrodiagn., 82: 15-16. Buchthal, F. and P. Rosenfalck (1973) On the structure of motor units. In: J. E. Desmedt (Ed.), New Developments in Electromyography and Clinical Neurophysiology, Vol. 1, Karger, Basel, pp. 71-85. Buchthal, F., F. Erminio and P. Rosenfalck (1959) Motor unit territory in different human muscles, Acta physioL scand., 45: 72-87. Buchthal, F., C. Gutd and P. Ro~nfalck (1957a) Volume conduction of the spike of the motor unit potential investigated with a new type of multielectrode, .4cta physiol, scand., 38: 331-354. Buchthal, F., C. Guld and P. Rosenfalck (1957b) Multielectrode study of the territory of a motor unit, Acta physioL scand., 39: 83-104.
301 Burke, R. E., D. N. Levine, F. E. Zajac, P. Tsaris and W. K. Engel (1971) Mammalian motor units Physiological-histochemical correlation in three types in rat gastrocnemius, Science, 174: 709-712. Doyle, A. M. and R. F. Mayer (1969) Studies of the motor units in the cat, Bull. Sch. Med. Maryland, 54: 11-17. Edstr6m, L. and E. Kugelberg (1968) Histochemical composition, distribution of fibres and fatiguability of single motor units, J. NeuroL Neurosurg. Psychiat., 31 : 424-433. Ekstedt, J. (1964) Human single fibre action potentials, Actaphysiol. scand., 61 : Suppl. 226, pp. 1-96. Engel, W. H. (1966) Histochemistry of neuromuscular diseases - - Significance of muscle fibre types. In : Neuromuscular Diseases, VoL 2 (Proceedings of the 8th International Congress of Neurology, Vienna), pp. 67-101. Hannertz, J. (1974) Discharge properties of motor units in relation to recruitment order in voluntary contraction, Acta physiol, scand., 91: 374-384. Jennekens, F. G. I., B. E. Tomlinson and J. N. Walton (1971) Data on the distribution of fibre types in five h u m a n limb muscles, J. neurol. Sci., 14" 245-257. Krnjevic, K. and R. Miledi (1958) Motor units in the rat diaphragm, J. Physiol. (Lond.), 140: 427439. Kugelberg, E. (1973) Properties of the rat hind-limb motor units. In: J. E. Desmedt (Ed.), New Developments in Electromyography and Clinical Neurophysiology, Vol. 1, Karger, Basel, pp. 213. Norris, F. H. and R. L. Irwin (1961) Motor unit area in rat muscle, Amer. J. Physiol., 200: 944-946. Schwartz, M. S., E. St~,lberg, H. H. Schiller and B. Thiele (1976) The reinnervated motor unit in man - - A single fibre E M G multielectrode investigation, J. neurol. Sci., 27: 303-312. Sthlberg, E. and B. Thiele (1973) Discharge pattern of motor neurones in humans. In: J. E. Desmedt (Ed.). New Developments in Electromyography and Clinical Neurophysiology, VoL 3, Karger, Basel, pp. 234-241. St~dberg, E. and B. Thiele (1975) Motor unit fibre density in the extensor digitorum communis muscles - - Single fibre electromyographic study in normal subjects at different ages, J. Neurol. Neurosurg. Psychiat., 38: 874-880. Sthlberg, E., J. Ekstedt and A. Broman (1970) The electromyographic jitter in normal human muscles, Electroenceph. clin. Neurophysiol., 31: 429438. Williamsson, E. and M. H. Brooke (1972) Myokymia and the motor unit, Acta physioL scand., 26: 11-16. Wuerker, E. B., A. M. McPhedran and E. Henneman (1965) Properties of motor units in a heterogenous pale muscle (m. gastrocnemius) of the cat, J. Neurophysiol., 28 : 85-99. -
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