This work was performed at Faculty of Veterinary Medicine, Utrecht University, The Netherlands. This work has been presented at BEVA 2009: Proceedings 28th BEVA congress, Equine Vet J Ltd, Newmarket.
Corresponding author: Dr I.D. Wijnberg, Specialist KNMvD Equine Internal Medicine, Dipl. ECEIM, Department of Equine Internal Medicine, Faculty of Veterinary Medicine, Utrecht University, Yalelaan 114, 3584 CM Utrecht, The Netherlands; e-mail: email@example.com.
Background: Reference values for quantitative electromyography (QEMG) in neck muscles of Royal Dutch Sport horses are lacking.
Objective: Determine normative data on quantitative motor unit action potential (QMUP) analysis of serratus ventralis cervicis (SV) and brachiocephalicus (BC) muscle.
Animals: Seven adult normal horses (mean age 9.5 standard deviation [SD] ± 2.3 years, mean height 1.64 SD ± 4.5 cm, and mean rectal temperature 37.6 SD ± 0.3°C).
Methods: An observational study on QMUP analysis in 6 segments of each muscle was performed with commercial electromyography equipment. Measurements were made according to formerly published methods. Natural logarithm transformed data were tested with ANOVA and posthoc testing according to Bonferroni.
Results: Mean duration, amplitude, phases, turns, area, and size index (SI) did not differ significantly among the 6 segments in each muscle. Mean amplitude, number of phases, and SI were significantly (P < .002) higher in SV than BC, 520 versus 448 μV, 3.0 versus 2.8 μV, and 0.48 versus 0.30 μV, respectively. In SV 95% confidence intervals (CI) for amplitude, duration, number of phases, turns, polyphasia area, and SI were 488–551 μV, 4.3–4.6 ms, 2.9–3.0, 2.4–2.6, 7–12%, 382–448, and 0.26–0.70, respectively; in BC this was 412–483 μV, 4.3–4.7 ms, 2.7–2.8, 2.4–2.6, 4–7%, 393–469, and 0.27–0.34, respectively. Maximal voluntary activity expressed by turns/second did not differ significantly between SV and BC with a 95% CI of 132–173 and 137–198, respectively.
Conclusions and Clinical Importance: The establishment of normative data makes objective QEMG of paraspinal muscles in horses suspected of cervical neurogenic disorders possible. Differences between muscles should be taken into account.
Motor unit action potential (MUP) analysis by quantitative electromyography (QEMG) has been used since the 1950s in human medicine. Since then, modern automated sampling techniques have expanded possibilities by adding new variables and examining sensitivity and specificity for different disorders.1–8 Semiquantitative MUP analysis of equine paraspinal neck muscles proved to be a helpful diagnostic tool in diagnosing for example cervical disease in horses with locomotion disturbances of unknown origin, stringhalt,9 or suspected cervical lesions.10 Traditionally, quantitative MUP analysis involves calculation of mean values of MUP variables such as amplitude, duration, number of phases and turns, and presence of polyphasic or complex MUPs. In addition, presence of pathological spontaneous activity (PSA; eg, positive sharp waves, fibrillation, complex repetitive discharges, or neuromyotonia) is evaluated, as well as insertional activity patterns.5,6,8,11–16 In addition to manual or (semi)automatic quantitative motor unit action potential analysis at weak effort,8 (auto) analysis of interference patterns at stronger effort are explored.1,4,5,16 The evaluation of above mentioned variables enables comparison with normative data. Former research indicated that QEMG could be useful in localizing and evaluating equine neck lesions that induced neurogenic damage of the lower motor neurons such as arthropathy of the cervical vertebral joints.9,10 However, these data were based on semiquantitative analyzing techniques because normative data were lacking. Qualitative visual analysis of MUPs and interference patterns may be diagnostic in clear cut changes, but may be misleading in patients with more subtle lesions.8,17 Other than the conventional MUP variables such as amplitude, duration number of phases and number of turns, newer variables such as MUP area, size index (SI), MUP thickness, and interference pattern analysis (IPA) are being studied in human medicine for diagnostic power. Their potential advantage is to be more reliable and less sensitive to signal noise or to interoperator differences.2,17,18
This indicated the need for more normative data than those established in several equine muscle such as subclavian,11–13 lateral vastus, triceps,12–14 and descending pectoral muscles19 so far. Normative data can be helpful in diagnosing or localizing neuromuscular disorders of the neck.9,10 This report describes normative data on automatic MUP analysis including recently introduced MUP variables such as SI, MUP area, and IPA in serratus ventralis cervicis (SV) and brachiocephalicus (BC) muscle enabling more detailed quantitative evaluation for cervical diseases.20
Materials and Methods
Seven adult clinically healthy Royal Dutch Sport horses (mean age 9.5 standard deviation [SD] ± 2.3 years, mean height 1.64 SD ± 4.5 cm, rectal temperature 37.6 SD ± 0.3°C) from the herd of Utrecht University were used. Exclusion criteria were abnormalities on neurological and orthopedic examination. In addition, radiological examination of cervical vertebrae had to be judged as normal by a Diplomat of the European College of Veterinary Diagnostic Imaging. In both muscles (M SV and M BC) 6 segments were measured in unsedated horses. Because the BC cannot be separated anatomically from the M omotransversarius (OT) in the caudal third of the muscle, the EMG data from this segment can originate from both BC or OT or a mixture. Segments between 2 vertebrae were based on palpation of the transverse processes (Fig 1). The study was approved by the Committee of Animal Welfare, Utrecht University.
Details on definition, materials, and methods of QEMG examination have been described in former publications.11–13 In brief, EMG signals were recorded with a portable apparatusa and 26 G concentric EMG needle electrodes.b Band pass was between 5 Hz and 10 kHz. Sweep speed was 10–20 ms/division. Amplifier gain was 50–100 μV for spontaneous activity and 100–500 μV for MUP recording. A surgical pad attached to the horse with a girdle and connected to the preamplifier served as the ground electrode.
Quantitative MUP Analysis
Insertional activity, PSA, MUPs, and satellite potentials were segmentally recorded in each muscle. PSA was assessed outside the endplate region in the same regions in which MUPs were obtained and included fibrillation potentials, positive sharp waves, complex repetitive discharges, or (neuro)myotonia. It was considered indicative of pathology if present in 2 or more locations. At least 3 insertions and 3 directions per insertion were made per investigated muscle. The needle was redirected several times, and by selecting sharp sounding MUPs while the needle was withdrawn with 3-mm increments, sampling was performed throughout the muscle. MUPs were selected partly in a semiautomatic way, using a trigger line that selects identical MUPs above chosen amplitude. The automatic MUPs selection was corrected where indicated manually off line (noice excluded). End point of MUP duration was corrected by on screen visual assessment.16,17 Amplitude, duration, number of phases, and number of turns were obtained from 20 to 30 MUPs per muscle, with a maximal rise time of 0.8 ms rise and identically firing at least 4 times. Mean percentage of polyphasic MUPs was calculated of both muscles. Numbers of turns were calculated manually using on screen analysis. In addition, not previously described parameters such as SI and MUP area were recorded and analyzed. MUP area was calculated automatically. SI was calculated automatically by the EMG software by the formula 2 × log (amplitude) + area/amplitude.
Low frequency filter was set at 20 Hz, high frequency filter at 10 KHz, and sampling frequency of at least 25 Hz.16 Thirty contractions at random force per segment in each muscle were recorded with standard concentric needle electrodesb and evaluated. Interference patterns were analyzed measuring maximal voluntary activity (MVA) expressed by turns/second and cloud analysis.4,5 The EMG software calculated these variables automatically.
Data on MUP variables were ANOVA with posthoc testing according to Bonferroni on natural logarithm transformed data was used as statistical method by SPPS 15.0/16.0. Significance was set at <0.05. Descriptive statistics were used to describe the data. Geometric mean values of MUP variables were derived from back transformation of the Ln transformed data.
In both SV and BC, prolonged or absent insertional activity was not measured nor were PSA or abundant satellite potentials identified. Ln transformation of data resulted in a normal distribution enabling statistical analysis. Within both muscles, mean duration, amplitude, number of phases and turns, area, and SI did not significantly differ among the 6 segments. However, in SV polyphasia was higher (P < .03) in the segment between the 3rd and 4th vertebrae compared with the other segments. Mean amplitude, number of phases, and SI were significantly (P < .002) higher (520 versus 448 μV, 3.0 versus 2.8, and 0.48 versus 0.30) in the SV muscle compared with the BC muscle (Fig 2, Table 1). Ninety-five percent CI intervals of amplitude, duration, number of phases, turns, and polyphasia were in the SV 488–551 μV, 4.3–4.6 ms, 2.9–3.0, 2.4–2 6, and 7–12%, respectively. In the BC this was 412–483 μV, 4.3–4.7 ms, 2.7–2.8, 2.4–2.6, and 4–7%, respectively (Fig 2). Complex MUPs (>5 turns) were absent. Of the newer variables, SI but not MUP area, was significantly higher in the SC than in BC. In SV 95% CI intervals of area and SI were 382–448 and 0.26–0.70, respectively. In BC they were 393–469 and 0.27–0.34, respectively.
Table 1. Geometric mean values of MUP variables of serratus ventralis cervicis and brachiocephalicus muscle.
IPA showed that MVA did not differ significantly among muscles with a 95% CI of 132–173 and 137–198 turns/second, respectively. Projecting 7 individual interference patterns expressed as amplitude/turns plotted against turns/second of all 7 horses per muscle on top of each other, which produced a cloud of measuring points. Each cloud was presented in the lower left corner of the figure (Fig 3). The center of gravity of measuring points seemed different between muscle (Fig 3B, Fig 4B). The normal limit in humans is set at less than 15% of the total number of points outside the normal boundaries.
Normative data and 95% intervals of conventional and modern MUP variables were established in the present study. Muscle differences, but not segmental difference, were present with exception for a higher percent polyphasic MUPs seen at C3-4 segment in the SV muscle. The difference was significant but compared with QEMG data in other muscles the values can still be in the physiological range. Ninety-five percent intervals of MUP amplitude and number of phases were higher in the SV than in the BC muscle. Number of phases was computed automatically in contrast to the former study where the autoanalysis of both number of phases and turns was manually corrected. Because the new software did not count number of turns, this was performed manually, resulting in lower turns than phases, which is not logical. As a consequence we can say that not only the muscle itself but also the technical equipment could contribute to differences of the data.6 An increase in number of turns is a variable that, together with MUP duration, is an early determinator of neurogenic disease. Diagnosing subtle neurogenic changes probably necessitates therefore using normative data “belonging” to the equipment used. It should be realized that the QEMG data in the caudal third of the muscle can be a mixture of BC (cleidobrachialis part) and OT muscle as mentioned in “Materials and Methods” section.
Looking at 1 MUP variable instead of the combination of all makes clinical use easier, resulting in human studies evaluating the sensitivity of sole variables in discriminating between neuro- and myopathies.2,3,15,21–23 MUP duration corresponds to a larger uptake zone (2.5 mm in radius) than area (2 mm) or MUP amplitude (0.5 mm) and thus is largely determined by the number of muscle fibers.17 Changes in number of contributing muscle fibers show up as changes in MUP duration. This fact is responsible for the finding that according to some authors MUP duration is the feature that best distinguished between neuropathy and myopathy even though others consider amplitude a better variable.3,17 However, the use of MUP duration as sole variable is time consuming if used in the correct and truly quantitative way.17 A sole robust MUP parameter is therefore searched in human studies.
It appears that the ratio area/amplitude, which is determined by MUP duration and the shape of the MUP (this ratio is expressed as the so-called MUP thickness), might be such a factor. MUP area is determined by the number of muscle fibers that are up to 2 mm from the recording electrode. It can be imagined as the product of height (amplitude) and thickness. Differences in MUP area should reflect differences in the number of muscle fibers in a rather large portion of the motor unit. The ratio area /amplitude is named thickness and its advantage is being less sensitive to electrode position than area or amplitude alone. It is considered a robust parameter that is less sensitive to the operator's judgment. By combining all, the diagnostic sensitivity and specificity can be further increased for myopathies.2,17
The SI describes the relation between amplitude and area and its value becomes larger when amplitude and the thickness increase. The SI is (automatically) calculated from the MUP amplitude and the area/amplitude ratio (thickness). The term size index was introduced by Sonoo and Stalberg17 and is the normalized thickness, eg, the area/amplitude that is expected if the needle position is manipulated so that the MUP amplitude will be 1 mV. It remains constant throughout needle movement, which makes it less dependent of operator skills. In practical QEMG this means that differentiation between normal and neurogenic MUP is possible by SI irrespective of needle position. In addition it enables making a diagnosis by measuring a small number of MUPs.17
In order to decide if and what kind of pathology is present and to give a prognosis, QEMG in at least 2 or 3 muscles by MUP analysis applied at weak effort and IPA at stronger effort is recommended.8,24 MUP variables as described above evaluate the MUP generated at low level force resulting in a bias toward fibers that are recruited 1st (type I fibers).25,26 Evaluation of muscle function at higher force necessitates analysis of maximal voluntary contraction at random and fixed force because this provides additional information about the character of disease. Weak muscles generate higher firing frequency or early recruitment in order to compensate for the loss of functional units.26 Another way of evaluation at higher force is IPA using automated turns/second analysis, amplitude/turn analysis, or plotting the mean amplitude as function of turns in the form of which the latter excludes the necessity of measurement of force.5,27 Turns/second analysis shows that a decrease in duration leads to less summation and cancellation of small spikes and therefore high number of turns/second, whereas a long duration leads to a low number of turns/second. The number of turns/second increases and mean amplitude per turn decreases in myopathic muscles, whereas the opposite has been found in neurogenic muscles in man. However, the sensitivity of this technique is better if force is standardized7,24 making this not suitable for application in the horse.
The so-called cloud analysis1,4,5,24 generated from IPA by plotting the mean amplitude as function of turns (Figs 3, 4) enables discrimination in myopathy, neuropathy, or normal and has the advantage that it is generated at random muscle force. In human medicine a muscle is considered abnormal if more than 10% of the analyzed epochs are outside the upper centile amplitude/activity cloud.1,5 IPA proved to be more sensitive than mean MUP duration analysis in discriminating between normal and abnormal and between neuropathy and myopathy if the normal limit was set at <15% of the total number of points outside the normal boundaries.5 In disorders directly affecting the muscle, the diseased muscle fibers have to fire at a higher frequency in order to maintain a given muscle force, leading to a full interference pattern of low amplitude at relatively low force. In neuropathies such as axonal loss or conduction block, if motor units are lost, the remaining motor units have to fire at a higher rate in order to compensate for the loss of potential to recruit motor unit activity resulting in a poor interference pattern with the few remaining motor units firing at high frequency although this phenomenon can also be seen in poor cooperation, upper motor neuron lesions, or end-stage myopathies.7,8,28 This sensitive application is interesting because in horses, signs similar to humans with cervical radiculopathy have been documented in association with radiographic abnormalities of the cervical vertebrae29–31 and needle EMG is a helpful tool in identifying the relevance of an abnormality found in, for example, SV and BC muscle.9,10,19
Compared with historical data in former studies in lateral vastus, triceps, and subclavian muscles of adult Royal Dutch Sport horses13 the amplitude is higher than measured in the formerly described muscles. This suggests the presence of a higher percentage of the larger type I muscle fibers in the measured neck muscles because amplitude is positively correlated with muscle fiber size.13 Data on muscle fiber typing of these muscles are lacking but it would be logical considering the function of the SV and BC and their almost continuous EMG activity resulting from the antigravity activity in relation to the head position.32 The larger contribution in the SV muscle compared with the BC muscle in antigravity function33 might be responsible for its largest MUP amplitude in the SV muscle.
The study presented above provides normative values for SV and BC muscles in adult horses and information on newer/more recently developed QEMG variables such as SI, MUP area, and IPA. The normative data can be helpful in diagnosing or localizing neuromuscular disorders of the neck. For example, neurogenic damage to motor nerves of the cervical paraspinal nerve roots (cervical part SV) or the distal part of the BC muscle (cleidobrachialis part that is innervated by axillary nerve originating from cervical nerve root segment C 7–8*) innervating the muscle segment will result in QEMG alterations.9,10 As indicated in the literature,6 normative data are preferably created for each institute, type of equipment, or both; however, we believe that these differences will be too small to intervene with clinical utility because the difference with disease is expected to be large. To what extent the area, SI, and cloud analysis will be superior above conventional QEMG in horses remains unclear until tested in clinical applications.
a Viking Quest EMG system, Nicolet Biomedical Inc, Madison, WI
b Length, 50 mm; diameter 0.45 mm; sampling area 0.07 mm2, Nicolet Biomedical Inc