Muscle Velocity Recovery Cycles to Examine Muscle Membrane Properties
Summary
Presented here is a protocol for the recording of muscle velocity recovery cycles (MVRCs), a new method of examining muscle membrane properties. MVRCs enable in vivo assessment of muscle membrane potential and alterations in muscle ion channel function in relation to pathology, and it enables the demonstration of muscle depolarization in neurogenic muscles.
Abstract
Although conventional nerve conduction studies (NCS) and electromyography (EMG) are suitable for the diagnosis of neuromuscular disorders, they provide limited information about muscle fiber membrane properties and underlying disease mechanisms. Muscle velocity recovery cycles (MVRCs) illustrate how the velocity of a muscle action potential depends on the time after a preceding action potential. MVRCs are closely related to changes in membrane potential that follow an action potential, thereby providing information about muscle fiber membrane properties. MVRCs may be recorded quickly and easily by direct stimulation and recording from multi-fiber bundles in vivo. MVRCs have been helpful in understanding disease mechanisms in several neuromuscular disorders. Studies in patients with channelopathies have demonstrated the different effects of specific ion channel mutations on muscle excitability. MVRCs have been previously tested in patients with neurogenic muscles. In this prior study, muscle relative refraction period (MRRP) was prolonged, and early supernormality (ESN) and late supernormality (LSN) were reduced in patients compared to healthy controls. Thereby, MVRCs can provide in vivo evidence of membrane depolarization in intact human muscle fibers that underlie their reduced excitability. The protocol presented here describes how to record MVRCs and analyze the recordings. MVRCs can serve as a fast, simple, and useful method for revealing disease mechanisms across a broad range of neuromuscular disorders.
Introduction
Nerve conduction studies (NCS) and electromyography (EMG) are the conventional electrophysiological methods used for the diagnosis of neuromuscular disorders. NCS enables detection of axonal loss and demyelination in the nerves1, while EMG can differentiate whether myopathy or neurogenic changes are present in the muscle due to nerve damage. However, NCS or EMG provide limited information about muscle fiber membrane properties and underlying disease mechanisms. This information can be achieved using intracellular electrodes in isolated muscles from muscle biopsies2,3,4. However, it is of clinical importance to use methodologies using recordings from intact muscles in patients.
The velocity of a second muscle fiber action potential changes as a function of the delay after the first5, and this velocity recovery function (or recovery cycle) has been shown to change in dystrophic or denervated muscles. The yield of such recordings from single muscle fibers was, however, too low to be of use as a clinical tool6. However, Z'Graggen and Bostock later found that multi-fiber recordings, obtained by direct stimulation and recording from the same bundle of muscle fibers, provide a fast and simple method of obtaining such recordings in vivo7. A sequence of paired pulse electrical stimuli with varying interstimulus intervals (ISIs) is used in this method7,8,9,10,11.
The evaluated MVRC parameters include the following: 1) muscle relative refractory period (MRRP), which is the duration after a muscle action potential until the next action potential can be elicited; 2) early supernormality (ESN); and 3) late supernormality (LSN). ESN and LSN are the periods after the refractory period in which the action potentials are conducted along the muscle membrane faster than normal. The depolarizing afterpotential, and potassium accumulation in the t-tubules of the muscle respectively, are hypothesized as the main causes for the two periods of supernormality.
The wide applicability of MVRCs to muscle disorders has been shown in detecting membrane depolarization in ischemia7,10,12 and renal failure13, as well as providing information about muscle membrane abnormalities in critical illness myopathy14 and inclusion body myositis15. Frequency ramp and intermittent 15 Hz and 20 Hz simulation protocols have since been introduced. MVRCs, together with these additional protocols, have demonstrated the different effects on muscle membrane excitability related to loss-of-function or gain-of-function mutations in various muscle ion channels in the inherited muscle ion channelopathies (i.e., sodium channel myotonia, paramyotonia congenita16, myotonic dystrophy17, Andersen-Tawil syndrome18, and myotonia congenita19,20).
In a recent study, the applicability of MVRCs to neurogenic muscles was shown for the first time. The term "neurogenic muscle" refers to the secondary changes in skeletal muscles that develop as denervation and reinnervation after any injury to the anterior horn cells or motor axons. Denervation is characterized in EMG as spontaneous activity (i.e., fibrillations [fibs] and positive sharp waves [psws]), while large motor unit potentials with prolonged duration and increased amplitude present reinnervation21. EMG changes are evident in denervated muscles, but the underlying cellular changes in muscle fiber membrane potentials have only been demonstrated in experimental studies on isolated muscle tissue2,3,4. MVRCs provide further insight into in vivo human muscle membrane properties regarding the denervation process.
This paper describes the methodology of MVRCs in detail. It also summarizes the changes in neurogenic muscles in a subgroup of patients from a previously reported study22 and healthy control subjects that enables determination of whether the method is appropriate for a planned study.
The recordings are performing using a recording protocol that is part of a software program. Other equipment used is an isolated linear bipolar constant current stimulator, 50 Hz noise eliminator, isolated electromyography amplifier, and analogue-to-digital converter.
Protocol
Representative Results
Discussion
MVRCs, as programmed in the recording software, is a highly automated procedure, but care is needed to obtain reliable results. In the recording stage, while adjusting the needles, it is important to avoid stimulating the end-plate zone or nerve. This usually leads to large twitches of the whole muscle, which increases the risk of displacement of the stimulation and/or recording needle during recording MVRCs. To date, the method has been applied to several muscles that have better described end-plate zone; however, the e…
Divulgaciones
The authors have nothing to disclose.
Acknowledgements
This study was financially supported mainly by the two grants from Lundbeck Foundation (Grant number R191-2015-931 and Grant number R290-2018-751). Additionally, the study was financially supported by Novo Nordisk Foundation Challenge Programme (Grant number NNF14OC0011633) as part of the International Diabetic Neuropathy Consortium.
Materials
| 50 Hz Noise Eliminator | Digitimer Ltd | Humbug | |
| Analogue-to-Digital Converter | National Instruments | NI-6221 | |
| Analysing software program | Digitimer Ltd (copyright Institute of Neurology, University College, London) | QtracP, MANAL9 | |
| Disposable concentric needle electrode, 25 mm x 30G | Natus | Dantec DCN | |
| Disposable monopolar needle electrode, 25 mm x 26G | Natus | TECA elite | |
| Isolated EMG amplifier | Digitimer Ltd | D440 | |
| Isolated linear bipolar constant-current stimulator | Digitimer Ltd | DS5 | |
| Software and recording protocol | Digitimer Ltd (copyright Institute of Neurology, University College, London) | QtracW software, M3REC3 recording protocol written by Hugh Bostock, Istitute of Neurology, London, UK) |
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