Functional Isolation of Single Motor Units of Rat Medial Gastrocnemius Muscle

Summary

This method allows the recording of the force of twitch and tetanic contractions and action potentials in three types of motor units in the rat medial gastrocnemius muscle. The functional isolation of a single motor unit is induced by electrical stimulation of the axon.

Abstract

This work outlines functional isolation of motor units (MUs), a standard electrophysiological method for determining characteristics of motor units in hindlimb muscles (such as the medial gastrocnemius, soleus, or plantaris muscle) in experimental rats. A crucial element of the method is the application of electrical stimuli delivered to a motor axon isolated from the ventral root. The stimuli may be delivered at constant or variable inter-pulse intervals. This method is suitable for experiments on animals at varying stages of maturity (young, adult or old). Moreover, this protocol can be used in experiments studying variability and plasticity of motor units evoked by a large spectrum of interventions. The results of these experiments may both augment basic knowledge in muscle physiology and be translated into practical applications. This procedure focuses on the surgical preparation for the recording and stimulation of MUs, with an emphasis on the necessary steps to achieve preparation stability and reproducibility of results.

Introduction

Motor units (MUs) are the smallest functional units of skeletal muscles. Therefore, understanding their function, plasticity and contractile properties, as well as the mechanisms of their force regulation, is crucial for progress in muscle physiology. The basic contractile properties of MUs and the proportions of their physiological types have been documented for numerous muscles, predominantly the hindlimb muscles in experimental animals. However, both the plasticity of MU properties and the mechanisms of MU force regulation are still not fully understood.

The principle of the described method is extensive denervation of the hindlimb muscl....

Protocol

All procedures need to be approved by the local ethics committee and adhered to the European Union guidelines on animal care as well as the national law on the protection of animals.

NOTE: Each experimenter involved in this procedure must be trained in basic surgical procedures and has to obtain a valid license for performing animal experiments.

1. Anesthesia

1. Anesthetize the rat with an intraperitoneal injection of sodium pentobarbital (an initial dose of 60 mg·kg^-1).
2. After approximately 5 min, check the depth of anesthesia by pinching the rat’s ear or forelimb with blu....

Results

Parameters of motor unit contractions and action potentials can be calculated on the basis of recordings when stable conditions of recordings are ensured. Figure 1 presents a representative recording of the single twitch of a fast MU. The upper trace shows the motor unit action potential. The delay between stimulus delivery and onset of the motor unit action potential is due to conduction time from ventral root to muscle. Figure 2 shows a representative recordin.......

Discussion

If performed correctly by experienced scientists, the surgical component of the described protocol should be completed within approximately two hours. One should take particular care to maintain stable physiological conditions of the animal during the surgery, particularly body temperature and depth of anesthesia, which should be systematically controlled by assessing pinna and withdrawal reflexes. Following the surgery, it should be possible to maintain stable recording conditions for at least six hours.

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
Force transducer	custom-made
Forceps	Fine Science Tools	No. 11255-20	Dumont #55 with extra light and fine shanks
Forceps	Fine Science Tools	No. 11150-10	Extra Fine Greafe Forceps
Forceps	Fine Science Tools	No. 11026-15	Special cupped pattern for superior grip
Forceps	Fine Science Tools	No. 11023-10	Slim 1x2 teeth
Forceps	Fine Science Tools	No. 11251-20	Dumont #5
Hemostats	Fine Science Tools	No. 13003-10	Hartman
Isolation Unit	Grass Instruments	S1U5A
Low Noise Bioamplifer	World Precision Instruments	Order code 74030
Needle holders	Fine Science Tools	No. 12503-15	With tungsten carbide jaws
Rongeurs	Fine Science Tools	No. 16021-14	Friedman-Pearson
Scissors	Fine Science Tools	No. 14101-14	Straight sharp/blunt with large finger loops
Scissors	Fine Science Tools	No. 14075-11	Curved blunt/blunt
Scissors	Fine Science Tools	No. 14084-08	Extra fine bonn
Scissors	Fine Science Tools	No. 15000-00	Straight, ideal for cutting nerves
Stimulator	Grass Instruments	S88	Dual Output Square Pulse Stimulator