In Vivo Electrophysiological Measurement of the Rat Ulnar Nerve with Axonal Excitability Testing

Axonal excitability techniques provide a powerful tool to examine pathophysiology and biophysical changes that precede irreversible degenerative events. This manuscript demonstrates the use of these techniques on the ulnar nerve of anesthetized rats.

The overall goal of this technique is to determine a range of excitability properties for the ulnar nerve in living rats. This method can help answer key questions in the field of neuroscience such as, can we measure the pathological changes of nerve function prior to irreversible neurodegenerative events? The main advantage of this technique is that it allows the indirect assessment of peripheral nerve function in vivo enabling repeat measurements for the characterization of disease progression.

Before beginning the procedure, confirm the appropriate level of sedation by pedal withdrawal and corneal reflex in a 12-week-old female Long-Evans rat. Place the rat on a feedback controlled heat mat and insert a rectal thermometer probe to maintain the body temperature at 37 degrees Celsius. To record the compound muscle action potential, insert the reference electrode through the dorsal aspect of the fourth digit and the recording electrode through the hypothenar muscle.

Place the ground electrode through the skin on the superior aspect of the forearm between the stimulating and recording electrodes. Then insert the percutaneous stimulating needle cathode approximately four millimeters distal to the cubital tunnel at the elbow and insert the anode approximately one centimeter proximally through the skin of the axilla region. Next, use a semi-automated computer controlled axonal excitability program linked to a constant current stimulator and an amplifier to apply a one millisecond square wave pulse to the ulnar nerve with the cathode needle electrode and to record the compound muscle action potential from the hypothenar muscle.

Carefully adjust the angle and/or position of the cathode until an optimal biphasic response curve with a consistent amplitude is achieved and stabilize the cathode with a repositionable electrode holder. For threshold tracking, it is critically important that there is a proportionate response to the stimulus. A successful response is indicated by a sigmoid shape in the stimulus response curve.

To record a stimulus response curve, incrementally increase the stimulus intensity of a one millisecond impulse until a maximum response is obtained. Record multiple axonal excitability parameters including the threshold electrotonus, current threshold relationship, and recovery cycle. Then transfer the rat into an individual cage with monitoring until full recumbency.

The inward and outward rectifying currents can be assessed by depolarizing and hyper polarizing currents in the current voltage parameter. Data regarding the internodal conductances and membrane potential can be obtained by long subthreshold depolarizing and hyper polarizing currents represented by the threshold electrotonus waveform. In this representative experiment, sequential axonal excitability testing on the left and right forelimbs within 35 minutes after the loss of the pedal withdrawal reflex revealed no significant differences between the left and right ulnar nerves in the standard nerve conduction parameters, compound muscle action potential amplitude and latency nor are differences observed in the nerve excitability variables including super excitability and threshold electrotonus hyper polarizing at 90 to 100 milliseconds.

Once mastered, this technique can be completed in 20 to 30 minutes if it is performed properly. While attempting this procedure, it is important to take care to place the electrodes in their appropriate positions to ensure an accurate stimulus response curve and successful threshold tracking. After its development, this technique paved the way for researchers in the field of neuroscience to explore the biophysical properties of peripheral nerves in both the clinical setting and in animal models.

After watching this video, you should have a good understanding of how to perform a nerve excitability protocol and to obtain a range of biophysical indices in the ulnar nerve.