Development of a Novel Technique for the Measurement of Neuromuscular Junction Functionality in Isotonic Conditions

Flavia Forconi; Ludovica Apa; Simona Pisu; Irene Casola; Antonio Musarò; Emanuele Rizzuto; Zaccaria Del Prete

doi:10.1007/s12195-022-00721-3

Development of a Novel Technique for the Measurement of Neuromuscular Junction Functionality in Isotonic Conditions

Cell Mol Bioeng. 2022 Apr 7;15(3):255-265. doi: 10.1007/s12195-022-00721-3. eCollection 2022 Jun.

Authors

Flavia Forconi¹, Ludovica Apa², Simona Pisu², Irene Casola¹, Antonio Musarò¹, Emanuele Rizzuto², Zaccaria Del Prete²

Affiliations

¹ DAHFMO - Unit of Histology and Medical Embryology, Sapienza University of Rome, Via Antonio Scarpa 16, Rome, Italy.
² Department of Mechanical and Aerospace Engineering, Sapienza University of Rome, Via Eudossiana 18, 00184 Rome, Italy.

Abstract

Introduction: The neuromuscular junction (NMJ) is a chemical synapse responsible for converting electrical pulses generated by the motor neuron into electrical activity in muscle fibers, and is severely impaired in various diseases, such as Amyotrophic Lateral Sclerosis (ALS). Here, we proposed a novel technique to measure, for the first time, NMJ functionality in isotonic conditions, which better reflect muscle physiological activity.

Methods: We employed the in-situ testing technique, studied a proper placing of two pairs of wire electrodes for nerve and muscle stimulation, developed an extensive testing protocol, and proposed a novel parameter, the Isotonic Neurotransmission Failure (INF), to properly capture the impairments in neurotransmission during isotonic fatigue. We employed wild-type mice to assess the feasibility of the proposed technique, and the ALS model SOD1^G93A mice to demonstrate the validity of the INF.

Results: Results confirmed the measurement accuracy in term of average value and coefficient of variation of the parameters measured through nerve stimulation in comparison with the corresponding values obtained for membrane stimulation. The INF values computed for the SOD1^G93A tibialis anterior muscles pointed out an impairment of ALS mice during the isotonic fatigue test, whereas, as expected, their resistance to fatigue was higher.

Conclusions: In this work we devised a novel technique and a new parameter for a deep assessment of NMJ functionality in isotonic conditions, including fatigue, which is the most crucial condition for the neuronal signal transmission. This technique may be applied to other animal models, to unravel the mechanisms behind muscle-nerve impairments in other neurodegenerative pathologies.

Keywords: Experimental optimal force; In-situ functional measurements; Isotonic fatigue; Isotonic neurotransmission failure; Maximum power; Membrane stimulation; Nerve stimulation; Neurotransmission failure.