Effect of dysautonomia on the morphofunctional aspects of skeletal muscle in mice with amyotrophic lateral sclerosis

• Grant number: 25/04792-6
• Support Opportunities: Scholarships in Brazil - Scientific Initiation
• Start date: June 01, 2025
• End date: May 31, 2026
• Field of knowledge: Biological Sciences - Physiology - Physiology of Organs and Systems
• Principal Investigator: Julio Cesar Batista Ferreira
• Grantee: João Pedro Batista Oliveira Miranda
• Host Institution: Instituto de Ciências Biomédicas (ICB). Universidade de São Paulo (USP). São Paulo , SP, Brazil

Abstract

Amyotrophic Lateral Sclerosis (ALS) is a lethal neurodegenerative disease that affects 0.01% of the global population. The disease causes intrinsic changes that lead to the death of motor neurons and muscle atrophy, resulting in muscle weakness and paralysis in more advanced stages. Clinical data show that ~50% of patients with ALS present non-motor symptoms, raising the hypothesis that dysfunction of other systems (extrinsic to the neuromuscular axis) may contribute to the onset and progression of ALS. Among the systems extrinsic to the neuromuscular axis that possibly affect the pathophysiology of ALS is an autonomic imbalance (dysautonomia). Both patients and experimental models of ALS present an autonomic imbalance resulting from hyperactivation of the sympathetic nervous system. These clinically relevant, yet still associative, data suggest that increased sympathetic nervous system activity is involved in the pathophysiology of ALS.Recent data from our laboratory demonstrate that mice with ALS induced by the expression of the mutant protein SOD1-G93A present dysautonomia in the final stage of the disease. However, the contribution of dysautonomia in the progression of ALS has not yet been defined. In this sense, considering that skeletal muscle is an important target of ALS, the present project aims to characterize the effect of chronic administration of isoproterenol (maximizing the phenotype of dysautonomia in ALS) on the functional, morphological and biochemical aspects of the skeletal muscles of mice with ALS induced by the expression of the mutant protein SOD1-G93A. Our hypothesis is that isoproterenol will aggravate the muscular effects of dysautonomia in ALS. Molecularly, this response will occur due to the desensitization of ¿2-adrenergic receptors in the skeletal muscles of animals with ALS.To this end, our experimental strategy involves the evaluation of functional, morphological and biochemical aspects of the skeletal muscles of wild-type male mice (C57BL/6J) and those with ALS (SOD1-G93A mutation), both treated daily with isoproterenol 10mg/kg/day or vehicle solution, between days 60 (pre-symptomatic phase) and 160 of the animals' lives. The evaluations will be performed in the initial and advanced stages of the disease, 80 and 160 days of life, respectively. Isoproterenol is a beta-adrenergic receptor agonist, with an action similar to norepinephrine produced by the sympathetic nervous system. Isoproterenol will be administered daily via peritoneal route at a concentration of 10 mg/kg/day, starting on day 60 of the animals' life, when ALS is asymptomatic. Our analytical strategy involves characterizing the muscle fiber typing in both the soleus (oxidative) and tibialis anterior (TA, glycolytic) muscles. We will also perform histological analyses with hematoxylin and eosin and picrosirius red to assess cross-sectional area, integrity, inflammation, and fibrosis in skeletal muscle. Finally, we will perform functional ex vivo muscle contraction tests on the EDL muscle, as previously standardized in the laboratory. The preliminary results of the project show that the proposal is feasible and compatible with a scientific initiation program. In addition, we will have the assistance of doctoral student Lisley Ramalho, who has experience in the subject and is proficient in the analytical strategies proposed in this research project.