Development of a Caenorhabditis elegans exercise model for future application of functional genomics

• Grant number: 16/25583-7
• Support Opportunities: Scholarships in Brazil - Scientific Initiation
• Start date: April 01, 2017
• End date: March 31, 2018
• Field of knowledge: Biological Sciences - Physiology - Physiology of Organs and Systems
• Principal Investigator: Julio Cesar Batista Ferreira
• Grantee: Carlos Alberto da Silva Gomes Ferreira
• Host Institution: Instituto de Ciências Biomédicas (ICB). Universidade de São Paulo (USP). São Paulo , SP, Brazil

Abstract

Physical inactivity is a risk factor for degenerative disease. For this reason, exercise is considered a critical tool for the maintenance of quality of life. Despite of its physiological benefits, the intracellular mechanisms related to the adaptive response induced by exercise remain unknown. There are several experimental models to study the impact of exercise in physiology (phenotype). However, none of them allows a longitudinal, functional, global and integrative analysis of the intracellular signaling involved in exercise adaptation. Measuring either transcripts or protein levels is the major global approach to identify potential candidates involved in exercise response. However, this transversal approach is generally associative, which impairs identification of new targets involved in exercise adaptation. Additionally, the high cost to genetically manipulate the experimental models available make the target validation even harder. According, the development of a more robust, fast, longitudinal and functional experimental model is essential for a better understanding of the intracellular mechanisms involved in exercise adaptation. This study aims to develop a Caenorhabditis elegans (C. elegans) exercise model. The establishment of an exercise model for this nematode will allow the study of exercise-related mechanisms. C. elegans has many advantages as experimental model, such as easy genetic manipulation, high homology with the human genome, transparency (allowing analysis of the integrated physiology in real time) and suitability for large-scale functional genomic analysis using RNAi libraries (genome-wide functional screens). In the first part of this study we will establish a platform for the evaluation of the C. elegans locomotor performance in a liquid environment. C. elegans is a soil nematode cultured in solid media in the laboratory. However, upon exposition to a liquid environment it performs wave swimming movements. To evaluate C. elegans locomotor performance in liquid environment, wild type worms (strain N2) will be individually transferred from culture plates (solid medium) to a 96 well plate containing M9 buffer (liquid medium). Then, we will record them for 10 min using a stereoscope. The acquired videos will be fragmented and the images analyzed by using the software worm-tracker and Image J. The follow parameters will be measured: 1. Frequency of body waves, 2. Maximum swimming distance, 3. Averaged speed and 4. Quiescence time. We will validate our findings by using genetic and pharmacologic interventions. Accordingly, locomotor capacity of C. elegans mutants nuo-6 (qm200) and isp-1 (qm150), which present deficits in mitochondrial oxidative capacity, and wild worms N2 incubated with different mitochondrial stressors (sodium azide and doxycycline) will be analyzed in order to verify whether the impairment of mitochondrial metabolism reduces the C. elegans locomotor performance. To further verify the efficiency of the platform for evaluation of the C. elegans locomotor performance, animals will be incubated with caffeine (positive control) during the test. In the second part of this study, we will evaluate the effect of continuous and intermittent (mimicking exercise) C. elegans exposure to the liquid medium on locomotor performance and metabolism. In the third part of this study, we will establish an exercise training program for C. elegans in the liquid environment. N2 wild type worm will be daily exposed to liquid medium throughout life (25 days). The effectiveness of exercise training program will be evaluated by measuring locomotor performance and life span. Our preliminary results are promising and suggest that the platform for evaluation of the C. elegans locomotor performance is effective. It is worth mentioning that the establishment of an exercise model for C. elegans will allow us perform a genome-wide functional screening to identify the genes involved in the adaptive response to exercise. (AU)