The University of Maine DigitalCommons@UMaine Electronic Theses and Dissertations Fogler Library 8-2020 Zebrafish as a Model ot Understand the Impact of Inactivity and Neuromuscular Electrical Stimulation on Duchenne Muscular Dystrophy Elisabeth Kilroy University of Maine, [email protected] Follow this and additional works at: https://digitalcommons.library.umaine.edu/etd Part of the Exercise Physiology Commons, Exercise Science Commons, and the Musculoskeletal Diseases Commons Recommended Citation Kilroy, Elisabeth, "Zebrafish as a Model ot Understand the Impact of Inactivity and Neuromuscular Electrical Stimulation on Duchenne Muscular Dystrophy" (2020). Electronic Theses and Dissertations. 3232. https://digitalcommons.library.umaine.edu/etd/3232 This Open-Access Thesis is brought to you for free and open access by DigitalCommons@UMaine. It has been accepted for inclusion in Electronic Theses and Dissertations by an authorized administrator of DigitalCommons@UMaine. For more information, please contact [email protected]. ZEBRAFISH AS A MODEL TO UNDERSTAND THE IMPACT OF INACTIVITY AND NEUROMUSCULAR ELECTRICAL STIMULATION ON DUCHENNE MUSCULAR DYSTROPHY By Elisabeth Ann Kilroy B.S. College of Charleston, 2014 A DISSERTATION SubmitteD in Partial Fulfillment of the Requirements for the Degree of Doctor of Philosophy (in BiomeDical Science) The GraDuate School The University of Maine August 2020 Advisory Committee: Clarissa Henry, Professor of Biological Sciences, University of Maine, Advisor Gregory Cox, Associate Professor, The Jackson Laboratory Thane Fremouw, Associate Professor of Psychology, University of Maine Roger Sher, Research Associate Professor of Neurobiology, Stony Brook University Calvin Vary, Faculty Scientist III, Maine Medical Center Research Institute Ó 2020 Elisabeth Ann Kilroy All Rights ReserveD ii ZEBRAFISH AS A MODEL TO UNDERSTAND THE IMPACT OF INACTIVITY AND NMES ON DUCHENNE MUSCULAR DYSTROPHY By Elisabeth Ann Kilroy Dissertation Advisor: Dr. Clarissa Henry An Abstract of the Dissertation Presented in Partial Fulfillment of the Requirements for the Degree of Doctor of Philosophy (in BiomeDical Science) August 2020 Skeletal muscle plasticity is imperative for functional aDaptation to changing Demands in activity. Although a great Deal is known about the structural anD functional plasticity of healthy skeletal muscle, far less is known about plasticity in DiseaseD muscle. Here, we combineD the power of the zebrafish moDel with the aDaptability of neuromuscular electrical stimulation (NMES) to study the basic mechanisms of plasticity in the zebrafish model of Duchenne Muscular Dystrophy (DMD). Four NMES paradigms, defineD by their frequency, delay, anD voltage, were designed to emulate the repetition and load schemes of human resistance training programs. AdDitionally, two inactivity paraDigms were Designed to emulate activity patterns in individuals with DMD. Three sessions of enDurance NMES improve muscle structure, increase swim velocity and distance traveleD, anD extenD survival. EnDurance NMES significantly increaseD the number anD length of branching for neuromuscular junctions. Nuclear surface area and volume also significantly increased following endurance NMES. Time-lapse imaging suggests less degeneration and improved regeneration of the fast-twitch muscle fibers. Conversely, three days of inactivity worsen muscle structure anD Decreases survival. Strikingly, inactivity followeD by a single session of endurance or power NMES obliterates muscle resilience. Therefore, our data clearly indicate that, at least in the zebrafish model, some resistance training is beneficial whereas inactivity is Deleterious for Dystrophic muscle. More importantly, though, our Data provide a new methodology with which to study muscle plasticity in healthy and diseased muscle. DEDICATION To all the chilDren anD aDults fighting back against muscular Dystrophy, to their caregivers anD to their aDvocates, this is for you. Your strength ignites a fiery passion inside me that will never, ever be dimmed until cures are found. Adapt anD overcome, always. iii ACKNOWLEDGEMENTS To the GraDuate School of BiomeDical Science anD Engineering, thank you. You provided me with a graduate experience that surpassed all of my expectations. You supported me as I ventureD Down unchartereD territories in graDuate eDucation anD professional development. To the Admissions Committee, thank you for looking past all the undergraduate courses I did not take and all the experiences I did not have. Thank you for believing in me anD my mission to cure muscular Dystrophy. If I haD to choose to attenD graDuate school all over again, I would choose GSBSE always. To my aDvisor, Dr. Clarissa Henry, and committee members, Drs. Greg Cox, Cal Vary, Roger Sher, anD Thane Fremouw, thank you for challenging me, for teaching me how to ask questions, and for showing me how to confiDently aDmit that I do not know the answer. Thank you for allowing me to chase this crazy idea of mine without any proof that it coulD work. Clarissa, thank you for all of the laughs anD tears, and for the aDventures in Italy anD Costa Rica, incluDing the bajillion hours spent in the airport. Thank you for pushing me to be inDepenDent from my very first day. Thank you for giving me time to pursue my passions for higher education, advocacy and spending time with those fighting muscular dystrophy. Thank you for always volunteering me for speaking engagements and tackling the toughest challenges. Finally, thank you for allowing me to have open access to your refrigerator, washer and dryer, hair brush, and snack drawer. I promise that you did not eat your snacks as fast as you thought you did. To all those who assisteD me on this project, these experiments woulD not be possible without you. To the members of the Henry Lab, both past and present, thank you for allowing me to share this space and grow into a scientist. I promise to never steal a pipette again. To all the unDergraDuate stuDents that worked alongside me, especially Kaylee Brann, Devon Varney, Amelie GermonD, anD Claire Schaffer, thank you for bringing your energy, your stories, anD your love for science to the lab each anD every Day. You are all exceptional human beings anD I am iv excited to continue watching you grow into the best versions of yourselves. Dr. Josh Kelley anD AhmeD Almaghasilah, thank you for your image analysis expertise anD proviDing me with superior methods for quantifying muscle health and neuromuscular junctions. Dr. Karissa Tilbury and Jordan Miner, thank you for sharing your biomedical engineering expertise with me and using SHG to provide an in-depth analysis of the sarcomeres. Drs. Ben King anD Joy-El Talbot, anD KoDey Silknitter, thank you for navigating the large data sets that accompanieD the RNAseq experiment and providing me numerous pathways to chase. Drs. JareD Talbot anD Susan Amacher, thank you for sharing your transgenic 3MuscleGlow zebrafish lines and making in vivo time-lapse analyses possible. To Mark Nilan, thank you for outstanding care and maintenance of the zebrafish facility. With your help I went from one male and one female zebrafish to way too many zebrafish. To my mother and stepfather, Julie and Russ Moser, thank you. Thank you for the car and moped that got me to and from the laboratory. Thank you for allowing me to go grocery shopping in your refrigerator and pantry and for the hundreds of loads of laundry that I did at your house. Momma, thank you for giving me pep talks, for always encouraging me to keep my head down and focused on my goals, and for the jackets and wool socks that I borrowed and faileD to return. Russ, thank you for finDing the errors in my scripts, for the reminDers to shut down my computer, and for the homemaDe beers and handcrafteD furniture. To my father, Dr. Timothy Kilroy, thank you for forcing me to apply to UMaine and paying for the application fee. Thank you for always showing me the importance of eDucation, networking and professional growth. Thank you for reminDing me to take breaks and to strive for progress over perfection. Let it be known that you are no longer the real Dr. Kilroy. To my brothers, Killian, Keegan and Konlon Kilroy, thank you for shaping me into the competitive and committed teammate that I am today. Keegan, thank you for helping me design the NMES parameters anD the gym. Thank you for always being one phone call away anD offering your engineering expertise to help me move this project forward. Killian, thank you for v paying for my cell phone bill to make these critical phone calls. Konlon, thank you for allowing me to Drive across the country with you. I love watching you grow and will never forget to turn up for the weekends. To Gabriel Jensen, thank you for everything. Your daily support, passion for science, and ability to make me belly laugh filleD my gratituDe cup every single day. To think that you were once a stranger crashing on my couch to now my fiancé and best best friend is an increDible feeling. For over three years, we woke up together, went to lab together, took classes together, ate lunch together, anD went home together. I never imagineD spenDing that much time with a single human being. You helpeD me work through my Data and experimental design, you cheered for me at every presentation, you cooked me dinner, anD you unDerstooD that five minutes in lab coulD mean an hour or more. I cannot wait to spenD the rest of my life with you. To Scott Delcourt anD Drs. KoDy Varahramyan and Joan Ferrini-Mundy, thank you for providing me with everything I needed to succeed as a graduate student and in my journey to becoming the best researcher that I can be. The commitment that you three have to growing and improving graduate education at UMaine is beyond anything I have ever witnesseD. Scott, thank you for teaching me how to be compassionate and how to be an exceptional mentor.