Oregon Health & Science University
Slow channel syndrome (SCS) is a congenital form of myasthenia resulting from point mutations in muscle nicotinic acetylcholine receptor (AChR) subunits. These mutations prolong the open duration of the AChR, and consequently increase the decay time constant of endplate currents. Clinically, SCS is characterized in patients by muscle weakness and fatigue, muscle degeneration and endplate myopathy. Although symptoms can be managed with pharmaceutical interventions, there is currently no cure for this progressively debilitating disease. The zebrafish motility mutant twister carries a point mutation in the alpha (Î±) subunit of the AChR that recapitulates both the aberrant single receptor properties and compromised voluntary movement seen in humans. Unlike the human condition, however, symptoms manifest transiently, and animals undergo a homeostatic recovery process that renders them asymptomatic for the remainder of their lifespan. The goal of this dissertation work is to progress our understanding of the pathogenesis of SCS by examining the consequences of the mutation in zebrafish and uncover the mechanism of this recovery process. Electrophysiological recording of synaptic currents during the period of compromised motility reveals that two unique kinetic components are introduced to the decay process. Addition of these two components results in persistent current entry and maintained muscle depolarization, preventing the cell from following the high-frequency trains of action potentials necessary for swimming. These components, termed intermediate and slow, have time constants that are approximately 10-and 100-fold longer than those recorded at wild-type neuromuscular junctions. The molecular basis for these two components was determined by reconstitution of single AChRs. These studies revealed that the gating of the embryonic AChR isoform accounts for the slow kinetic component; gating of the adult AChR isoform underlies the intermediate component. Additionally, both receptor subtypes displayed an increased apparent affinity to acetylcholine, and significant numbers of spontaneous openings were observed in the embryonic form. Recovery from SCS, and improvement of swimming function in zebrafish occurs through the natural developmental switch from the embryonic (Î³) to the adult (Îµ) subunit. Reducing the fraction of embryonic AChRs at the synapse effectively accelerates synaptic current decay and allows the muscle cell to follow high-frequency trains of action potentials. When Î³-subunit translation was blocked using morpholinos, the recovery process was accelerated, supporting the finding that the embryonic subunit is responsible for the motility defect. This distinction makes twister both a genetic model for SCS and a functional model for Escobar Syndrome, a myasthenic syndrome resulting from mutations in the y-subunit. Although gene knockdown technology was effective in the zebrafish, it has limited application in humans. Therefore, we tested quinidine, a long-lived open-channel blocker currently administered to human patients. These studies demonstrated that quinidine improved neuromuscular function in zebrafish as well. We further demonstrate that the mode of quinidine action is an acceleration of synaptic current decay to near wild type levels. Additionally, quinidine was shown to be an effective blocker of the persistently open embryonic AChR.
Program in Neuroscience
School of Medicine
Walogorsky, Michael D., "Self repair in a zebrafish model for slow-channel myasthenic syndrome" (2012). Scholar Archive. 856.