Differential muscle‐driven synaptic remodeling in the neuromuscular junction after denervation

Abstract

We used knock-in mice that express green fluorescent protein (GFP)-labeled embryonic-type acetylcholine receptors to investigate postsynaptic responses to denervation of fast-twitch and slow-twitch muscle fibers, and to visualize the integration of newly synthesized GFP-labeled embryonic-type receptors into adult synapses. The embryonic-type receptors are transiently expressed and incorporated into the denervated endplates. They replaced synaptic adult-type receptors in a directed fashion, starting from the endplate's periphery and proceeding to its central regions. The progress of embryonic-type receptor expression with respect to transcriptional control is a transient, short-term activation mechanism. The less pronounced increase in the expression levels of the GFP-labeled receptors revealed a differential shift in the integration and degradation processes that constitute the dynamic equilibrium of the synaptic receptor pool. Therefore, we were able to model the changes in the total receptor load of the neuromuscular endplate following denervation as a function of the abundance of available receptors and the initial receptor load of the endplate.

Role of autophagy, SQSTM1, SH3GLB1, and TRIM63 in the turnover of nicotinic acetylcholine receptors

“…11,12 After their arrival at the membrane, CHRN can either persist there or get endocytosed in vesicular carriers 13,14 from where it might be either recycled back to the membrane or get degraded [15][16][17] in lysosomes. 18 While the expression of CHRN displays strong upregulation under muscle wasting conditions, 19,20 such as immobilization and denervation, its metabolic stability decreases at the same time, 10,21-23 implicating post-transcriptional regulation in CHRN turnover under stress. Indeed, CHRN is subject to a wealth of regulatory post-translational modifications ranging from phosphorylation 24,25 to ubiquitination.…”

The Neuromuscular Junction: Measuring Synapse Size, Fragmentation and Changes in Synaptic Protein Density Using Confocal Fluorescence Microscopy

“…The method is limited, however, by the requirement that all samples are processed for immunofluorescence then imaged during the same confocal session. A recent study by Yampolsky et al 5 described a method for measuring endplate AChR density that might help overcome this limitation. In this study, the same fields of endplates were imaged at several different laser power settings.…”

“…In this study, the same fields of endplates were imaged at several different laser power settings. The slope of the relationship between laser power and rhodamine-BGT fluorescence intensity was used to assess relative changes in AChR density at endplates in different mice 5 . This method might be useful for comparing AChR intensity in samples imaged at different times over the course of a prolonged study.…”

“…It is required for all voluntary movement. Fluorescence microscopy has long been used to study the effects of transgenes on the mouse NMJ [1][2][3] or to compare the effects of age, diet, exercise and disease upon rodent NMJs [4][5][6][7][8][9][10][11] . Such studies have taught us much about the physiology and pathophysiology of the NMJ, but the diverse parameters reported (e.g., AChR area, endplate area, perimeter length, fragmentation indices) often make it difficult to compare the findings of these studies.…”

Expression profile of plakin cross-linking proteins in short-term denervated mouse hindlimb skeletal muscle

“…In denervated skeletal muscle fibers, the expression of AChR-ε subunit is suppressed, and that of the embryonic AChR-γ variant is transiently reexpressed in extrasynaptic regions, peaking at ~5 days postdenervation and returning to levels observed in innervated muscle after 20 days postdenervation. [52][53][54] The fact that denervation appears to "reboot" the developmental expression profile of MACF, even transiently, may suggest that MACF potentially contributes to organizing elements of the postsynaptic region of neuromuscular junctions prior to innervation. Additional studies are required to confirm this notion.…”