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Wei He, Yang Sun; Single particle tracking of nicotinic acetylcholine receptors (AChRs): implication for the pathogenesis of ocular myasthenia gravis (MG). Invest. Ophthalmol. Vis. Sci. 2018;59(9):333.
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© ARVO (1962-2015); The Authors (2016-present)
Myasthenia gravis (MG) is a neuromuscular junction (NMJ) disease caused by autoimmune targeting of the acetylcholine receptors (AChRs), which are accumulated at high concentrations in the postsynaptic muscle membrane. Eighty percent of patients with myasthenia gravis (MG) initially present with the ocular muscles, leading to double vision. This study investigates the accumulation of the AChRs in the NMJ in the primary cell cultures for molecular regulations of MG.
The dynamics of AChRs on live muscle cells isolated from the Xenopus primary culture were examined using the single particle tracking (SPT) technique. Single AChRs were labeled with quantum dots coated with streptavidin, which binds to biotinylated α-bungarotoxin. The NMJ was induced on the muscle cells by the heparin-binding growth-associated molecule (HB-GAM) coated silica beads. After 2-4 hours of HB-GAM bead treatment, multiple AChRs were recorded on the muscle membrane. As a control for the polarization of cortical actin in a parallel pattern, fibronectin was stamp-printed in square shapes with different sizes and the polarized pattern for actin filaments was examined with phalloidin staining.
AChRs travel to the synaptic area in three different patterns, per SPT. Most strikingly, around 9% AChR trajectories showed clear linear trajectories toward the HB-GAM beads.The linear trajectories are localized to the plasma membrane rather than endocytosed, as shown by the quenching of the surface quantum dots induced by cell membrane impermeable quenching dyes.The linear track is guided by the cortical actin pattern. Although the linear trajectories exhibit a radiant pattern towards the beads, line shape trajectories show the parallel pattern on the muscle cells with parallel polarized cortical actin patterns.
The cortical actin networks control the accumulation of the AChRs towards the NMJ. Given that AChR signaling in NMJ is conserved between the ocular muscle and the skeletal muscle, this study shows the potential for understanding the role of cortical actins in ocular MG.
This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.
Figure 1 Single particle tracking on muscle cells coated with HB-GAM. Differently colored trajectories are superimposed on the cluster image in grayscale, cell boundaries are indicated by white lines, and the bead locations are marked by open green circles.
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