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. 2022 Apr 27;70(9):1605–1629. doi: 10.1002/glia.24184

FIGURE 3.

FIGURE 3

PSC mAChRs activation is not reduced at denervated NMJs in YFP.SOD1G37R animals. (a) False color confocal images of two NMJs from an YFP.SOD1G37R animal (P460), expressing yellow fluorescent protein (YFP) in all motor axons and nerve terminals (green). During the in situ Ca2+ imaging experiment, postsynaptic nAChRs were labeled with α‐bungarotoxin (α‐BTX, red) to discriminate between innervated and denervated NMJs. Note the small partially innervated area (arrow) of the NMJ at the bottom. (b) The amplitude of the EPP (nerve stimulation, lower trace) at the partially innervated NMJ (Innervated‐P, red trace) was comparable to the one generated by a nearby completely innervated NMJ (Innervated‐C; black trace). (c) Immunohistochemical staining of the same two NMJs shown in (a) with two presynaptic markers (NF‐M+ SV2, green) to further confirm the state of innervation. PSCs were labeled using S100 antibody (cyan). (d) False color confocal images of two NMJs from another YFP.SOD1G37R animal (P460), expressing YFP (green). Postsynaptic nAChRs were labeled with α‐bungarotoxin (α‐BTX, red) during the Ca2+ imaging experiments. Note that the NMJ on the left (yellow circle) was denervated as indicated by the absence of the YFP signal over the α‐BTX staining. (e) Lack of evoked EPP (nerve stimulation, lower trace) at this denervated NMJ (red trace) while EPPs were evoked at the neighboring innervated NMJ (black trace). (f) Immunohistochemical staining of the same two NMJs shown in (d) was performed with two presynaptic markers (NF‐M+ SV2, green) to further confirm the state of innervation. PSCs were labeled using S100 antibody (cyan). (g) Mean PSC Ca2+ responses (solid line) ± SEM (dotted lines) induced by local application of muscarine (10 μM) in PSCs associated with innervated (black traces) or denervated (red traces) NMJs. No significant difference was observed between the amplitude of Ca2+ responses. (h) Similar experiments on the same cells but with local applications of ATP (10 μM). Note ATP‐elicited responses were smaller at denervated NMJs. Scale bars = 20 μM. **p < .01. NMJ, neuromuscular junction; PSC, perisynaptic Schwann cell.