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Pre- and postsynaptic changes in the neuromuscular junction in dystrophic mice.

Pratt SJ, Valencia AP, Le GK, Shah SB, Lovering RM - Front Physiol (2015)

Bottom Line: To examine mdx- and age-dependent changes in the relative localization of pre- and postsynaptic structures, we calculated NMJ occupancy, defined as the ratio of the footprint occupied by presynaptic vesicles vs. that of the underlying motor endplate.Finally we found an almost two-fold increase in the number of nuclei and an increase in density (nuclei/area) underlying the NMJ.These outcomes suggest substantial remodeling of the NMJ during dystrophic progression.

View Article: PubMed Central - PubMed

Affiliation: Department of Orthopaedics, University of Maryland School of Medicine Baltimore, MD, USA.

ABSTRACT
Duchenne muscular dystrophy (DMD) is a devastating neuromuscular disease in which weakness, increased susceptibility to muscle injury, and inadequate repair appear to underlie the pathology. While most attention has focused within the muscle fiber, we recently demonstrated in mdx mice (murine model for DMD) significant morphologic alterations at the motor endplate of the neuromuscular junction (NMJ) and corresponding NMJ transmission failure after injury. Here we extend these initial observations at the motor endplate to gain insight into the pre- vs. postsynaptic morphology, as well as the subsynaptic nuclei in healthy (WT) vs. mdx mice. We quantified the discontinuity and branching of the terminal nerve in adult mice. We report mdx- and age-dependent changes for discontinuity and an increase in branching when compared to WT. To examine mdx- and age-dependent changes in the relative localization of pre- and postsynaptic structures, we calculated NMJ occupancy, defined as the ratio of the footprint occupied by presynaptic vesicles vs. that of the underlying motor endplate. The normally congruent coupling between presynaptic and postsynaptic morphology was altered in mdx mice, independent of age. Finally we found an almost two-fold increase in the number of nuclei and an increase in density (nuclei/area) underlying the NMJ. These outcomes suggest substantial remodeling of the NMJ during dystrophic progression. This remodeling reflects plasticity in both pre- and postsynaptic contributors to NMJ structure, and thus perhaps also NM transmission and muscle function.

No MeSH data available.


Related in: MedlinePlus

Neuronal morphology. (A1,B1) Overlay images of wild type (WT, A1–A3) and dystrophic (mdx, B1–B3) NMJs showing the terminal aspect of the neuron (purple, neurofilament) and postsynaptic acetylcholine receptors (red, BTX). (A2,B2) Shows examples of the neuron before processing, followed by skeletonization of digital images post-processing (A3,B3). Binary images were skeletonized and the corresponding histograms describing the discontinuity (C) and branching (D) were generated as described in the Methods. Data collected from 48 NMJs (30 WT, 18 mdx). *Indicates significance compared to wild type (p < 0.05). Scale bar equals 10 μm.
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Figure 4: Neuronal morphology. (A1,B1) Overlay images of wild type (WT, A1–A3) and dystrophic (mdx, B1–B3) NMJs showing the terminal aspect of the neuron (purple, neurofilament) and postsynaptic acetylcholine receptors (red, BTX). (A2,B2) Shows examples of the neuron before processing, followed by skeletonization of digital images post-processing (A3,B3). Binary images were skeletonized and the corresponding histograms describing the discontinuity (C) and branching (D) were generated as described in the Methods. Data collected from 48 NMJs (30 WT, 18 mdx). *Indicates significance compared to wild type (p < 0.05). Scale bar equals 10 μm.

Mentions: We also examined the terminal aspect of the neuron (using antibodies against neurofilament, Figure 4, purple) in close proximity to the motor endplate (using BTX, Figure 4, red). Composite images of confocal Z-stacks were processed and analyzed using Image J. We found intra-terminal and extra-terminal (aka pre-terminal) branching (axonal sprouting) as reported by others (Santo Neto et al., 2003; Marques et al., 2007) (Figure 4B, purple). However, quantification in the current study provides significant data showing a lack of continuity (Figure 4C) and an increase in branching (Figure 4D). All morphological data are summarized in Table 1.


Pre- and postsynaptic changes in the neuromuscular junction in dystrophic mice.

Pratt SJ, Valencia AP, Le GK, Shah SB, Lovering RM - Front Physiol (2015)

Neuronal morphology. (A1,B1) Overlay images of wild type (WT, A1–A3) and dystrophic (mdx, B1–B3) NMJs showing the terminal aspect of the neuron (purple, neurofilament) and postsynaptic acetylcholine receptors (red, BTX). (A2,B2) Shows examples of the neuron before processing, followed by skeletonization of digital images post-processing (A3,B3). Binary images were skeletonized and the corresponding histograms describing the discontinuity (C) and branching (D) were generated as described in the Methods. Data collected from 48 NMJs (30 WT, 18 mdx). *Indicates significance compared to wild type (p < 0.05). Scale bar equals 10 μm.
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Figure 4: Neuronal morphology. (A1,B1) Overlay images of wild type (WT, A1–A3) and dystrophic (mdx, B1–B3) NMJs showing the terminal aspect of the neuron (purple, neurofilament) and postsynaptic acetylcholine receptors (red, BTX). (A2,B2) Shows examples of the neuron before processing, followed by skeletonization of digital images post-processing (A3,B3). Binary images were skeletonized and the corresponding histograms describing the discontinuity (C) and branching (D) were generated as described in the Methods. Data collected from 48 NMJs (30 WT, 18 mdx). *Indicates significance compared to wild type (p < 0.05). Scale bar equals 10 μm.
Mentions: We also examined the terminal aspect of the neuron (using antibodies against neurofilament, Figure 4, purple) in close proximity to the motor endplate (using BTX, Figure 4, red). Composite images of confocal Z-stacks were processed and analyzed using Image J. We found intra-terminal and extra-terminal (aka pre-terminal) branching (axonal sprouting) as reported by others (Santo Neto et al., 2003; Marques et al., 2007) (Figure 4B, purple). However, quantification in the current study provides significant data showing a lack of continuity (Figure 4C) and an increase in branching (Figure 4D). All morphological data are summarized in Table 1.

Bottom Line: To examine mdx- and age-dependent changes in the relative localization of pre- and postsynaptic structures, we calculated NMJ occupancy, defined as the ratio of the footprint occupied by presynaptic vesicles vs. that of the underlying motor endplate.Finally we found an almost two-fold increase in the number of nuclei and an increase in density (nuclei/area) underlying the NMJ.These outcomes suggest substantial remodeling of the NMJ during dystrophic progression.

View Article: PubMed Central - PubMed

Affiliation: Department of Orthopaedics, University of Maryland School of Medicine Baltimore, MD, USA.

ABSTRACT
Duchenne muscular dystrophy (DMD) is a devastating neuromuscular disease in which weakness, increased susceptibility to muscle injury, and inadequate repair appear to underlie the pathology. While most attention has focused within the muscle fiber, we recently demonstrated in mdx mice (murine model for DMD) significant morphologic alterations at the motor endplate of the neuromuscular junction (NMJ) and corresponding NMJ transmission failure after injury. Here we extend these initial observations at the motor endplate to gain insight into the pre- vs. postsynaptic morphology, as well as the subsynaptic nuclei in healthy (WT) vs. mdx mice. We quantified the discontinuity and branching of the terminal nerve in adult mice. We report mdx- and age-dependent changes for discontinuity and an increase in branching when compared to WT. To examine mdx- and age-dependent changes in the relative localization of pre- and postsynaptic structures, we calculated NMJ occupancy, defined as the ratio of the footprint occupied by presynaptic vesicles vs. that of the underlying motor endplate. The normally congruent coupling between presynaptic and postsynaptic morphology was altered in mdx mice, independent of age. Finally we found an almost two-fold increase in the number of nuclei and an increase in density (nuclei/area) underlying the NMJ. These outcomes suggest substantial remodeling of the NMJ during dystrophic progression. This remodeling reflects plasticity in both pre- and postsynaptic contributors to NMJ structure, and thus perhaps also NM transmission and muscle function.

No MeSH data available.


Related in: MedlinePlus