Adaptations of diaphragm neuromuscular junction following inactivity

We hypothesized that differences exist in the morphological adaptations of neuromuscular junctions (NMJs) on different fiber types in response to prolonged inactivation. Two weeks of inactivity of both phrenic motoneurons and diaphragm muscle was induced by spinal cord hemitransection at C2 (spinal isolation; SI). A three-color fluorescent immunocytochemical technique, combined with laser-scanning confocal microscopy, was used to create two- (2D) and three-dimensional (3D) images of NMJs and obtain morphological information concerning: (1) innervating axons and presynaptic nerve terminals; (2) motor endplates (postsynaptic apparatus consisting of acetylcholine receptors), and (3) myosin heavy chain (MHC) phenotype of muscle fibers. In both sham controls (CTL) and SI animals, planar (2D) and surface (3D) areas of motor endplates and nerve terminals on type II muscle fibers (anti-fast MHC immunoreactive) were smaller than on type I (nonimmunoreactive to anti-fast MHC) fibers, when normalized for fiber diameter. The number of branches, total branch length and perimeter of both motor endplates and nerve terminals were greater for NMJs on type II fibers than on type I fibers. The extent of overlap between nerve terminal and endplate was greater on type I fibers than on type II fibers. After SI, there was a significant expansion of NMJs on type II fibers. Planar and surface areas of motor endplates and nerve terminals, number of endplate and nerve terminal branches, total branch length, and perimeter were all increased on type II fibers following SI. The extent of overlap of nerve terminal and endplate increased on type II fibers, approaching that observed in type I fiber NMJs. These results indicate that neuromuscular inactivation leads to a selective expansion of type II fiber NMJs through addition of new terminal area, and elongation of existing terminal branches. These changes may represent a compensatory effort to improve neuromuscular transmission.