Morphology of diaphragm neuromuscular junctions on different fibre types

Abstract

We hypothesize that the morphology of the neuromuscular junction on different muscle fibre types varies, reflecting differences in activation history. In the rat diaphragm muscle, we used a three-colour fluorescent immunocytochemical technique to simultaneously visualize (1) innervating axons and presynaptic nerve terminals, (2) motor endplates, and (3) myosin heavy chain isoform expression (muscle fibre type). Laser-scanning confocal microscopy was then used to optically section the triple-labelled muscle fibres, and create three-dimensional views of the neuromuscular junction. Type I fibres were innervated by the smallest axons, and type IIa, IIx and IIb fibres by progressively larger axons. Absolute planar areas of nerve terminals and endplates progressively increased from type I, IIa, IIx to IIb fibres. When normalized for fibre diameter, planar areas of nerve terminals were largest on type I fibres, with no difference among type II fibres. The normalized planar area of endplates were larger for type I and IIb fibres, compared to type IIa and IIx fibres. The three-dimensional surface area of endplates was largest on type I fibres, with no differences across type II fibres. When normalized for fibre diameter, endplate surface areas increased progressively from type I, IIa, IIx to IIb fibres. The branching patterns of both nerve terminals and endplates varied across fibre types. The number of nerve terminal and endplate branches increased progressively from type I, IIa, IIx to IIb fibres. Conversely, individual branch length was longest on type I fibres, and shortest on type IIb fibres. The extent of overlap of pre- and postsynaptic elements of the neuromuscular junction decreased progressively on type I, IIa, IIx and IIb fibres. We conclude that these morphological differences at the neuromuscular function of different fibre types reflect differences in activation history and may underlie phenotypic differences in neuromuscular transmission.

Acetylcholine Release at Neuromuscular Junctions of Adult Tottering Mice Is Controlled by N-(Cav2.2) and R-Type (Cav2.3) but Not L-Type (Cav1.2) Ca2+ Channels

“…Localization of the different Ca 2ϩ channel ␣ 1 subunits at tg and WT mouse motor nerve terminals was compared using fluorescence immunohistochemistry in the extensor digitorum longus (EDL) and triangularis sterni (TS) muscles from animals whose diaphragm was used for pharmacological studies. The EDL is a homogeneous fast twitch type muscle; thus, concerns associated with myofiber-type-dependent differences in structure or function of the neuromuscular junctions were minimized (Gertler and Robbins, 1978;Prakash et al, 1996). The thinness of the TS muscle allowed us to label neuromuscular junction structures without using cryosectioning techniques that involved prolonged exposure of the preparation to chemicals such as sucrose, yet permitted high-quality images to be obtained.…”

Fiber Types in Mammalian Skeletal Muscles

“…Combined electrophysiological recordings and determination of the loss of the fluorescent membrane dye FM1-43 loaded into synaptic vesicles yield values of 178,000 vesicles for the EDL fibers and 252,000 vesicles for the soleus fibers (73). In rat diaphragm, slow fibers are innervated by the smallest axons, whereas type 2A, 2X, and 2B fibers receive progressively larger axons (622). Absolute areas of nerve terminals and endplates progressively increase from type 1, 2A, 2X, to 2B fibers; however, when normalized for fiber diameter, the areas of nerve terminals are largest in type 1 fibers, with no difference among type 2 fibers (622).…”

Late onset muscle plasticity in the whisker pad of enucleated rats

“…Metabolic properties, myosin isoform subtypes and plate size in striate muscles are all correlated positively (22)(23)(24). In general, oxidative slow twitch type I muscle fibers have larger motor plates than those observed in association with glycolytic fast twitch type II muscle fibers (25)(26)(27); although see (28)(29)(30). We evaluated the area of the presynaptic and postsynaptic elements of the motor plate in control and enucleated rats at different ages (Fig.…”