Abstract
The principles of synapse formation have been largely unraveled by investigations at the somatic neuromuscular junction (NMJ). Such experiments are commonly performed on placental mammals (e.g. mouse), where all four limbs develop on a similar timescale. By contrast, some Australian marsupials have very segmented limb development, where forelimbs are precociously developed to assist the journey to the pouch, while hindlimbs are comparatively unformed. We explored the implications of this novel developmental pattern for the morphological development of marsupial NMJs. Western Grey Kangaroo neonates (n = 8) aged postnatal day (P) 0-100 were ethically sourced from licensed shooters. Multiple (5-9) muscles from each specimen were embedded, cryosectioned, and stained immunohistochemically for NMJ compartment proteins. Confocal micrographs of adult kangaroo NMJs revealed small oval endplates with a central internal perforation. Standard indices of synapse maturation (e.g. ACh receptor clustering) confirmed significant NMJ development from P0-P100. Notably, forelimb NMJs were considerably more developed than hindlimb junctions at P0. From P0-100, kangaroo forelimb NMJs developed at a slower rate than hindlimb NMJs, so that all limbs were similarly developed by P100. Interestingly, within the hindlimb, a detailed comparison of 5 muscles suggested a single, homogenous developmental profile, in contrast with recent studies in mouse suggesting both fast and delayed synapsing muscle populations (Pun et al., 2002). The segmented nature of NMJ development in Western Grey Kangaroos combined with the relative accessibility provided by a protracted maturation in the pouch, suggest that marsupials may present novel opportunities for studying synapse maturation compared with placental mammals.