Abstract
Purpose: The somatic neuromuscular junction (NMJ) is a valuable experimental model of synapse formation. Such experiments are most 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 have compared the morphological development of the NMJ in kangaroo hindlimbs and forelimbs, to investigate how NMJ development proceeds in an animal with this novel developmental pattern.
Methods: Three Western Grey Kangaroo neonates aged postnatal day (P) 0, 26 and 100 were ethically sourced from licensed shooters in South-Western Australia. 5 forelimb and 4 hindlimb muscles from each specimen were embedded, cryosectioned, and stained immunohistochemically for NMJ compartment proteins.
Results: Confocal micrographs of adult kangaroo NMJs revealed an unusual oval endplate with a internal perforation at its centre. When markers of NMJ development described in rodents (e.g. neurofilament distribution, ACh receptor clustering) were analysed in P0 neonates, forelimb NMJs were considerably more developed than hindlimb junctions. However, between P0 and 100, NMJs in kangaroo forelimbs developed at a slower rate than hindlimb NMJs, so that all limbs were similarly developed by P100.
Conclusions: Our preliminary data indicates that the segmented limb development in Western Grey Kangaroos is reflected at the level of the NMJ. This segmented NMJ development, combined with the relative accessibility of kangaroo neonates (which undergo a large degree of maturation in the pouch), present novel opportunities for studying synaptic development compared with placental mammals.