Changes in morphology and behaviour of retinal growth cones before and after crossing the midline of the mouse chiasm – a confocal microscopy study

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Abstract
The growth of retinal axons was investigated in different regions of the optic chiasm in C57 pigmented mouse embryos aged embryonic day 13 (E13) to E15. Individual retinal axons and their growth cones were labelled anterogradely by DiI and imaged using a confocal imaging system. In aldehyde‐fixed embryos, retinal growth cones display a simple form in the optic nerve and become more complex in morphology in the chiasm. The complex form is particularly prominent in those axons that turn to the ipsilateral tract in the premidline region of chiasm. Moreover, complex growth cones are also commonly found in axons in the postmidline chiasm, which are markedly different in morphology from those axons in the premidline region, suggesting that the postmidline chiasm contains a novel environment for the pathfinding of retinal axons. In another experiment, the dynamic growth of retinal axons is studied in a brain slice preparation of the living retinofugal pathway. Retinal axons show an intermittent growth across the premidline and postmidline chiasm. Extensive remodelling of growth cone form followed by a shift in growth direction is commonly seen during the pause periods, indicating that signals that guide axon growth across the chiasm are not restricted to the midline, but are laid down throughout the chiasm. Moreover, dramatic changes in axon trajectory are noted first at the premidline chiasm where the uncrossed axons segregate from the crossed axons, and second at the postmidline chiasm where specific sorting of retinal axons according to their position in the dorsal ventral retinal axis and their ages are known to take place. These results show that there are two distinct environments, separated by the midline in the chiasm, where axons show different responses to local guidance cues and develop the distinct fibre orders.