We have used organotypic cultures of embryonic rat spinal cord and dorsal root ganglia (DRG) to study the development of central projections of primary sensory afferent axons that express calcitonin gene-related peptide (CGRP). In vivo, small- and medium-diameter CGRP-positive primary afferents terminate in laminae I, II, and V of the spinal cord and do not enter the ventral horn. A similar pattern of CGRP-positive axonal projections was observed in spinal cord slices of Day 16 embryos (E16) maintained in culture for 6 days. Both intact and dissociated DRG neurons showed the same pattern of central arborization, indicating that complex intercellular interactions between DRG neurons are not required for laminar specific targeting. Furthermore, targeting to the dorsal horn and avoidance of the ventral horn was observed in isolated dorsal and ventral hemicords, suggesting that separate mechanisms mediate the avoidance of CGRP-positive axons from the ventral horn and the elaboration of the afferent arbors within the dorsal horn. CGRP-positive afferents can grow into the dorsal horn only during a brief time window. Cultures of age-matched (isochronic) DRG and spinal cord from E14, E16, and E18 animals showed the characteristic pattern of CGRP-positive axon arborization, while cultures from E20 and neonatal animals did not. Heterochronic cultures indicate that it is the age of the spinal cord, and not the age of the DRG, that determines the ability of the CGRP-positive afferents to arborize within the dorsal horn. Together these results demonstrate that cues intrinsic to the spinal cord can direct sensory projections to appropriate locations in the spinal cord.
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