Ontogeny and effect of activity on proenkephalin mRNA expression during development of the chick spinal cord

Numerous studies have shown in the adult nervous system that mRNA expression can be regulated by neuronal activity. To examine the effect of activity during embryogenesis, the ontogeny of proenkephalin mRNA expression and expression following activity blockade was investigated during development of chick spinal cord. A cDNA fragment (ca. 0.5 kb) coding for chick proenkephalin was cloned and sequenced. With this cDNA, a cRNA probe was made to examine proenkephalin mRNA expression in the spinal cord during embryogenesis. Proenkephalin mRNA was expressed in spinal cord in clusters of cells located in the developing dorsal horn and intermediate lamina at the earliest stages examined (stage 22; E4). Proenkephalin-positive cells in the intermediate lamina were located immediately adjacent to the ventricular zone. At stage 28 (E6) an additional cluster of proenkephalin mRNA-positive cells was seen at the lateral border of the developing intermediate lamina. At stage 33 (E7.5-5-8) the pattern of hybridization positive cells was similar to earlier stages, but individual cells could be identified. At stage 39 (E13) densely labeled cells were seen throughout the dorsal horn and intermediate laminae including the column of Terni. To determine whether neural activity affects proenkephalin mRNA expression, d-tubocurarine (an inhibitor of neural activity) was injected into developing embryos. Following administration of d-tubocurarine a dramatic decrease was seen in proenkephalin mRNA hybridization in the dorsal horn and intermediate lamina of the spinal cord. This study demonstrates in vivo that changes in the level of neural activity can alter gene expression during embryogenesis and suggests that activity is required for expression of nervous system-specific genes.