The development of interneurons in the chick embryo spinal cord following in vivo treatment with retinoic acid

Abstract

To investigate the role of retinoic acid (RA) in the development of interneurons in the spinal cord, we examined the expression of cellular retinoic acid binding protein type I (CRABP I). The earliest developing interneurons in the chick spinal cord can be divided into two major groups: circumferential (C) neurons and primitive longitudinal (PL) neurons. In brachial segments, both types of interneurons began to express CRABP I at stage (st.) 13+ of the V. Hamburger and H.L. Hamilton (1951, J. Morphol. 88:49-92) stage series, which is before the onset of axonogenesis. Subsequently, with the onset of axonal outgrowth, C neurons and PL neurons expressed CRABP I in their cell bodies, axons, and growth cones. The expression of CRABP I was developmentally regulated. CRABP I immunoreactivity gradually decreased after st. 36 (embryonic day [E] 10) such that no interneurons expressed this protein by E21. The transient expression of CRABP I during a period of intensive axonal growth suggested that RA may be involved in the development of interneurons. To test this idea, we implanted an all-trans RA-containing ion exchange bead into either rostral segments of the spinal cord at st. 12-13 or into caudal segments at st. 15-16, all stages that are well before the appearance of CRABP-I-positive neurons in these segments. In the RA-treated spinal cord, increased numbers of pyknotic cells were found predominantly in dorsal regions, presumably reflecting the death of neuroepithelial cells, C neurons and premigratory neural crest cells. Surviving C neurons in the RA-treated spinal cord extended their axons ventrally toward the floor plate as in control embryos. PL neurons also projected their axons rostrally or caudally in the RA-treated spinal cord, similarly to control embryos. However, the proportion of caudally projecting PL neurons was significantly increased in segments rostral to the RA-containing bead. These results suggest that RA may regulate the survival and axonal orientation (directionality) of subpopulations of spinal interneurons.