Excess Retinoic Acid Induces Fusion of Centra by Degenerating Intervertebral Ligament Cells in Japanese flounder, Paralichthys olivaceus

In marine aquaculture fish, excessive supplement of vitamin A (VA) to zooplanktons for larval culture and experimental exposure of larvae to retinoic acid (RA: active form of VA) have been known to cause vertebral deformity. However, the tissues in the developing vertebral column that are affected by RA and the progression of vertebral deformity remain undetermined. To examine these questions, we histologically traced the progress of vertebral deformity induced by RA in Japanese flounder (Paralichthys olivaceus). Larvae were exposed to RA for 3 days at mid-metamorphosis (G-stage), a critical stage for vertebral deformity. Intervertebral ligament, which is known to form intervertebral joints in cooperation with the notochord, was severely degenerated by RA, leading to fusion of centra. During further development to adult, growth of centra was severely suppressed in an anterior–posterior direction in RA-treated fish and the notochord tissue was lost from fused centra, resulting in complete loss of intervertebral joints and fusion of centra. We conclude that RA initially damages the intervertebral ligaments, and these defects lead to fusion, narrowing of centra, and loss of intervertebral joints in the vertebral column. The cumulative effect of these modifications is a truncated body form.