Expression and localization of two isoforms of brain ankyrin, 440- and 220-kDa ankyrinB, were studied in the developing nervous system of the rat fetus. The 440-kDa ankyrinB appeared on as early as embryonic day 13, and its level increased progressively toward the day of birth, which was similar to the expression pattern of growth-associated protein (GAP)-43, a well-established axonal protein. On the other hand, 220-kDa ankyrinB was expressed at a low level but constitutively throughout the latter prenatal period and was a major isoform even before embryonic day 14. Whereas the localization of 440-kDa ankyrinB was essentially confined to the axons, judging from the similarity with that of GAP-43, 220-kDa ankyrinB showed a rather general distribution in neural tissue. The localization of L1, known as an ankyrinB-binding protein, was similar to that of 440-kDa ankyrinB in the brain tissue, whereas it was similar to that of 220-kDa ankyrinB in cultured neurons, suggesting that the interaction of L1 with brain ankyrins in neurons is affected by their environment.