Toxic and teratologic effects of verapamil on early chick embryos: Evidence for the involvement of calcium in neural tube closure

Toxic and teratologic effects of verapamil, a calcium antagonist, on chick embryos explanted at stage 8 (four‐somite stage) and cultured for 6–8 hours were investigated. In general, embryos responded to verapamil in a dose‐related manner. Concentrations lower than 2 μg/ml had no apparent effect on the development of embryos. A concentration of 15 μg/ml significantly increased the incidence of embryos (approximately 80% of viable embryos) with neural tube closure defects and less numerous somites. Higher concentrations (e.g., 30 μg/ml) were embryotoxic and over 90% of the embryos were either severely malformed or dead after 8 hours of incubation. Compared to controls, verapamil‐treated neuroepithelial cells had smoother apical surfaces and less conspicuous microfilament bundles. The deleterious effects of verapamil (15 μg/ml) could be reversed by subculturing the affected embryos, within 3 hours of treatment, on nutrient medium alone or on nutrient medium containing 25 μg/ml chlorotetracycline (CTC), a calcium agonist, the latter being more effective provided that treatment did not exceed 4 hours. Exposure of the developing neuroepithelium to 15 μg/ml verapamil for 3–4 hours resulted in a significant reduction in free Ca²⁺ levels, as revealed by the pyroantimonate precipitation method, throughout neuroepithelial cells. Overall results suggest that verapamil causes neural tube closure defects by reducing intracellular free Ca²⁺ levels, thereby relaxing apical microfilament bundles of developing neuroepithelial cells.