Initial evidence that electrical excitability is both an early aspect of neuronal differentiation and a developmentally regulated property was obtained from recordings of action potentials in vivo. Subsequently, the analysis of the underlying voltage-dependent currents during early stages of embryogenesis was facilitated by investigation of dissociated neurons and muscle cells differentiating in culture. Calcium and potassium currents play a major role in the differentiation of the action potential of Xenopus spinal neurons, and calcium influx triggers specific features of neuronal differentiation. However, the extent to which differentiation of currents in vitro parallels that in vivo is uncertain. We have undertaken a study of in vivo differentiation of these macroscopic currents in Xenopus embryos. Spinal cords were isolated from embryos at several early stages of neurogenesis. Neurons in these isolated spinal cords were accessible to patch-clamp electrodes. Neuronal currents were recorded within 1 hr to assure that the characteristics of the currents resulted from developmental events occurring in vivo prior to the experiment. Whole-cell voltage-clamp recordings from neurons in these acutely isolated and intact embryonic spinal cords demonstrate that both the delayed-rectifier and inactivating potassium current and a low-voltage-activated calcium current mature in a manner closely parallel to that observed in culture. The results validate those from the culture system and indicate that the spinal cord is another region of the CNS accessible to cellular analysis in an intact preparation.