Endogenous electric current is associated with normal development of the vertebrate limb

A steady ionic current is driven out of both developing and regenerating am phibian limbs. In the developing limbs of anurans and urodeles, focal outwa rdly directed current (0.5-2 muA/cm(2)) predicts the location of mesenchyme accumulations producing the early bud. Here, we report measurements of a s imilar outwardly directed ionic current associated with the development of the limb bud in the mouse and chick embryo by using a noninvasive, self-ref erencing electrode for the measurement of extracellular current. In both th e mouse and chick embryo, flank currents were usually inwardly directed the direction of Na+ uptake by ectoderm. Outward currents associated with the mouse limb bud ranged from 0.04-10.8 muA/cm(2). Mouse limb bud and flank cu rrents were similar to those measured in amphibian larvae, because they wer e reversibly collapsed and/or reversed by application of 30 muM amiloride, a Na+ channel blocker. Unlike the amphibian embryos, flank ectoderm adjacen t to the mouse limb bud in the anterior/posterior axis was usually associat ed with outwardly directed ionic current. This raises the possibility of a different, or changing, gradient of extracellular voltage experienced by me senchyme cells in this plane of development than that observed in other reg ions of the limb bud. In the chick flank caudal to the somites, a striking reversal of the inwardly directed flank currents to very large (similar to 100 muA/cm(2)) outwardly directed currents occurred three developmental sta ges before limb bud formation. We tested the relevance of this outwardly di rected ionic current to limb formation in the chick embryo by reversing it by using an artificially applied "countercurrent" pulled through a microele ctrode inserted just beneath the caudal ectoderm of the embryo. This applic ation was performed for approximately 6 hr 2.5-3 developmental stages befor e hindlimb bud formation. This method resulted in abnormal limb formation b y the tenth day of gestation in some embryos, whereas all control embryos d eveloped normally. These data suggest an early physiological control of lim b development. (C) 2001 Wiley-Liss, Inc.