The effect of deuterium oxide on junctional membrane permeability to dichlorofluorescein was examined to determine the mode of transfer of the dye from one cell interior to another in the septate giant axon of earthworm. Dichlorofluorescein was shown to diffuse through the nexus passively and in a hydrated form. Additionally, evidence suggested an alteration of the cell-to-cell channel structure by deuterium/hydrogen exchange. Dichlorofluorescein was rendered impermeant at 6 degrees C in D2O and 4 degrees C in H2O. Action potentials, however, were capable of propagation from cell to cell at 4 degrees C in D2O and H2O. The results are consistent with a hydrophilic channel where solute molecules diffuse through the junction (nexus) in a hydrated form. The temperature blocks are presumably brought about by increasing hydration shells around solute and channel proteins with cooling until the solute is rendered too large to diffuse.