To clarify the result that marked diffusional anisotropy had been found in nomyelinated nerve, and in completion of an evaluation of the role of all longitudinal axonal structures, we report NMR measurements of water diffusion in the giant axon of the squid, where diffusional anisotropy is determined by the neurofilamentary structure. The diffusion coefficients of water parallel and perpendicular to the long axis of the squid giant axon at 20 degrees C are (1.61 +/- 0.06) x 10(-5) cm2 s-1 and (1.33 +/- 0.09) x 10(-5) cm2 s-1, respectively, which yield an anisotropic diffusion ratio of 1.2 +/- 0.1. Water diffusion in the squid giant axon is therefore quite rapid and nearly isotropic, thus eliminating the possibility of a significant role for the longitudinally oriented neurofilaments in producing diffusional anisotropy within the axoplasm. In conjunction with our work on garfish nerves therefore, only membranes, either as numerous axonal membranes or as myelin (if present), remain to fulfill the role of the primary determinant of anisotropic water diffusion in nerve and in white matter.