In 2,5-hexanedione (2,5-HD)-induced axonal neuropathy, the rate of neurofilament (NF) transport increases in optic axons. To test the prediction that increases in the rate of polymer transport in any one locality of the axon lead directly to a decrease in the number of NF in that locality, NF and microtubules (MT) were quantitatively analyzed in axonal cross sections. In 2,5-HD axons the number of NF was 38% of that in control axons while the number of MT was not significantly changed; it appears that the drug treatment decreases NF number in the proximal axon regions, most directly through an increase in rate of NF transport. In those regions, the cross-sectional areas of the 2,5-HD-treated axons were 45% smaller than those of control axons; although the axons had shrunk in diameter, they retained their normal cylindrical shapes as measured by the index of circularity. Reduced internal expansive forces in the axon, working in conjunction with the normal external compressive forces, appear to reduce the radius of the axon. Quantitative analyses demonstrated that the average and the maximum lateral spacings between NF-NF, NF-MT, and MT-MT were all 30% larger in 2,5-HD-treated axons than in control axons. This suggests that polymers are relatively free to move laterally away from one another and to fill the available space within the axon. These observations are not consistent with models wherein 2,5-HD acts to crosslink the NF into an immobile network that can no longer advance within the axon. Instead, it appears more likely that 2,5-HD acts selectively on the interaction between some NF and the slow transport mechanism to increase the rate of NF transport.