The role of microtubules in axoplasmic transport in vivo

To verify whether microtubules are involved in the mechanism of axoplasmic transport in vivo, [3H]leucine was injected into ventral horns of rats, and 3 h later Ca2+ or other drugs injected into sciatic nerves. The injection of 50-200 mM Ca2+, raising intra-axoplasmic Ca2+ levels, blocked transport above the intraneural injecting site and decreased microtubular density. Conversely, injection of 10 mM EGTA lowering the intra-axoplasmic Ca2+ induced the same changes. By combining the injection of 50 mM colchicine with 25 mM Ca2+ or 5 mM EGTA, the effects were additive in that transport was weakened further or even blocked and microtubules disappeared. Therefore, microtubules seemed to be a mediator between the injected drug and the blockade of transport and Ca2+ to be a regulator of axoplasmic transport in vivo. Tubulin, a subunit of microtubules, contains SH groups and Cd2+ is a chelate of them. By injection of 50-100 mM Cd2+, transport was weakened or blocked. The sulfhydryl inhibitor, N-ethylmaleimide increased, but the sulfhydryl donor, dimercaptosuccinate, abolished the effect of Cd2+ on transport. N-ethylmaleimide also amplified the Cd2+ effect on decreasing SH group content of sciatic nerve homogenate. There were 8.7 SH groups per tubulin monomer isolated from rabbit brain. The SH groups of tubulin in vitro and microtubular density in vivo were decreased with the increase of Cd2+ concentration. All these results indicated that microtubules play a role in the mechanism of axoplasmic transport.