The axoplasmic flow has been the growing interests for scientists who intend to study the neural function. In order to make clear the mechanism of axoplasmic flow in its relation to other physiological phenomena of the axon, it seems necessary to establish a method to study the axoplasmic flow in vitro. For this purpose, an special incubation chamber separated into 6 compartments by the wedge of septums (3 mm thickness), was used in present experiments.
The rat sciatic system, including L₄-L₆ spinal cord to common peroneal as well as tibial nerve, was excised and placed in this chamber and was incubated at 37°C in 4 ml of KrebsRinger bicarbonate buffer with 1 mM of leucine. At the proximal compartment, ¹⁴C-leucine (40 mμCi/4 ml of medium) was added and the radioactivity in the distal ones was measured at a definite interval.
The results are as follows :
(1) The fast and slow transports were recognized like as in vivo. The velocities of slow and fast transports were about 4 and 576 mm/day respectively. The ratio of labeled compound in slow and fast transports was 3 to 1.
(2) The elution profiels were clearly biphasic in colum chromatography (SephadexG 25) of the labeled materials in the medium of distal compartments. The first peak was estimated the soluble protein more than 5, 000 (M.W.) and the second peak was presumabely the free leucine.
(3) The autoradiographic pictures directly demonstrate the labeled materials confined to intra-axonal spaces and that is clear-cut “plug” of radioactive materials within the axon.
(4) Some amino acids were also eligible for transport at different rates, which was highest in L-glutamate and lowest in a-aminoisobutylate. Although, L-glutamate accelerated the leucine transport, the effect of GABA on transport was variable and both glycine and D-glutamate lowered it.
(5) Leucine transport was inhibited slightly by colchicine, vinblastine or vincristine and significantly decreased under hypoxia, low glucose media or hypothermia and markedly was inhibited by cyanide or iodoacetic acid and was diminished by cycloheximide or puromycin.
(6) Chinoform, chlorpromazine, imipramine, amitriptyline streptomycin, kanamycin markedly inhibited the slow transport.
(7) This leucine transport was significantly promoted by high potassium ions and even more intensified by additional deprivation of calcium ions. These observations imply that axoplasmic flow might associate with membraneous mechanism related to calcium ions.