Axonal Migration of Proteins in the Central Nervous System and Peripheral Nerves as Shown by Radioautography.

The "axonal flow" theory holds that the nerve cell body is continuously forming new cytoplasm which enters the axon and travels along its length. This theory was formulated by Weiss and collaborators (Weiss and Hiscoe, '48) to account for the accumulation of material in front of a nerve constriction and for the high concentration of enzymes appearing in the proximal stump of a sectioned nerve (Sawyer, '46; Hebb and Waites, '56; Friede, '59). For instance, since the so-called neurosecretory substance of the hypothalamus accumulates in the central stump of the severed pituitary stalk, this substance is believed to migrate down into the posterior pituitary (Scharrer and Scharrer, '54). Techniques that do not damage the nerves have been used in a search for direct evidence in support of the axonal flow theory. Thus, radioactive substances were given, which it was hoped would be taken up into the nerve cell body and flow down the axon: phosphate-PJ2 (Samuels et al., '51; Ochs and Burger, '58; Ochs et al., '62), glucose-Ci4 (Waelsch, quoted by Weiss, '60) , and labeled amino acids (Waelsch, '58; Koenig, '58b; Schultze and Oehlert, '58; Verne and Droz, '60). However, the radioactive substances used were taken up not only by nerve cell bodies, but also by Schwann cells. Hence the difference in radioactivity in successive segments of nerve trunks was small and Weiss ('60) commented that, while the results obtained "were expected to lend deeper insight into the processes involved, this expectation has thus far remained disappointingly unfulfilled.'' In fact, several authors hold the view that there is no axonal migration at all (Schultze and Oehlert, '58; Koenig and Koelle, '60; Miani, '60). At the present time, an experimental approach is needed which will either prove or disprove the axonal flow theory. It was thought that the attempts at tracing amino acids into the proteins of neurons might be repeated, but in such a way that the radioactivity of individual axons could be distinguished from that of glial cells. This aim might be achieved with tritium labeled amino acids, since a high radioautographic resolution is obtained with tritium. The first step was to make sure of the sites of protein synthesis. That such synthesis takes place continuously in the central nervous system was shown biochemically by the incorporation of labeled amino acids into the protein fraction (Friedberg et al., '48; Gaitonde and Richter, '56; Palladin, '57; Vladimirov, '57; Lajtha et al., '57). Radioautography after injection of a labeled amino acid indicated that the protein synthesis takes place in the body of nerve cells (Cohn et al., '54; Fisher et al., '56; Flanigan et al., '57; Leblond et al., '57; Koenig, '58a; Oehlert et al., '58; Droz and Verne, '59). However, these experiments were carried out using amino acids labeled with isotopes which do not allow good radioautographic resolution (such as and the animals were sacrificed several hours after injection, that is, too late to be sure that the newly-formed proteins had not migrated out of their sites of synthesis (Warshawsky et al., '63). It was, therefore, decided to reinvestigate the problem using mainly tritium labeled amino acids and sacrificing the animals very soon after injection, so that newlyformed proteins would still be at their site of synthesis. Once the sites of synthesis of protein were known with certainty, it was possible