Partial Isolation of Two Classes of Dopamine β‐Hydroxylase‐Containing Particles Undergoing Rapid Axonal Transport in Rat Sciatic Nerve

Abstract: The rapid bidirectional transport of dopamine β‐hydroxylase (DBH) in adrenergic axons provides a means of analyzing the life cycle of adrenergic storage vesicles. We compared the physical characteristics of DBH‐containing particles traveling to or returning from the terminal varicosities of ligated rat sciatic nerves. Density gradient centrifu‐gation and Sephacryl S1000 gel‐permeation chromatography were used to fractionate extracts from nerve segments proximal or distal to the ligatures. A series of experiments indicated the existence of at least two populations of rapidly transported DBH‐containing particles, a “light” 85‐nm particle and a larger “dense” 120‐nm particle. The 85‐nm particles were prevalent in unligated nerve, but accounted for only one‐third of the total anterogradely transported DBH activity accumulated after 18 h. The 120‐nm particles were barely detectable in unligated nerve, but they accumulated at twice the rate of the 85‐nm particles and accounted for the rest of the anterogradely transported particulate DBH activity. These two populations of particles were readily isolated from proximal nerve extracts by sucrose density gradient centrifugation. Similar‐appearing dense and light peaks of particulate DBH activity were obtained from distal nerve extracts. Much of the retrogradely transported DBH of the extracts, however, was associated with large particles (>300 nm) not resolved by Sephacryl SI000. Retrogradely transported exogenous NGF was found only in the dense sucrose gradient peak. We propose that the 85‐nm DBH‐containing particles correspond to “large dense‐cored vesicles,” and that the 120‐nm particles are derived from the dense tubules visualized in adrenergic nerves by the chromaffin reaction.