The seperation of catecholamine storage vesicles from rat heart

The concept that acetylcholine is released from certain presynaptic nerve terminals in packets or quanta of uniform size led to the speculation that the transmitter might be stored in synaptic vesicles (1,2). The isolation of acetylcholine-containing pinched-off nerve endings (synaptosomes) from brain (3) and more recently the synaptic vesicles themselves (4,5) have stimulated investigations of the subcellular localization of biogenic amines in peripheral and central nervous tissue. In guinea pig brain the storage particles that contain norepinephrine (NE) and 5-hydroxytryptamine have been found to differ from those containing acetylcholine in that the first two have sedimentation characteristics more dense than the acetylcholine containing vesicles (6). Recent studies have shown that catecholamines in heart (7,8), brain (6,9) and splenic nerve (10) are stored in subcellular vesicles. Potter and Axelrod (7,11) reported that 30 minutes after the intravenous administration of DL-norepinephrine-7-H³ to rats a sizable fraction of the labeled compound in several peripheral tissues appeared in a fraction associated with microsomes. These authors suggested that the radioactive NE is taken up from the circulation and fixed at the anatomical sites which bind endogenous NE. The morphological characterization of these sites was not possible with the Potter and Axelrod preparation. A modification of the density gradient centrifugation procedure of Potter and Axelrod (11) has made it possible to isolate NE containing particles of considerably greater homogeneity. Both endogenous NE and administered H³NE are localized in a particulate fraction which equilibrates in that part of the gradient which corresponds in density ot 0.45-0.5 M sucrose. Electronmicroscopic examination of this fraction has revealed numerous small vesicles approximately 500 Å in diameter, some microsomal fragments but essentially no other subcellular organelles.