Cholinergic synaptic vesicles are metabolically and biophysically heterogeneous even in resting terminals

Abstract

The metabolic heterogeneity of synaptic vesicles in the cholinergic nerve terminals of the electromotor neurons of Torpedp marmoratta has been studied in resting tissue by evaluating the molecular acetylcholine content (MAC) of synaptic vesicles after vesicles extraction from frozen and crushed tissue and high-resolution centrifugal density gradient separation in a zonal rotor. Although vesicular acetylcholine was distributed in the gradient as a single, more or less symmetrical peak, 3 subpopulations of synaptic vesicles could be identified: a small, relatively light subpopulation of low MAC on the ascending limb of the acetylcholine peak, designated V 0, a main population of fully charged vesicles designated V 1, and a small, denser subpopulation also of low MAC on the descending limb of the acetylcholine peak, designated V 2. The mean proportions and MACs of the 3 pools were: V 0, 13%, 58,00; V 1, 53%, 246,000; V 2, 34%, 79,000. When triated acetate was perfused through excised blocks of electric organ for 1–2 h before vesicle isolation, the specific radioactivity of thr acetylcholine in the V 0 and V 2 pools was 10–30 times higher than in the V 1 pool. This suggest that both the V 0 and V 2 pools are not generated by the isolation procedure but are present in the intact endings and are functionally active. On the basis of their density and uptake of newly synthesized acetylcholine, the V 0 and V 2 pools were identified with the previously described VP 0 pool of axonal vesicles and the VP 2 pool of recycling vesicles in stimulated nerve terminals respectively. Since stimulation of electromotor nerve terminals is known to generate large proportions VP 2 vesicles, variations in the proportion of V 2 vesicles in unstimulated tissue are attributed to varying amounts of adventitous stimulation of the tissue during dissection and perfussion.