TURNOVER OF TRANSMITTER AND SYNAPTIC VESICLES AT THE FROG NEUROMUSCULAR JUNCTION

TLDR

Frog neuromuscular preparations were stimulated at 10 Hz or 2 Hz, end‑plate potentials recorded, and terminals fixed or treated with venom to quantify remaining transmitter, while horseradish peroxidase or dextran tracers were used to detect axolemmal vesicle formation. Stimulation produced extensive vesicle turnover: after 4 h at 2 Hz many new vesicles formed and transmitter secretion exceeded the initial store, while after 20 min at 10 Hz terminals swelled and were depleted of both vesicles and transmitter, yet a 1‑h rest restored morphology, generated new vesicles, and partially replenished transmitter stores, indicating vesicles fuse with and reform from the terminal membrane and can store and release transmitter.

Abstract

Curarized cutaneous pectoris nerve-muscle preparations from frogs were stimulated at 10/s or at 2/s for periods ranging from 20 min to 4 h. End plate potential were recorded intracellularly and used to estimate the quantity of transmitter secreted during the period of stimulation. At the ends of the periods of stimulation the preparations were either fixed for electron microscopy or treated with black widow spider venom to determine the quantities of transmitter remainind in the terminal. Horseradish peroxidase or dextran was added to the bathing solution and used as a tracer to detect the formation of vesicles from the axolemma. During 4 h of stimulation at 2/s many new vesicles were formed from the axolemma and the quantity of transmitter secreted was several times greater than the quantity in the initial store. After this period of stimulation, the terminals were severely depleted of transmitter, but not of vesicles, and their general morphological organization was normal. During 20 min of stimulation at 10/s the nerve terminals swelled and were severely depleted both of vesicles and of transmitter. During a subsequent hour of rest the changes in morphology were largely reversed, many new vesicles were formed from the axolemma and the stores of transmitter were partially replenished. These results suggest (a) that synaptic vesicles fuse with, and re-form from, the membrane of the nerve terminal during and after stimulation and (b), that the re-formed vesicles can store and release transmitter.