Activity-induced changes in synaptic release sites at the crayfish neuromuscular junction

Abstract

Crustacean motor axons provide a model in which activity-dependent changes in synaptic physiology and synaptic structure can be concurrently observed in single identifiable neurons. In response to a train of stimulation, crustacean neuromuscular junctions undergo pronounced facilitation of transmitter release. The effects of maintained high-frequency stimulation may persist for at least several hours (“long-term facilitation”). Electrophysiological studies suggest that the number of “active” synapses contributing transmitter quanta at low frequencies of stimulation increases during and after a train of high-frequency stimulation. However, at different terminal recording sites the effect of stimulation varies, and it was observed that not all released quanta produce a voltage change in the postsynaptic muscle cell. Electron microscopic examinations of serial sections from nerve terminals subjected to stimulation were made to determine whether changes in synaptic structure could be correlated with activity-induced long-lasting enhancement of transmission. A procedure was introduced for marking a recorded terminal with fluorescent polystyrene microspheres, which are visible in electron micrographs of the recording site. Crustacean nerve terminals possess a large number of discrete synapses, a small fraction of which have multiple presynaptic “active zones” (dense bodies with clustered synaptic vesicles, thought to represent sites of evoked transmitter release). In terminals previously stimulated, the proportion of synapses with multiple “active zones” is greater than in control unstimulated terminals. Total synaptic vesicle counts and readily releasable vesicles at synapses are not significantly different in previously stimulated terminals and controls. In terminals fixed during stimulation a few synapses show evidence of division in “active zones,” and synaptic vesicle counts are lower than in controls.(ABSTRACT TRUNCATED AT 250 WORDS)

Assessing ultrastructure of crustacean and insect neuromuscular junctions

“…Recent studies using the combined approach have enhanced the view that many terminals possess silent synapses, and that the number of active synapses can be adjusted during short-term and long-term facilitation to Ž . meet physiological requirements Wojtowicz et al, 1994 . In addition, there is reason to believe that the number of synapses on a terminal is only one of the factors responsible for synaptic efficacy in both Drosophila and crus-Ž taceans Kurdyak et al, 1994;. Cooper et al, 1995c . As shown here, such studies may be subject to errors affecting the estimates of the sizes of small structures, but the general features of synaptic proliferation and enlargement are not qualitatively altered by these considerations.…”

“…These data can be used in a three-dimensional reaction-diffusion model of calcium ions entering at Ž . active zones Winslow et al, 1994b to determine whether it is likely that the calcium domains from closely associated active zones interact during nerve terminal depolariza-Ž tion Wojtowicz et al, 1994;Cooper . et al, , 1996Winslow et al, 1994a .…”

“…The beads remain on the surface of the tissue as it is processed for electron microscopy, and they are visible within the resin block under fluorescence Ž . microscopy Wojtowicz et al, 1994;Cooper et al, 1995c . The recording site can then be rapidly located by sectioning to the beads, and locating them in thick sections once the site is being approached. Serial thin sections are then cut and checked for the beads, which are readily apparent in transmission electron microscopy.…”

“…quantal content increases frequency facilitation , indicating recruitment of some silent synapses to the responsive Ž . pool Wojtowicz et al, 1991Wojtowicz et al, , 1994 . Probably this situation exists in many neuronal systems.…”

“…This approach has been used successfully in studies of crus-Ž tacean nerve terminals Wojtowicz et al, 1994;Cooper et . al., 1995c , and could be applied to advantage elsewhere.…”