Analysis of Multiquantal Transmitter Release From Single Cultured Cortical Neuron Terminals

Abstract

Analysis of multiquantal transmitter release from single cultured cortical neuron terminals. Application of single synapse recording methods indicates that the amplitude of postsynaptic responses of single CNS synapses can vary greatly among repeated stimuli. To determine whether this observation could be attributed to synapses releasing a variable number of transmitter quanta, we assessed the prevalence of multiquantal transmitter release in primary cultures of cortical neurons with the action potential (AP)-dependent presynaptic turnover of the styryl dye FM1–43 ( Betz and Bewick 1992 , 1993 ; Betz et al. 1996 ). It was assumed that if a high proportion of vesicles within a terminal were loaded with FM1–43 the amount of dye released per stimulus would be proportional to the number of quanta released and/or the probability of release at a terminal. To rule out differences in the amount of release (between terminals) caused by release probability or incomplete loading of terminals, conditions were chosen to maximize both release probability and terminal loading. Three-dimensional reconstruction of terminals was employed to ensure that bouton fluorescence was accurately measured. Analysis of the relationship between the loading of terminals and release indicated that presumed larger terminals (>FM1–43 uptake) release a greater amount of dye per stimulus than smaller terminals, suggesting multiquantal release. The distribution of release amounts across terminals was significantly skewed toward higher values, with 13–17% of synaptic terminals apparently releasing multiple quanta per AP. In conclusion, our data suggest that most synaptic terminals release a relatively constant amount of transmitter per stimulus; however, a subset of terminals releases amounts of FM1–43 that are greater than that expected from a unimodal release process.