Visualization of synaptic activity in hippocampal slices with FM1-43 enabled by fluorescence quenching

Fluorescence imaging of presynaptic uptake and release of styryl dyes such as FM1-43 has provided valuable insights into synaptic function. However, i n studies of CNS neurons, the utility of these dyes has been severely limit ed by nonsynaptic background fluorescence. This has thwarted the use of FM dyes in systems more intact than dissociated neuronal cultures. Here, we de scribe an approach to selectively reduce undesired fluorescence through que nching of the surface-bound FM1-43 signal. The introduction of sulfo-rhodam ine, a fluorophore that is not taken up by synaptic vesicles, selectively r educed the nonsynaptic fluorescence in FM1-43-labeled hippocampal cultures. When applied to rat hippocampal slices, this procedure allowed us to obser ve activity-dependent staining and destaining of functional synapses. Exten ding the usefulness of styryl dyes to slice preparations may help make func tional synaptic networks amenable to optical measurements.