For more than a century the diffraction barrier has prevented neuroscientists from resolving cellular structures smaller than around 200 nm by light microscopy. However, within this decade, several live cell compatible microscopy techniques have been developed that allow imaging beyond this limit. These techniques are closing the gap between existing electron microscopy and fluorescence microscopy modalities, by allowing researchers to study the nanoscale morphology of living cells. This provides an opportunity to study details of the smallest morphological entities of synapses in live tissue, including dendritic spines and axons, as well as processes from astrocytes and microglia.

Our recent work has focused on dendritic spines; minute bulbous protrusions of the cell membrane consisting of a spine head harboring the excitatory postsynapse, which is connected to the dendrite via a thin spine neck. The small sizes of the spine head and neck have hindered geometrical analyses by conventional 2-photon and confocal fluorescence microscopy, and the structure-function relationship of spines is therefore poorly understood.

To relate spine structure to function we combined superresolution STED imaging with fluorescence recovery after photo-bleaching (FRAP) measurements, glutamate uncaging, and whole-cell patch-clamp recordings in organotypic hippocampal slice cultures. This approach allowed us to resolve the detailed morphology of live spines on hippocampal pyramidal cells, and to relate morphological details directly to functional readouts. We investigated how functional plasticity is paralleled by morphological dynamics, and found conceptually new aspects of the structure-function relationship of spines.

In this seminar I will briefly introduce the most widely applied superresolution microscopy techniques (STED, SIM, STORM/PALM) and emphasize their respective strengths and weaknesses for neuroscience research, before presenting our experimental work on dendritic spines.