Function and EM
Using Femtonics microscopes and careful procedures it is possible to correlate functional properties measured by calcium imaging with the ultrastructure obtained by electron microscopy.
Most functional studies suffer from the lack of precise anatomical characterization of the cell or tissue. The high spatial resolution of Femtonics microscopes enables easier identification of the functionally characterized structure in electron microscopy in order to correlate anatomical structure with functional properties.
An elegant example has been shown by Holderith and colleges: Authors included biocytin in the intracellular solution in addition to the fluorescent dyes allowing post hoc light microscopic and subsequent electron microscopic visualization of the imaged areas. Their results showed clear correlation between the functional and the ultra-structural properties.
Left, up) Ca2+ transients measured with line scans in a representative spine head of a CA3 pyramidal cell basal dendrite. Two responses were evoked by extracellular stimulation (100 ms inter-stimulus interval, the time of the stimuli is indicated by the arrowheads) of local axon collaterals in the str. oriens. Successful transmission for the first and second stimuli are colored in blue and green, respectively (failures are shown in black). Right, up) Electron microscopic images of an axonal bouton that established excitatory synapses on pyramidal cell spines (arrowheads demarcate the post synaptic density) Bottom) Total fluxed calcium, calculated from the peak Ca2+ transients and the volume of the boutons, correlates tightly (p < 0.001) with the active zone area. Green circle represent the bouton shown in upper panel
Image courtesy of Noemi Holderith & Zoltan Nusser, IEM HAS, Budapest, Hungary
Related scientific publication:
Holderith et al, Nature Neuroscience (2012)