Figure 2.

Mapping connectivity utilizing photostimulation: (A) Presynaptic neurons (black) are stimulated sequentially in the slice-preparation to fire action potentials by focused uncaging of glutamate via short pulses of UV-light. These APs, in turn, evoke synaptic potentials in the postsynaptic neuron (red), the membrane potential of which is monitored via a patch-clamp electrode (for detailed methods, see Boucsein et al., 2005). (B) Overlay of a picture of a neocortical brain slice and the results from scanning the tissue for functional connections to the postsynaptic neuron (reconstruction shown in red). Squares denote the positions of presynaptic sites, the stimulation of which elicited a postsynaptic current. Colored squares refer to selected sites, highlighted in the corresponding color in (C,D). (C) Principle of dynamic photostimulation (DPS). Using the fast DPS-system (Boucsein et al., 2005; Nawrot et al., 2009), presynaptic neurons can be activated dynamically to produce patterns of spatiotemporal synaptic input to the postsynaptic cell (red). (D) Dendritic integration in the postsynaptic neuron. Three repetitions of the same stimulation sequence (tics, bottom) resulted in repeated sub-threshold membrane potential fluctuations. Voltage traces for selected presynaptic sites are colored for the first 80 ms following presynaptic photostimulation, corresponding to the sites, indicated in (B,C). Note the remarkable reproducibility of the voltage fluctuations across trials. The gray trace shows the response to the same stimulation after application of TTX, confirming the purely synaptic origin of the recorded potentials.