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. Author manuscript; available in PMC: 2017 Feb 8.
Published in final edited form as: Cold Spring Harb Protoc. 2016 Jun 1;2016(6):pdb.prot091306. doi: 10.1101/pdb.prot091306

FIGURE 2.

FIGURE 2

Finding connected neurons using quadruple recordings. (A) Flattened two-photon-imaging stack of four neighboring L5 PCs filled with Alexa 594 (scale bar: 25 μm). (B) Thirty Hertz trains are evoked (top, scale bars: 200 msec, 10 mV) to identify responding postsynaptic cells (bottom, scale bars: 100 msec, 0.25 mV). Sweeps should be repeated 10–40 times every 10–20 sec, and are then averaged. Note short-term depressing connection from cell 3 (black) to cell 2 (gray). (C) Monosynaptic connections have a jitter of less than 1 msec (blue arrows); larger jitter suggests that the responses are polysynaptic. Fifty spike-triggered traces (gray) from cell 2 are represented, whereas the presynaptic action potential (black) is represented by a single sweep (scale bars: 2 msec, 20 mV [black]/300 μV [gray]). (D) Monosynaptic connections also have submillisecond latency between presynaptic spike and 10% of EPSP peak (vertical dashed lines). EPSP trace (gray) is a spike-triggered average of 50 sweeps, whereas the presynaptic action potential (black) is represented by a single sweep (scale bars: 1 msec, 20 mV [black]/200 μV [gray]).