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. Author manuscript; available in PMC: 2017 Sep 5.
Published in final edited form as: Science. 2015 May 28;348(6238):1007–1013. doi: 10.1126/science.aaa5542

Figure 1. Synaptic Plasticity and Connectivity of Engram Cells.

Figure 1

(A) Mice taken Off DOX 24 hrs before contextual fear conditioning (CFC) and dispatched 24 hrs post training. Saline (SAL) or anisomycin (ANI) administered immediately after training.

(B) AAV8-CaMKIIα-ChR2-EYFP and AAV9-TRE-mCherry viruses injected into the entorhinal cortex and dentate gyrus, respectively, of c-fos-tTA mice.

(C) Paired recordings of engram (red) and non-engram (grey) DG cells during optogenetic stimulation of ChR2+ perforant path (PP) axons.

(D) Representative image of a pair of recorded biocytin-labeled engram (mCherry+) and non-engram (mCherry−) DG cells. Note ChR2+ PP axons in green.

(E) (Top) Example traces of AMPA and NMDA receptor-dependent postsynaptic currents in mCherry+ and mCherry− cells, evoked by light activation of ChR2+ PP axons. (Bottom) EPSC amplitudes and AMPA/NMDA current ratios of mCherry+ and mCherry− cells of the two groups are displayed as means (columns) and individual paired data points (grey lines). Paired t-test * p < 0.05, ** p < 0.001. SAL group compared with the ANI group, unpaired t-test * p < 0.05.

(F) (Left) Representative confocal images of biocytin filled dendritic fragments derived from SAL and ANI groups for ChR2+ and ChR2− cells (arrow heads: dendritic spines). (Right) Average dendritic spine density showing an increase occurring exclusively in ChR2+ fragments. Data are represented as mean ± SEM. Unpaired t tests ** p < 0.01, *** p < 0.001.

(G) Engram Connectivity. (Top left) AAV9-TRE-ChR2-EYFP and AAV9-TRE-mCherry viruses, injected into the DG and CA3, respectively, of c-fos-tTA mice. (Bottom left) Example of mCherry+ (1) and mCherry− (2) biocytin-filled CA3 pyramidal cells. Note ChR2+ mossy fibers (MF) in green. (Top Right) mCherry+ cell but not mCherry− cell displayed EPSPs in response to optogenetic stimulation of MF. (Bottom Right) Probability of connection of DG ChR2+ engram axons and CA3 mCherry+ and mCherry− cells. Error bars are approximated by binomial distribution. Fisher’s exact test: * p < 0.05.