Rapid activation of plasticity-associated gene transcription in hippocampal neurons provides a mechanism for encoding of one-trial experience

Abstract

The hippocampus is hypothesized to support rapid encoding of ongoing experience. A critical prerequisite for such function is the ability to readily recruit enduring synaptic plasticity in hippocampal neurons. Hippocampal long-term potentiation (LTP) and memory consolidation require expression of the immediate-early gene (IEG) Arc. To determine whether Arc transcription could be driven by limited and controlled behavioral experience, we used a rectangular track paradigm. In past electrophysiological studies, pyramidal neurons recorded from rats running in one direction on similar tracks typically exhibited a single firing field. Using fluorescence in situ hybridization, we show that the behavioral activity associated with a single lap around the track was sufficient to trigger Arc transcription in complete CA3 neuronal ensembles, as predicted given the role of CA3 in one-trial learning. In contrast, Arc transcription in CA1 ensembles was recruited incrementally, with maximal activation achieved after four laps a day for 4 consecutive days. To test whether Arc transcription is linked to learning and plasticity, or merely elicited by location-specific firing, we inactivated the medial septum, a treatment that compromises hippocampus-dependent learning and LTP but spares location-specific firing in CA1 neurons. Septal inactivation abolished track training-induced Arc transcription in CA1 and CA3 neurons, showing that Arc transcription requires plasticity-inducing stimuli. Accordingly, LTP induction activated Arc transcription in CA1 neurons in vivo. These findings demonstrate for the first time that a single brief experience, equivalent to a single crossing of a firing field, can trigger IEG expression required for long-term plasticity in the hippocampus.

Figure 1.

Track training design. a, A schematic and photograph of the track training procedure. At each training session, a rat was placed on the starting corner of the track and guided to walk in the same direction. b, A schematic of the experimental design. Rats walked around the track in one direction either once or four times on a single day or over 4 consecutive days. sac, The time point at which rats were killed.

Figure 2.

Activation of Arc, zif268, and c-fos transcription in dorsal hippocampus by limited experience on the track revealed by RT-qPCR. For all genes analyzed, training on the track induced significantly higher levels of primary transcript relative to the caged controls (*p < 0.05). Of note, the fold differences between the track-trained groups did not differ for any of the genes. n = 3, except for 4L-1D (n = 2).

Figure 3.

Dynamics of hippocampal ensemble recruitment in the behaving rat: robust activation of CA3 by limited experience and incremental activation of CA1 with extended experience. Quantification of Arc-positive CA3 and CA1 neurons from rats trained on the rectangular track. a, A confocal projection image of the CA3 region from a TC, 1L-1D, and 4L-4D rat. Yellow arrows indicate cells containing Arc transcription foci (green color) within a nucleus (red color). Scale bars, 20 μm. b, A confocal projection image of the CA1 region from a TC, 1L-1D, and 4L-4D rat. c, The percentages of Arc⁺ CA3 neurons (mean ± SEM) for the six groups. Arc⁺ neurons of the track-trained rats were significantly higher than those of both control groups but did not differ from each other. *p < 0.05, different from all of the track-trained groups; n = 5–9 rats per group. d, The percentages of Arc⁺ CA1 neurons (mean ± SEM) for the six groups. Similar to CA3, the proportions of Arc⁺ neurons of the track-trained animals were significantly higher than those of controls. However, in contrast to CA3, the proportion of Arc⁺ neurons incremented with increased experience on the track. *p < 0.05, different from all track-trained groups; ^Δp < 0.05, different from 1L-1D; n = 5–9 rats per group. e, The proportion of Arc⁺ cells in CA3 did not correlate with number of laps completed on the track. Pearson's correlation coefficient and p values are shown. f, The proportion of Arc⁺ cells in CA1 correlated with number of laps completed on the track. Pearson's correlation coefficient and p values are shown.

Figure 4.

Activation of Arc transcription in single CA3 and CA1 neurons. a, b, The comparison of integrated fluorescent intensities (median and interquartile range) for Arc⁺ and Arc⁻ cells from the track trained groups in CA3 (a) and CA1 (b). Note the virtual lack of signal in Arc⁻ cells and the dramatic levels in Arc⁺ cells. Whereas the intensities of Arc⁺ signals did not differ across the training groups in CA3, Arc⁺ signal in 1L-1D group was significantly lower than the other three training groups in CA1 (*p < 0.0001 compared with all other three groups; ^Δp < 0.05 compared with 4L-1D and 1L-4D).

Figure 5.

Medial septal inactivation by infusion of tetracaine abolished behavioral induction of Arc in CA3 and CA1 neurons. a, A schematic of the experimental design indicating the time course of training, infusion, and death. sac, The time point at which rats were killed. b, c, Confocal projection images from CA3 (b) and CA1 (c) from a VEH-TRACK and TETRA-TRACK rat. Yellow arrows indicate cells containing Arc transcription foci (green color) within a cell nucleus (red color). Scale bars, 20 μm. d, e, In both CA3 (d) and CA1 (e), the percentage of Arc⁺ neurons (mean ± SEM) in the VEH-TRACK group was significantly higher than in all three other groups. *p < 0.0001, different from all other behavioral groups in a; *p < 0.001 in b; n = 3–6 rats per group.

Figure 6.

Activation of IEG transcription in CA1 neurons by LTP-inducing stimulation. a, b, Traces illustrating negative-going population EPSPs recorded by a micropipette positioned in CA1 contralateral from the stimulating electrode in CA3 at baseline (a) and after HFS (b; 2 100-Hz trains separated by 30 s). c, Population-spike amplitude as a percentage of baseline before and after HFS stimulation. d, Confocal projection images of the contralateral CA1 show that LTP stimulation (HFS) activated transcription of Arc and zif268, whereas the test stimulus did not. Similar results were seen in ipsilateral CA1 neurons (data not shown). Yellow arrows indicate cells containing Arc transcription foci (green color) within a nucleus (red color).