Scientific Understanding of Consciousness
Hippocampal Engram of Sparse Population for Specific Memory
Nature 484, 381–385 (19 April 2012)
Optogenetic stimulation of a hippocampal engram activates fear memory recall
Xu Liu, Steve Ramirez, Petti T. Pang, Corey B. Puryear, Arvind Govindarajan, Karl Deisseroth & Susumu Tonegawa
RIKEN-MIT Center for Neural Circuit Genetics at the Picower Institute for Learning and Memory, Department of Biology and Department of Brain and Cognitive Sciences, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139, USA
Department of Bioengineering and Department of Psychiatry and Behavioral Sciences, Stanford University, Stanford, California 94305, USA
A specific memory is thought to be encoded by a sparse population of neurons. These neurons can be tagged during learning for subsequent identification and manipulation. Moreover, their ablation or inactivation results in reduced memory expression, suggesting their necessity in mnemonic processes. However, the question of sufficiency remains: it is unclear whether it is possible to elicit the behavioural output of a specific memory by directly activating a population of neurons that was active during learning. Here we show in mice that optogenetic reactivation of hippocampal neurons activated during fear conditioning is sufficient to induce freezing behaviour. We labelled a population of hippocampal dentate gyrus neurons activated during fear learning with channelrhodopsin-2 (ChR2) and later optically reactivated these neurons in a different context. The mice showed increased freezing only upon light stimulation, indicating light-induced fear memory recall. This freezing was not detected in non-fear-conditioned mice expressing ChR2 in a similar proportion of cells, nor in fear-conditioned mice with cells labelled by enhanced yellow fluorescent protein instead of ChR2. Finally, activation of cells labelled in a context not associated with fear did not evoke freezing in mice that were previously fear conditioned in a different context, suggesting that light-induced fear memory recall is context specific. Together, our findings indicate that activating a sparse but specific ensemble of hippocampal neurons that contribute to a memory engram is sufficient for the recall of that memory. Moreover, our experimental approach offers a general method of mapping cellular populations bearing memory engrams.
An important question in neuroscience is how a distinct memory is formed and stored in the brain. Recent studies indicate that defined populations of neurons correspond to a specific memory trace, suggesting a cellular correlate of a memory engram. Selective ablation or inhibition of such neuronal populations erased the fear memory response, indicating that these cells are necessary for fear memory expression. However, to prove that a cell population is the cellular basis of a specific fear memory engram it is necessary to conduct a mimicry experiment to show that direct activation of such a population is sufficient for inducing the associated behavioural output.
The hippocampus is thought to be critical in the formation of the contextual component of fear memories. Modelling and experimental studies have demonstrated an essential role of the dentate gyrus (DG) of the hippocampus in discriminating between similar contexts. Cellular studies of immediate early gene expression showed that sparse populations of DG granule cells (2–4%) are activated in a given context. Moreover, although the same population of DG granule cells is activated repeatedly in the same environment, different environments or different tasks activate different populations of DG granule cells. These lines of evidence point to the DG as an ideal target for the formation of contextual memory engrams that represent discrete environments.
We have shown that optical activation of hippocampal cells that were active during FC elicits freezing behaviour. To our knowledge, this is the first demonstration that directly activating a subset of cells involved in the formation of a memory is sufficient to induce the behavioural expression of that memory. Our results and previous studies that addressed the necessity of similarly sparse cell populations in the amygdala argue that defined cell populations can form a cellular basis for fear memory engrams. The memory engram that we selectively labelled and manipulated is probably contextual in nature, as previous studies have demonstrated that hippocampal interventions affect conditioned freezing responses to a context but not a tone. Indeed, recent findings show that optogenetic inhibition of the hippocampal CA1 region during training or testing inhibited the recall of a contextual fear memory, while leaving auditory-cued fear memory recall intact. However, we cannot completely rule out the possibility that the fear memory recalled in our experiments may have some tone memory component.
Although we have demonstrated the sufficiency of a DG memory engram for the behavioural expression of a fear memory, we cannot conclude that this engram is necessary for behavioural recall. During contextual fear conditioning (FC), it is likely that multiple contextual memory engrams are formed in a series of hippocampal regions. Each of these engrams may contribute to the formation of the complete fear memory in the basolateral amygdala (BLA) and may also be capable of reactivating it independently, as we observed in the case of the DG engrams. Because the hippocampus is not a linear feed-forward network but contains several parallel circuits, inhibiting the formation or activation of contextual engrams in one region may not necessarily block the expression of the fear memory. For instance, disruption of contextual memory engrams in the DG could be circumvented by CA1 engrams, which could be generated through the direct input from the entorhinal cortex and may be sufficient to activate the fear memory engram in the BLA. Indeed, we recently generated a mouse mutant, which permitted us to demonstrate that the DG input to the CA3 is dispensable in the formation and retrieval of contextual fear memory.
The approach and methods described in this work will be a powerful tool for mapping multiple component engrams, each contributing to an overall memory. A multifaceted analysis of these engrams and their interplay will reveal the nature of the overall memory engram.
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