Artificial association of memory events by optogenetic stimulation of hippocampal CA3 cell ensembles.

Previous gain-of-function studies using an optogenetic technique showed that manipulation of the hippocampal dentate gyrus or CA1 cell ensembles is important for memory reactivation and to generate synthetic or false memory. However, gain-of-function study manipulating CA3 cell ensembles has not been reported. The CA3 area of the hippocampus comprises a recurrent excitatory circuit, which is thought to be important for the generation of associations among the stored information within one brain region. We investigated whether the coincident firing of cell ensembles in one brain region, hippocampal CA3, associates distinct events. CA3 cell ensembles responding to context exploration and during contextual fear conditioning were labeled with channelrhodopsin-2 (ChR2)-mCherry. The synchronous activation of these ensembles induced freezing behavior in mice in a neutral context, in which a foot shock had never been delivered. The recall of this artificial associative fear memory was context specific. In vivo electrophysiological recordings showed that 20-Hz optical stimulation of ChR2-mCherry-expressing CA3 neurons, which is the same stimulation protocol used in behavioral experiment, induced long-term potentiation at CA3-CA3 synapses. Altogether, these results demonstrate that the synchronous activation of ensembles in one brain region, CA3 of the hippocampus, is sufficient for the association of distinct events. The results of our electrophysiology potentially suggest that this artificial association of memory events might be induced by the strengthening of synaptic efficacy between CA3 ensembles via recurrent circuit.

Overlapping memory trace indispensable for linking, but not recalling, individual memories.

Memories are not stored in isolation from other memories but are integrated into associative networks. However, the mechanisms underlying memory association remain elusive. Using two amygdala-dependent behavioral paradigms-conditioned taste aversion (CTA) and auditory-cued fear conditioning (AFC)-in mice, we found that presenting the conditioned stimulus used for the CTA task triggered the conditioned response of the AFC task after natural coreactivation of the memories. This was accompanied through an increase in the overlapping neuronal ensemble in the basolateral amygdala. Silencing of the overlapping ensemble suppressed CTA retrieval-induced freezing. However, retrieval of the original CTA or AFC memory was not affected. A small population of coshared neurons thus mediates the link between memories. They are not necessary for recalling individual memories.

Cellular tagging as a neural network mechanism for behavioural tagging.

Behavioural tagging is the transformation of a short-term memory, induced by a weak experience, into a long-term memory (LTM) due to the temporal association with a novel experience. The mechanism by which neuronal ensembles, each carrying a memory engram of one of the experiences, interact to achieve behavioural tagging is unknown. Here we show that retrieval of a LTM formed by behavioural tagging of a weak experience depends on the degree of overlap with the neuronal ensemble corresponding to a novel experience. The numbers of neurons activated by weak training in a novel object recognition (NOR) task and by a novel context exploration (NCE) task, denoted as overlapping neurons, increases in the hippocampal CA1 when behavioural tagging is successfully achieved. Optical silencing of an NCE-related ensemble suppresses NOR-LTM retrieval. Thus, a population of cells recruited by NOR is tagged and then preferentially incorporated into the memory trace for NCE to achieve behavioural tagging.

Artificial association of pre-stored information to generate a qualitatively new memory.

Memory is thought to be stored in the brain as an ensemble of cells activated during learning. Although optical stimulation of a cell ensemble triggers the retrieval of the corresponding memory, it is unclear how the association of information occurs at the cell ensemble level. Using optogenetic stimulation without any sensory input in mice, we found that an artificial association between stored, non-related contextual, and fear information was generated through the synchronous activation of distinct cell ensembles corresponding to the stored information. This artificial association shared characteristics with physiologically associated memories, such as N-methyl-D-aspartate receptor activity and protein synthesis dependence. These findings suggest that the association of information is achieved through the synchronous activity of distinct cell ensembles. This mechanism may underlie memory updating by incorporating novel information into pre-existing networks to form qualitatively new memories.