Hippocampus and amygdala fear memory engrams re-emerge after contextual fear relapse.

The formation and extinction of fear memories represent two forms of learning that each engage the hippocampus and amygdala. How cell populations in these areas contribute to fear relapse, however, remains unclear. Here, we demonstrate that, in male mice, cells active during fear conditioning in the dentate gyrus of hippocampus exhibit decreased activity during extinction and are re-engaged after contextual fear relapse. In vivo calcium imaging reveals that relapse drives population dynamics in the basolateral amygdala to revert to a network state similar to the state present during fear conditioning. Finally, we find that optogenetic inactivation of neuronal ensembles active during fear conditioning in either the hippocampus or amygdala is sufficient to disrupt fear expression after relapse, while optogenetic stimulation of these same ensembles after extinction is insufficient to artificially mimic fear relapse. These results suggest that fear relapse triggers a partial re-emergence of the original fear memory representation, providing new insight into the neural substrates of fear relapse.

Minian, an open-source miniscope analysis pipeline.

Miniature microscopes have gained considerable traction for in vivo calcium imaging in freely behaving animals. However, extracting calcium signals from raw videos is a computationally complex problem and remains a bottleneck for many researchers utilizing single-photon in vivo calcium imaging. Despite the existence of many powerful analysis packages designed to detect and extract calcium dynamics, most have either key parameters that are hard-coded or insufficient step-by-step guidance and validations to help the users choose the best parameters. This makes it difficult to know whether the output is reliable and meets the assumptions necessary for proper analysis. Moreover, large memory demand is often a constraint for setting up these pipelines since it limits the choice of hardware to specialized computers. Given these difficulties, there is a need for a low memory demand, user-friendly tool offering interactive visualizations of how altering parameters at each step of the analysis affects data output. Our open-source analysis pipeline, Minian (miniscope analysis), facilitates the transparency and accessibility of single-photon calcium imaging analysis, permitting users with little computational experience to extract the location of cells and their corresponding calcium traces and deconvolved neural activities. Minian contains interactive visualization tools for every step of the analysis, as well as detailed documentation and tips on parameter exploration. Furthermore, Minian has relatively small memory demands and can be run on a laptop, making it available to labs that do not have access to specialized computational hardware. Minian has been validated to reliably and robustly extract calcium events across different brain regions and from different cell types. In practice, Minian provides an open-source calcium imaging analysis pipeline with user-friendly interactive visualizations to explore parameters and validate results.

Consistent population activity on the scale of minutes in the mouse hippocampus.

Neurons in the hippocampus fire in consistent sequence over the timescale of seconds during the delay period of some memory experiments. For longer timescales, the firing of hippocampal neurons also changes slowly over minutes within experimental sessions. It was thought that these slow dynamics are caused by stochastic drift or a continuous change in the representation of the episode, rather than consistent sequences unfolding over minutes. This paper studies the consistency of contextual drift in three chronic calcium imaging recordings from the hippocampus CA1 region in mice. Computational measures of consistency show reliable sequences within experimental trials at the scale of seconds as one would expect from time cells or place cells during the trial, as well as across experimental trials on the scale of minutes within a recording session. Consistent sequences in the hippocampus are observed over a wide range of time scales, from seconds to minutes. The hippocampal activity could reflect a scale-invariant spatiotemporal context as suggested by theories of memory from cognitive psychology.

Dynamic and heterogeneous neural ensembles contribute to a memory engram.

In the century since the notion of the 'engram' was first introduced to describe the physical manifestation of memory, new technologies for identifying cellular activity have enabled us to deepen our understanding of the possible physical substrate of memory. A number of studies have shown that memories are stored in a sparse population of neurons known as a neural ensemble or engram cells. While earlier investigations highlighted that the stability of neural ensembles underlies a memory representation, recent studies have found that neural ensembles are more dynamic and fluid than previously understood. Additionally, a number of studies have begun to dissect the cellular and molecular diversity of functionally distinct subpopulations of cells contained within an engram. We propose that ensemble fluidity and compositional heterogeneity support memory flexibility and functional diversity.

Hippocampal spatial memory representations in mice are heterogeneously stable.

The population of hippocampal neurons actively coding space continually changes across days as mice repeatedly perform tasks. Many hippocampal place cells become inactive while other previously silent neurons become active, challenging the idea that stable behaviors and memory representations are supported by stable patterns of neural activity. Active cell replacement may disambiguate unique episodes that contain overlapping memory cues, and could contribute to reorganization of memory representations. How active cell replacement affects the evolution of representations of different behaviors within a single task is unknown. We trained mice to perform a delayed nonmatching to place task over multiple weeks, and performed calcium imaging in area CA1 of the dorsal hippocampus using head-mounted miniature microscopes. Cells active on the central stem of the maze "split" their calcium activity according to the animal's upcoming turn direction (left or right), the current task phase (study or test), or both task dimensions, even while spatial cues remained unchanged. We found that, among reliably active cells, different splitter neuron populations were replaced at unequal rates, resulting in an increasing number of cells modulated by turn direction and a decreasing number of cells with combined modulation by both turn direction and task phase. Despite continual reorganization, the ensemble code stably segregated these task dimensions. These results show that hippocampal memories can heterogeneously reorganize even while behavior is unchanging.

Neurophysiological coding of space and time in the hippocampus, entorhinal cortex, and retrosplenial cortex.

Neurophysiological recordings in behaving rodents demonstrate neuronal response properties that may code space and time for episodic memory and goal-directed behaviour. Here, we review recordings from hippocampus, entorhinal cortex, and retrosplenial cortex to address the problem of how neurons encode multiple overlapping spatiotemporal trajectories and disambiguate these for accurate memory-guided behaviour. The solution could involve neurons in the entorhinal cortex and hippocampus that show mixed selectivity, coding both time and location. Some grid cells and place cells that code space also respond selectively as time cells, allowing differentiation of time intervals when a rat runs in the same location during a delay period. Cells in these regions also develop new representations that differentially code the context of prior or future behaviour allowing disambiguation of overlapping trajectories. Spiking activity is also modulated by running speed and head direction, supporting the coding of episodic memory not as a series of snapshots but as a trajectory that can also be distinguished on the basis of speed and direction. Recent data also address the mechanisms by which sensory input could distinguish different spatial locations. Changes in firing rate reflect running speed on long but not short time intervals, and few cells code movement direction, arguing against path integration for coding location. Instead, new evidence for neural coding of environmental boundaries in egocentric coordinates fits with a modelling framework in which egocentric coding of barriers combined with head direction generates distinct allocentric coding of location. The egocentric input can be used both for coding the location of spatiotemporal trajectories and for retrieving specific viewpoints of the environment. Overall, these different patterns of neural activity can be used for encoding and disambiguation of prior episodic spatiotemporal trajectories or for planning of future goal-directed spatiotemporal trajectories.

The brain in motion: How ensemble fluidity drives memory-updating and flexibility.

While memories are often thought of as flashbacks to a previous experience, they do not simply conserve veridical representations of the past but must continually integrate new information to ensure survival in dynamic environments. Therefore, 'drift' in neural firing patterns, typically construed as disruptive 'instability' or an undesirable consequence of noise, may actually be useful for updating memories. In our view, continual modifications in memory representations reconcile classical theories of stable memory traces with neural drift. Here we review how memory representations are updated through dynamic recruitment of neuronal ensembles on the basis of excitability and functional connectivity at the time of learning. Overall, we emphasize the importance of considering memories not as static entities, but instead as flexible network states that reactivate and evolve across time and experience.

The role of intrinsic excitability in the evolution of memory: Significance in memory allocation, consolidation, and updating.

Memory is a dynamic process that is continuously regulated by both synaptic and intrinsic neural mechanisms. While numerous studies have shown that synaptic plasticity is important in various types and phases of learning and memory, neuronal intrinsic excitability has received relatively less attention, especially regarding the dynamic nature of memory. In this review, we present evidence demonstrating the importance of intrinsic excitability in memory allocation, consolidation, and updating. We also consider the intricate interaction between intrinsic excitability and synaptic plasticity in shaping memory, supporting both memory stability and flexibility.

Trajectory-modulated hippocampal neurons persist throughout memory-guided navigation.

Trajectory-dependent splitter neurons in the hippocampus encode information about a rodent's prior trajectory during performance of a continuous alternation task. As such, they provide valuable information for supporting memory-guided behavior. Here, we employed single-photon calcium imaging in freely moving mice to investigate the emergence and fate of trajectory-dependent activity through learning and mastery of a continuous spatial alternation task. In agreement with others, the quality of trajectory-dependent information in hippocampal neurons correlated with task performance. We thus hypothesized that, due to their utility, splitter neurons would exhibit heightened stability. We find that splitter neurons were more likely to remain active and retained more consistent spatial information across multiple days than other neurons. Furthermore, we find that both splitter neurons and place cells emerged rapidly and maintained stable trajectory-dependent/spatial activity thereafter. Our results suggest that neurons with useful functional coding exhibit heightened stability to support memory guided behavior.

Retrosplenial Cortical Representations of Space and Future Goal Locations Develop with Learning.

Recent findings suggest that long-term spatial and contextual memories depend on the retrosplenial cortex (RSC) [1-5]. RSC damage impairs navigation in humans and rodents [6-8], and the RSC is closely interconnected with brain regions known to play a role in navigation, including the hippocampus and anterior thalamus [9, 10]. Navigation-related neural activity is seen in humans [11] and rodents, including spatially localized firing [12, 13], directional firing [12, 14, 15], and responses to navigational cues [16]. RSC neuronal activity is modulated by allocentric, egocentric, and route-centered spatial reference frames [17, 18], consistent with an RSC role in integrating different kinds of navigational information [19]. However, the relationship between RSC firing patterns and spatial memory remains largely unexplored, as previous physiology studies have not employed behavioral tasks with a clear memory demand. To address this, we trained rats on a continuous T-maze alternation task and examined RSC firing patterns throughout learning. We found that the RSC developed a distributed population-level representation of the rat's spatial location and current trajectory to the goal as the rats learned. After the rats reached peak performance, RSC firing patterns began to represent the upcoming goal location as the rats approached the choice point. These neural simulations of the goal emerged at the same time that lesions impaired alternation performance, suggesting that the RSC gradually acquired task representations that contribute to navigational decision-making.

Hippocampal Place Fields Maintain a Coherent and Flexible Map across Long Timescales.

To provide a substrate for remembering where in space events have occurred, place cells must reliably encode the same positions across long timescales. However, in many cases, place cells exhibit instability by randomly reorganizing their place fields between experiences, challenging this premise. Recent evidence suggests that, in some cases, instability could also arise from coherent rotations of place fields, as well as from random reorganization. To investigate this possibility, we performed in vivo calcium imaging in dorsal hippocampal region CA1 of freely moving mice while they explored two arenas with different geometry and visual cues across 8 days. The two arenas were rotated randomly between sessions and then connected, allowing us to probe how cue rotations, the integration of new information about the environment, and the passage of time concurrently influenced the spatial coherence of place fields. We found that spatially coherent rotations of place-field maps in the same arena predominated, persisting up to 6 days later, and that they frequently rotated in a manner that did not match that of the arena rotation. Furthermore, place-field maps were flexible, as mice frequently employed a similar, coherent configuration of place fields to represent each arena despite their differing geometry and eventual connection. These results highlight the ability of the hippocampus to retain consistent relationships between cells across long timescales and suggest that, in many cases, apparent instability might result from a coherent rotation of place fields.

The Same Hippocampal CA1 Population Simultaneously Codes Temporal Information over Multiple Timescales.

It has long been hypothesized that a primary function of the hippocampus is to discover and exploit temporal relationships between events. Previously, it has been reported that sequences of "time cells" in the hippocampus extend for tens of seconds. Other studies have shown that neuronal firing in the hippocampus fluctuates over hours and days. Both of these mechanisms could enable temporal encoding of events over very different timescales. However, thus far, these two classes of phenomena have never been observed simultaneously, which is necessary to ascribe broad-range temporal coding to the hippocampus. Using in vivo calcium imaging in unrestrained mice, we observed sequences of hippocampal neurons that bridged a 10 s delay. Similar sequences were observed over multiple days, but the set of neurons participating in those sequences changed gradually. Thus, the same population of neurons that encodes temporal information over seconds can also be used to distinguish periods of time over much longer timescales. These results unify two previously separate paradigms of temporal processing in the hippocampus that support episodic memory.