Hippocampus

Spatial memory is crucial for navigation and adapting to changing environmental conditions. Known neurophysiological mechanisms of spatial memory center on the importance of hippocampal activity and its spatial tuning. Yet, the behavioral strategies that support adaptive spatial encoding remain poorly understood. We have shown that dorsal hippocampal activity during rearing is necessary for spatial working memory, highlighting a role of information seeking behaviors for spatial memory encoding. Similarly, spatial tuning by dorsal hippocampal neurons is substantially updated during another information seeking behavior: attentive head scanning. However, the functional relationship between these behaviors is unknown. Here, to assess the relevance of environmental context for the expression of these behaviors, we quantified rearing and head scanning in a radial-arm-maze spatial working memory task while manipulating the height of the maze walls. Our goal was to test whether the stereotyped patterns of rearing that rats generate with tall walls are replaced with attentive head scanning when the walls are short enough to reach the top without rearing. We found that rats reared significantly less often when the walls were shortened and, instead, exhibited frequent attentive head scanning. The head scanning was done when and where the rats had previously exhibited stereotyped rearing. These results support the hypothesis that rearing and head scanning are functionally related behaviors. Future work should test two key inferences: 1) Head scanning is a critical epoch of spatial memory encoding, and 2) Spatial tuning by hippocampal neurons is updated during rearing. Significance statementSpatial memory is a core cognitive function, essential for healthy independent living. Though the hippocampus is critical for spatial memory, it remains unclear when and how. Separate prior studies link rearing and lateral head scanning to key periods of hippocampal processing, suggesting both behaviors support sensory information gathering for updating cognitive maps. However, their relationship is unresolved. Here, we test whether these behaviors are functionally interchangeable, with environmental structure determining expression. In a radial-arm maze, rats reared frequently with 21 cm walls but showed reduced rearing when walls were shortened to 4.6 cm, instead increasing head scanning at similar locations. These findings suggest rearing and head scanning share underlying motivations and provide a basis for comparing hippocampal activity during exploration.

The hippocampus is organized along dorsal-ventral and left-right axes, but whether and how these axes interact within defined neuronal populations across behavioral states remains unresolved. Here, we combined within-animal slice electrophysiology with dual-site fiber photometry to compare dorsal and ventral CA1 activity across contralateral hemispheric configurations in mice expressing CaMKII-jGCaMP8s and SynI-jRCaMP1b at distinct longitudinal sites. Ventral CA1 pyramidal neurons exhibited greater intrinsic excitability and stronger AMPAR-mediated synaptic responses than dorsal CA1 neurons. In vivo, CaMKII-defined pyramidal recordings during home cage rest revealed a left-biased event-rate asymmetry within dorsal but not ventral CA1, with no comparable asymmetry in pan-neuronal SynI recordings. Apparent dorsal-ventral differences in spontaneous event rate were therefore configuration-dependent and resolved into a hemispheric, cell-type-specific effect restricted to the CaMKII-defined population. Lead-lag analysis showed that dorsal-ventral temporal coordination was likewise reorganized across configurations and was restricted to pyramidal-cell-biased recordings. During open-field center entries, dorsal CA1 was preferentially recruited before entry across both configurations, whereas non-coordinated entries revealed a relative post-entry suppression of contralateral ventral CA1. Together, these findings suggest that dorsal-ventral CA1 organization cannot be inferred from hemisphere-pooled designs and identify a pyramidal-cell-specific left dorsal CA1 asymmetry as a structural feature that shapes both spontaneous activity and behaviorally driven recruitment along the longitudinal hippocampal axis. Significance StatementThe hippocampus is widely understood to differ along its long axis, with dorsal regions supporting spatial processing and ventral regions supporting emotional behavior. Whether this organization interacts with the left-right axis between hemispheres has remained essentially untested, because most studies pool hemispheres or record unilaterally. Using bilateral fiber photometry in mice, we show that spontaneous activity in dorsal CA1 is left-biased and that this asymmetry is specific to excitatory pyramidal neurons. The asymmetry explains apparent dorsal-ventral differences that appear configuration-dependent under conventional analysis, and it reshapes how dorsal and ventral CA1 are recruited during open-field exploration. These findings reframe hemispheric configuration from a methodological detail into an organizational variable that should be considered when interpreting hippocampal long-axis function.

In mouse hippocampal area CA1, excitatory pyramidal neurons referred to as "place cells" fire at specific locations in spatial environments during navigation. Many of the excitatory inputs to place cells are themselves spatially tuned, and prior work has shown that synaptic plasticity at those inputs strongly contributes to a selective increase in excitatory synaptic conductance when an animal is inside a cells "place field." It is less clear whether inhibitory inputs to place cells vary with spatial position. Recent studies have investigated whether place cells receive spatially tuned inhibitory conductances by recording place cell activity in vivo and using computational models to help interpret experimental perturbations. One prior study used inhibitory optogenetics to suppress inhibitory neuron firing rates and observed a uniform depolarization of place cells across spatial locations, supporting a model with spatially uniform synaptic inhibition. In apparent conflict, other studies used excitatory optogenetics to depolarize place cells and observed a selective increase in excitability within place fields, supporting a model with a spatially localized decrease in inhibition. However, the latter studies overlooked the contribution of voltage-gated NMDA-type glutamate receptors (NMDARs) to synaptic integration, which are expected to contribute to the balance of excitatory and inhibitory synaptic currents. Here we show that when NMDARs are included at excitatory synapses in simple CA1 place cell models, all experimentally-observed properties of place cells can be recapitulated regardless of whether inhibition increases, decreases, or remains constant inside a place field. Significance StatementThe hippocampus is a brain region required for the formation of new spatial and episodic memories (what happened where and when). Investigating the cellular and circuit mechanisms of memory recall could identify targets for therapies to combat memory decline associated with aging or neurodegeneration. Here we compare the results of computational models of the hippocampus to experimental recordings from mice to better understand the contribution of inhibitory neurons to the expression of spatial memories. We find that a special type of glutamate receptor, the NMDA receptor, helps to maintain the spatial selectivity of excitatory neurons in the hippocampus by counter-balancing fluctuations in the magnitude of inhibitory synaptic currents.

Interval timing (IT) is the ability to time events in the range from seconds to a few minutes, allowing animals to organize behavior in time at short durations. IT relies on two cognitive functions: 1) Measuring the passage of time; 2) Storing and retrieving temporal memories in a context appropriate manner. The hippocampus (HC) and medial prefrontal cortex (mPFC) have been shown critical to the accuracy and precision of time-contingent instrumental responses in IT. The anatomy supporting mPFC-HC interactions, required for memory encoding and retrieval, include projections from HC to mPFC, and indirect bidirectional connections through the ventral midline thalamus (VMT), most notably reuniens. Here, we explored VMTs role in retrieving fixed-interval (FI) temporal memories. Rats were trained on a 5s FI signaled by an auditory cue and demonstrated temporal memory by poking predominantly at the time of the expected reward. Timing responses on individual trials were classified into on-time, early, and random response. Across sessions, random response trials decreased following training. Next, we switched training to longer intervals (20s or 80s; daily sessions for weeks). To probe the role of the VMT in temporal memory retrieval, we infused the GABAA-agonist muscimol, or saline, before training sessions. Results show that VMT muscimol infusions decreased timing precision. Also, at both intervals, the number of on-time response trials decreased, and the number of random response trials significantly increased. The number of early response trials had no significant change at 20s, and significantly decreased at 80s. Overall, our results suggest that the VMT is critical for precise retrieval of temporal memories. We also describe per-trial response patterns with characteristics consistent across all trained intervals, suggesting multiple behavioral strategies at play during interval timing.

A major outstanding question in neuroscience is whether the neocortex uses the same powerful learning algorithm as current AI models: error backpropagation. One way this could be accomplished is as a function of the temporal derivative (i.e., differences in neural activity states over time), which can closely approximate the backpropagated error gradient. We tested the hypothesis that the direction of synaptic plasticity is a function of the temporal derivative in synaptic activity over the course of a 200 ms (5 Hz) theta cycle. Using mouse hippocampal slices, we drove presynaptic activity across the two 100 ms halves of a 200 ms window at either 25 Hz or 50 Hz, combined with corresponding low and high magnitudes of postsynaptic depolarization, testing all four 2x2 combinations of these low and high activity levels, while measuring the resulting effects on synaptic efficacy (as measured by EPSP amplitude to standard test probes). Consistent with the computational hypothesis, a positive temporal derivative (low to high) resulted in LTP (increased synaptic strength), while a negative temporal derivative (high to low) resulted in LTD. Critically, both no-change conditions (stable low or high across 200 ms) resulted in no net synaptic change, even though the high no-change condition had the highest overall synaptic activity levels. Possible biochemical mechanisms that could support these results are discussed.

The dentate gyrus (DG) is thought to play a key role in the formation of dissociable memory representations for similar contexts. Neurons in the DG receive highly processed spatial and nonspatial sensory information from the medial and lateral entorhinal cortices, respectively. Changes in spatially tuned firing patterns of DG place cells occur after spatial changes to an environment, but the degree to which DG place cells respond to ethologically relevant nonspatial stimuli is largely unknown. Spatial and nonspatial information is thought to be transmitted to the DG during discrete local field potential events called dentate spikes. Here, we tested the extent to which different spatial and nonspatial stimuli modulate place cell firing patterns and dentate spike dynamics. We performed extracellular recordings of DG place cells and local field potentials in rats of both sexes exploring a familiar spatial environment, in which social stimuli and nonsocial odors of varying ethological relevance were presented, and a novel spatial environment. As expected, DG place cells exhibited different firing patterns between familiar and novel environments. Significant changes in firing were not observed, however, with any of the nonspatial stimuli. Surprisingly, the occurrence of dentate spikes associated with lateral entorhinal cortex input increased during exploration of ethologically relevant stimuli, and this increase was greater for social stimuli. Altogether, these results suggest that the DG preferentially responds to social stimuli at the network level, providing novel insights into how spatial and nonspatial information is processed in the DG. Significance StatementThe dentate gyrus (DG) encodes spatial and nonspatial sensory information. Here, we investigated how place cells in the DG respond to changes in spatial and nonspatial cues in familiar and novel environments in rats. We found that DG place cell firing patterns significantly changed in a novel spatial environment but did not significantly change when nonspatial stimuli were presented in a familiar environment. Conversely, discrete dentate spike events reflecting presumed nonspatial inputs from the lateral entorhinal cortex increased during investigation of ethologically relevant nonspatial stimuli. These findings suggest novel mechanisms of nonspatial information processing in the DG.

The hippocampus participates in crucial functions such as memory consolidation, spatial processing and emotional regulation that require diverse input from multiple cortical areas that is funneled through the upper layers of the entorhinal cortex (EC), mostly from layer II to the dentate gyrus (DG). Traditional models described 200,000 EC layer II neurons projecting to 1 million granule cells (GCs) in the rat, rendering low divergence (1:5), with each EC neuron establishing about 18,000 synapses with GCs and each GC receiving about 4,000 synapses from EC neurons. In this manuscript, we update this model of connectivity incorporating new features described in the last three decades that include updated populations of EC layer II neurons obtained with design-based stereology, a revised definition of EC layer II based on molecular criteria and selecting reelin expressing neurons as the only layer II neurons projecting to the hippocampus. The updated model shows [~]80,000 neurons from EC layer II projecting to the DG, [~]45,000 from the medial entorhinal cortex (MEC) and [~]35,000 from the lateral entorhinal cortex (LEC) with high divergence of 1:20 and 1:30. We also show that EC layer II neurons may establish [~]90,000-115,000 synapses on GCs, while GCs receive about 8,000 synapses from EC layer II neurons. We estimate a [~]25% redundancy in the connectivity, so each EC neuron may contact [~]68,000-86,000 GCs and each GC would be contacted by [~]3000 neurons from MEC and 3,000 from LEC. In addition, we quantitatively assess a potential projection of mossy cells to the medial molecular layer described in mice, which could have a potential impact on GC inhibition. Overall, we produced a detailed, complete, and updated quantitative model of EC projections to the DG that reveals a much more divergent and richer projection than previously described, with implications for functional models (e.g.: pattern separation) and more widely for building realistic hippocampal models or establishing comparisons across species.

Dopamine signaling through dopamine 1 receptors (D1R) and dopamine 2 receptors (D2R) regulates hippocampal synaptic plasticity underlying learning and memory, yet their subcellular localization within the hippocampus is unknown. Here we performed electron microscopic immunocytochemistry to elucidate the distribution of D1R and D2R in subregions of the mouse hippocampus. In CA1 and CA3 stratum radiatum (SR), D1R- and D2R-immunoreactivity was found primarily on pyramidal cell dendritic spines and unmyelinated axons, and to a lesser extent in axon terminals and glia. In both regions, D1R-labeled terminals formed predominantly asymmetric (excitatory-type) synapses on dendritic spines, whereas D2R-labeled terminals formed mainly symmetric (inhibitory-type) synapses on pyramidal cell dendritic shafts. In the dentate gyrus (DG) hilus, D1R-labeling was almost exclusively found in unmyelinated axons and glia. D2R immunoreactivity in the hilus similarly was present in unmyelinated axons and glia but was also detected in dendritic spines originating from mossy cells and in terminals forming symmetric synapses. These findings indicate that dopamine receptors are positioned to influence excitatory and inhibitory signaling in the murine hippocampus. As D1R and D2R exert opposing effects on neuronal signaling, their localization on pyramidal neuron compartments provides a structural substrate for bidirectional modulation of synaptic plasticity and pyramidal cell activity. In addition, the presence of D2Rs on inhibitory terminals contacting pyramidal neurons and hilar interneurons suggests a role in regulating inhibitory circuitry within the hippocampus.

Background: Norepinephrine (NE) released from locus coeruleus (LC) projections regulates astrocyte structure and function through adrenergic receptor signaling. We previously showed that increasing noradrenergic tone with the NE reuptake inhibitor desipramine increases astrocyte ramification in the molecular layer of the dentate gyrus. However, whether tonic LC-derived noradrenergic tone is required to maintain astrocyte morphological complexity in vivo, and whether {beta}-adrenergic receptor activation is the effector pathway, remained unclear. Methods: Adult male C57BL/6J mice received DSP-4 (50 mg/kg X 3 days i.p.), a selective LC neurotoxin, with or without concurrent isoproterenol that continued for 21 additional days post cessation of DSP-4 treatment (ISO; 2 mg/kg/day X 24 days), or saline (n = 4 mice per group). Animals were sacrificed 22 days after the final DSP-4 injection. Noradrenergic denervation was confirmed by dopamine {beta}-hydroxylase (DBH) immunostaining. GFAP-immunostained astrocytes in the molecular layer of the dentate gyrus were morphologically characterized using Sholl analysis. Astrocyte density was quantified by SOX9 immunostaining. Results: DSP-4 produced >83% reduction in DBH fiber coverage in the molecular layer. Sholl analysis revealed significant reductions in astrocyte branching complexity in both treatment groups, with the reductions concentrated at distances of 5-15 m from the soma. The maximum number of intersections was also significantly reduced in both groups. Unexpectedly, ISO did not rescue morphological complexity. While DSP-4 alone did not alter astrocyte density, as measured by the number of SOX9-expressing astrocytes, DSP-4+ISO increased SOX9-positive cell density, dissociating the effects of adrenergic signaling on morphology from those on cell numbers. Conclusions: LC-derived noradrenergic tone is required for the maintenance of astrocyte arbour complexity in the dentate gyrus molecular layer. {beta}-adrenergic receptor activation alone is insufficient to restore structural integrity following noradrenergic denervation, yet promotes astrocyte density independently of structural remodeling. These findings have implications for understanding how LC neurodegeneration in Alzheimer's disease and depression may compromise hippocampal astrocyte structure and function.

The lateral septum (LS) integrates afferents from multiple brain regions, including the raphe nuclei. The organization of these inputs contributes to the regionalization of LS functions, for example spatial coding in dorsal LS and emotional regulation in ventral LS. Raphe-LS projections include glutamatergic axons expressing the vesicular glutamate transporter type 3 (VGLUT3), which often form pericellular baskets around LS neurons. This study provides an anatomical characterization of the organization and origins of VGLUT3-positive (VGLUT3+) raphe inputs to the LS. We mapped VGLUT3+ axon terminal density across the rostro-caudal extent of the LS and quantified colocalization with serotonin (5-HT) using immunohistochemistry. Our results showed that VGLUT3 density was highest in the ventral LS, whereas VGLUT3/5-HT colocalization was strongest in the dorsal LS. Retrograde viral vector-mediated tracing identified predominant inputs from the median raphe and B9 neuron group. Interestingly, the ventral hippocampus, a functionally related region which is known to also receive raphe VGLUT3 inputs, showed collaterals with the LS. Additional VGLUT3+ inputs to the LS arose from the interpeduncular nucleus, bed nucleus of the stria terminalis, nucleus incertus and pontine central gray. Anterograde tracing revealed that inputs from these brain regions target distinct and largely non-overlapping domains in the LS. Our findings highlight multiple sources of VGLUT3+ inputs to the LS, beyond the raphe nuclei, and suggest that distinct VGLUT3 circuits could contribute to LS functional specialization.

Memory impairment is a common and sometimes overlooked feature of major depressive disorder, and cognitive deficits may precede the onset of depressive symptoms in some cases. However, the cognitive benefits of first-line treatments such as SSRIs are mixed. Tianeptine is an atypical antidepressant and cognitive enhancer that neither interacts with monoamine receptors nor inhibits the reuptake of their neurotransmitters. Its antidepressant efficacy in animal models requires activation of the mu-opioid receptor (mu-OR) and phosphorylation of the AMPA receptor. However, the receptors that mediate its memory enhancing actions have never been investigated. We therefore tested the ability of tianeptine to improve spatial memory in a cross-maze task in wild-type (WT) mice compared to its effects in mice with global knockout of either the mu-OR or delta-OR. In parallel, we assessed the effects of tianeptine on hippocampal oscillatory activity and spontaneous locomotion in the same genotypes. Adult male and female WT, mu -/-, and delta -/- mice on a C57BL/6J background were implanted with hippocampal electrodes for the recording of local field potential (LFP) oscillations. Consistent with our previous observations in anaesthetised rats, injection of tianeptine (10 mg/kg and 30 mg/kg SC) caused a dose-dependent increase in beta-frequency power in WT mice that was maximal at circa 25 Hz. The same effect was observed in delta -/- mice, but the increase in beta was completely absent in mu -/- animals. As others have reported previously, tianeptine also caused a mu-OR-dependent increase in spontaneous locomotor activity, but with a time-course that was distinct from the increase in beta power. Separate groups of WT, mu -/-, and delta -/- mice were tested for their ability to learn a food-rewarded spatial memory task in a cross-maze. Over a 20-day training period, sub-groups of each genotype received either tianeptine (10 mg/kg SC) or vehicle injection 30 min before testing. Tianeptine increased the percentage of correct trials and the number of allocentric (place) responses in WT mice, but did not enhance memory in either mu -/- or delta -/- mice, even though both genotypes were able to learn the task. These results indicate that the ability of tianeptine to drive hippocampal beta oscillations is dependent on the mu-OR, whereas its memory-enhancing actions require the presence of both mu- and delta-ORs. The latter result is consistent with the actions of tianeptine on postsynaptic AMPA receptors, and we are currently exploring the signalling pathways involved in this process.

Why are some experiences remembered better than others? Leading theories propose that hippocampal replay prioritizes memories for consolidation according to their expected future value. We recorded hippocampal activity while rats experienced a sequence of novel environments associated with different reward values. We found that behavioral state, rather than reward, predicted replay rates during the task, and replay shifted away from reflecting current experiences as more experiences accumulated. During sleep, recent experiences were preferentially replayed, regardless of reward value. We integrate these findings into a model where each replay strengthens an episode-specific priority trace that decays over time but that can be refreshed by remote replay. We also identify a role for local replay in map stabilization, in support of the compositional mapping theory. Together, our findings demand a reinterpretation of utility-driven theories of prioritized replay and provide a framework to integrate temporal decaying episode strength into theories of memory triage and consolidation.

Long-term potentiation (LTP) is a key cellular mechanism underlying learning and memory, but its conservation across species remains unclear. Using nonhuman primates (NHPs), we examined hippocampal synaptic plasticity at Schaffer collateral- CA1 synapses. Theta-burst stimulation (TBS) reliably induced LTP in NHPs, comparable to rodents. However, unlike rodents, TBS in NHPs readily engaged synaptic tagging and capture (STC), indicating a lower threshold for associative plasticity. This was accompanied by increased expression of plasticity-related proteins, including PKM{zeta} and BDNF, suggesting enhanced recruitment of protein synthesisdependent stabilization mechanisms. These findings reveal a species-specific divergence in the molecular regulation of persistent synaptic plasticity and identify an evolutionary specialization in mechanisms supporting associative memory. Together, our results highlight limitations of rodent models in fully capturing human-relevant memory processes and underscore the importance of primate systems for translational neuroscience.

Hippocampal area CA2 has recently emerged as a critical region for social recognition memory. Furthermore, this understudied region has been implicated in psychiatric diseases and neurodegenerative diseases. There has been accumulating evidence indicating that the pyramidal neurons (PNs) in area CA2 exhibit functional specializations that correlate with somatic position in stratum pyramidale (sp). In this study, we investigated the morphological differences in dendritic architecture of CA2 PNs with a focus on the radial gradient, i.e., along the deep-superficial axis of the sp. We conducted a comprehensive morphological analysis including Sholl intersection profiles, branching order distributions, root angle distributions, and dendritic cable lengths. We found that CA2 PNs have fewer oblique dendrites and a larger number of tuft-like dendrites as compared to CA1 PNs. Furthermore, within the CA2 population, we found that many of the dendritic structural features gradually changed along the radial axis from deep to superficial somatic location, indicating a continuum of dendritic morphology rather than two sharply defined subtypes of pyramidal neurons. This morphological characterization may serve as a starting point to better understand the corresponding functional organization of CA2. The gradual difference between deeper and superficial CA2 PNs suggests a continuum of their computational capabilities beyond two binary functional classes. In briefUsing several methods, we examine the dendritic morphology of over 130 CA2 and CA1 pyramidal neurons and find that many properties such as the cable length and terminal numbers of the dendritic arbors vary as a with the location of the soma in the pyramidal layer. HighlightsO_LIWe use scholl analysis, graph theory and machine learning techniques to quantify the different dendritic morphologies of CA2 pyramidal neurons. C_LIO_LIMany properties of CA2 pyramidal neuron apical dendrites vary as a function of somatic location in the pyramidal layer. C_LIO_LIMore superficial CA2 pyramidal neurons have longer oblique apical dendrites, and shorter tuft dendrites. C_LI

Navigation requires perception: location must be inferred from noisy and ambiguous egocentric sensory inputs, as in visual estimation of distance. However, many classical models of spatial representation implicitly assume that allocentric location is directly observable, thereby neglecting perceptual uncertainty. Here, we compare such a model with a Bayesian ideal observer that explicitly incorporates perceptual inference. We find that the Bayesian observers beliefs over location more accurately reproduce key properties of place cell activity, including place field width, area, and density, within and across environments. Using analytic arguments and numerical simulations, we show that recurrent neural networks trained to predict the next egocentric sensory input learn representations resembling Bayesian beliefs and yield place cell-like activity in both familiar and unfamiliar environments, outperforming autoencoders trained to reproduce the current input. Together, these results suggest that hippocampal circuits may construct Bayesian cognitive maps from experience through predictive perceptual learning.

The hippocampus and its projection targets constitute hippocampal output circuits that are essential for memory, navigation, and emotional regulation. Although cortical and subcortical regions receiving hippocampal projections have been well characterized, it remains unclear how hippocampal signals are processed within these projection target regions. To precisely understand information processing in hippocampal output circuits, it is therefore important to gain genetic access to neuronal subpopulations receiving direct hippocampal inputs for structural and functional analyses. Anterograde transsynaptic transduction using adeno-associated virus (AAV) serotype 1 has emerged as a powerful approach for targeting postsynaptic neurons, but its application is limited by unintended retrograde transport, which leads to false-positive labeling in reciprocally connected circuits. Here, we developed a direction-selective anterograde transsynaptic transduction by combining AAV vectors with distinct infection properties and an intersectional gene expression system. Using the hippocampal-entorhinal circuit, we identified an optimal viral combination that enables predominantly anterograde transsynaptic labeling. We further applied this method to map hippocampal projections to reciprocally connected regions, including the amygdala, providing a robust approach for dissecting complex hippocampal output circuits.

The hippocampus is characterized by a stereotypical macroscopic structure, where the nuclei are densely and heterogeneously packed among different subregions of the hippocampus. Despite the fact that tissue-specific cellular organization has been implicated in neural function, it has been technically challenging to quantitatively analyze mesoscopic cellular organization in the hippocampus due to its high cellular density. To overcome this technical hurdle, we developed Computational Biophysical Histomorphometry Software (CBHS), an automated image-analysis pipeline, aimed at quantifying nuclear shape and the order of the cellular ensemble in high-density areas. When applied to the subfields of hippocampus, we found that denser regions, most notably the dentate gyrus, were the most positionally, but least orientationally ordered. Nuclear shape exhibited a dependence on the local environment in a packing-dependent manner. This association was cell-type specific, with neurons, but not astrocytes displaying nuclear shape that varied with neighbour proximity, although astrocytes demonstrated greater intrinsic shape variance. The results reveal the presence of reproducible mesoscale cell packing order in hippocampal tissue, and are consistent with a nucleus-driven mechanical coupling between neighbouring cells. The present study provides a quantitative framework with which to understand mesoscopic tissue organization, thus enabling the formulation of testable hypotheses for future investigation.

Sleep is thought to promote memory consolidation through the offline reactivation and reorganization of newly acquired information. However, most studies assess memory shortly after sleep, leaving unresolved whether an initial post-learning sleep episode produces enduring modifications that influence how memories respond to later reactivation. Importantly, the absence of behavioral differences after prolonged retention intervals does not necessarily imply that sleep failed to modify the original memory. Instead, sleep-dependent changes may persist in latent forms that are not readily captured by conventional memory assessments. Here, we investigated whether post-learning sleep produces lasting changes in declarative memories that influence their subsequent response to reactivation. In Study 1, participants learned a declarative memory task and were assigned to either a short nap, a wake condition, or an exploratory long-nap condition that included both NREM and REM sleep. Memory was assessed one week later. Despite substantial forgetting across the retention interval, no significant differences in memory performance were observed between groups. In Study 2, participants learned the same task and subsequently underwent either a short nap or wakefulness. Memory was reactivated six days after learning using an incomplete reminder previously shown to induce memory updating in human declarative memory, and memory was tested one day later. Under these conditions, participants who slept after learning showed better memory performance than wake controls. Moreover, sleep physiological measures predicted the magnitude of the post-reactivation memory benefit. These findings suggest that post-learning sleep induces enduring modifications in declarative memories that are not readily detectable through delayed memory testing alone. Instead, these sleep-dependent changes become evident when memories are challenged through subsequent reactivation. Our results indicate that sleep-dependent consolidation influences the future expression of memory, shaping how memories respond to later reactivation experiences and providing new insight into the relationship between consolidation and reconsolidation.

Learning from aversive experiences often generalizes beyond the context in which they occurred. In rodents, a strong aversive event can induce retrospective memory linking (RLI), whereby fear generalizes to a previously neutral context encountered days earlier. Although prior work has shown that RLI is associated with increased co-activity of hippocampal CA1 neurons across neutral and aversive contexts, it remains unclear how broader representational changes support generalization without affecting the ability to discriminate between contexts. Here, we reanalyzed calcium imaging data from dorsal CA1 during RLI to examine how hippocampal representational geometry changes during fear generalization. Using robust, non-parametric measures of population similarity, we show that in mice exhibiting RLI, the representation of the neutral context not only changes over time but becomes more similar to the aversive context during recall. Beyond this similarity increase, we provide evidence for a higher-dimensional geometric transformation consistent with a shared "fear" operation that can be applied across contexts while preserving their identity. Crucially, these two representational signatures dissociate by behavioral state: similarity to the aversive context emerges during freezing, whereas a shared transformation is expressed during active exploration. Together, these findings demonstrate that hippocampal representations support retrospective fear generalization through state-dependent geometric transformations, highlighting representational geometry as a key computational mechanism to resolve the apparent tension between generalization and discriminability.

High-frequency "ripple" oscillations support learning and memory across species, yet it has been argued that putative ripples in awake human recordings are false positives produced when algorithms misread aperiodic (1/f) fluctuations as ripple-band oscillations. We show that this conclusion arises from an artifact of evaluating detection algorithms on surrogate data containing only aperiodic activity. Ripple detectors are adaptive, setting their threshold from the amplitude statistics of the signal, so applying them to surrogate data that contains only aperiodic activity lowers the threshold and inflates false positives (median 62%). Adding real ripple-band events back to the surrogate corrects this threshold shift and eliminates most false detections across multiple standard algorithms. Using multivariate classifiers, we show aperiodic fluctuations can reproduce the power of ripples but not their timing or spectral content. These findings indicate care needs to be taken when using surrogates to evaluate ripple detection algorithms. Thus, under realistic signal properties, human hippocampal ripples remain distinguishable from aperiodic activity.