Astrocyte-neuron lactate transport is required for long-term memory formation.

We report that, in the rat hippocampus, learning leads to a significant increase in extracellular lactate levels that derive from glycogen, an energy reserve selectively localized in astrocytes. Astrocytic glycogen breakdown and lactate release are essential for long-term but not short-term memory formation, and for the maintenance of long-term potentiation (LTP) of synaptic strength elicited in vivo. Disrupting the expression of the astrocytic lactate transporters monocarboxylate transporter 4 (MCT4) or MCT1 causes amnesia, which, like LTP impairment, is rescued by L-lactate but not equicaloric glucose. Disrupting the expression of the neuronal lactate transporter MCT2 also leads to amnesia that is unaffected by either L-lactate or glucose, suggesting that lactate import into neurons is necessary for long-term memory. Glycogenolysis and astrocytic lactate transporters are also critical for the induction of molecular changes required for memory formation, including the induction of phospho-CREB, Arc, and phospho-cofilin. We conclude that astrocyte-neuron lactate transport is required for long-term memory formation.

Learning-induced gene expression in the heads of two Nasonia species that differ in long-term memory formation.

BACKGROUND: Cellular processes underlying memory formation are evolutionary conserved, but natural variation in memory dynamics between animal species or populations is common. The genetic basis of this fascinating phenomenon is poorly understood. Closely related species of Nasonia parasitic wasps differ in long-term memory (LTM) formation: N. vitripennis will form transcription-dependent LTM after a single conditioning trial, whereas the closely-related species N. giraulti will not. Genes that were differentially expressed (DE) after conditioning in N. vitripennis, but not in N. giraulti, were identified as candidate genes that may regulate LTM formation. RESULTS: RNA was collected from heads of both species before and immediately, 4 or 24 hours after conditioning, with 3 replicates per time point. It was sequenced strand-specifically, which allows distinguishing sense from antisense transcripts and improves the quality of expression analyses. We determined conditioning-induced DE compared to naïve controls for both species. These expression patterns were then analysed with GO enrichment analyses for each species and time point, which demonstrated an enrichment of signalling-related genes immediately after conditioning in N. vitripennis only. Analyses of known LTM genes and genes with an opposing expression pattern between the two species revealed additional candidate genes for the difference in LTM formation. These include genes from various signalling cascades, including several members of the Ras and PI3 kinase signalling pathways, and glutamate receptors. Interestingly, several other known LTM genes were exclusively differentially expressed in N. giraulti, which may indicate an LTM-inhibitory mechanism. Among the DE transcripts were also antisense transcripts. Furthermore, antisense transcripts aligning to a number of known memory genes were detected, which may have a role in regulating these genes. CONCLUSION: This study is the first to describe and compare expression patterns of both protein-coding and antisense transcripts, at different time points after conditioning, of two closely related animal species that differ in LTM formation. Several candidate genes that may regulate differences in LTM have been identified. This transcriptome analysis is a valuable resource for future in-depth studies to elucidate the role of candidate genes and antisense transcription in natural variation in LTM formation.

The role of retinoic acid in long-term memory formation and synaptic plasticity in the mollusc Lymnaea stagnalis

The active metabolite of vitamin A, retinoic acid (RA), is involved in memory formation and hippocampal plasticity in vertebrates. A similar role for retinoid signaling in learning and memory formation has not previously been examined in an invertebrate species. However, the conservation of retinoid signaling between vertebrates and invertebrates is supported by the presence of retinoid signaling machinery in invertebrates. For example, in the mollusc Lymnaea stagnalis the metabolic enzymes and retinoid receptors have been cloned from the CNS. In this study I demonstrated that impairing retinoid signaling in Lymnaea by either inhibiting RALDH activity or using retinoid receptor antagonists, prevented the formation of long-term memory (LTM). However, learning and intermediate-term memory were not affected. An additional finding was that exposure to constant darkness (due to the light-sensitive nature of RA) itself enhanced memory formation. This memory-promoting effect of darkness was sufficient to overcome the inhibitory effects of RALDH inhibition, but not that of a retinoid receptor antagonist, suggesting that environmental light conditions may influence retinoid signaling. Since RA also influences synaptic plasticity underlying hippocampal-dependent memory formation, I also examined whether RA would act in a trophic manner to influence synapse formation and/or synaptic transmission between invertebrate neurons. However, I found no evidence to support an effect of RA on post-tetanic potentiation of a chemical synapse. Retinoic acid did, however, reduce transmission at electrical synapses in a cell-specific manner. Overall, these studies provide the first evidence for a role of RA in the formation of implicit long-term memories in an invertebrate species and suggest that the role of retinoid signaling in memory formation has an ancient origin.

Location, location, location: The many addresses of memory formation

Memory formation, like real estate, can be summarized succinctly—location, location, location. It is an emergent property involving different anatomical regions in the brain, sets of neuronal circuits, and cellular and molecular interactions between and within those neurons. At each of these levels of description, location continues to be a major organizing principle guiding researchers. The difficulty in the field is the integration of information between the various levels of analyses, and it is proposed that molecular reporters may help to fill that void.

Expectation Modulates Episodic Memory Formation via Dopaminergic Circuitry

Episodic memory formation is shaped by expectation. Events that generate expectations have the capacity to influence memory. Additionally, whether subsequent events meet or violate expectations has consequences for memory. However, clarification is still required to illuminate the circumstances and direction of memory modulation. In the brain, the mechanisms by which expectation modulates memory formation also require consideration. The dopamine system has been implicated in signaling events associated with different states of expectancy; it has also been shown to modulate episodic memory formation in the hippocampus. Thus, the studies included in this dissertation utilized both functional magnetic resonance imaging (fMRI) and behavioral testing to examine when and how the dopaminergic system supports the modulation of memory by expectation. The work aimed to characterize the activation of dopaminergic circuitry in response to cues that generate expectancy, during periods of anticipation, and in response to outcomes that resolve expectancy. The studies also examined how each of these event types influenced episodic memory formation. The present findings demonstrated that novelty and expectancy violation both drive dopaminergic circuitry capable of contributing to memory formation. Consistent with elevated dopaminergic midbrain and hippocampus activation for each, expected versus expectancy violating novelty did not differentially affect memory success. We also showed that high curiosity expectancy states drive memory formation. This was supported by activation in dopaminergic circuitry that was greater for subsequently remembered information only in the high curiosity state. Finally, we showed that cues that generate high expected reward value versus high reward uncertainty differentially modulate memory formation during reward anticipation. This behavioral result was consistent with distinct temporal profiles of dopaminergic action having differential downstream effects on episodic memory formation. Integrating the present studies with previous research suggests that dopaminergic circuitry signals events that are unpredicted, whether cuing or resolving expectations. It also suggests that contextual differences change the contribution of the dopaminergic system during anticipation, depending on the nature of the expectation. And finally, this work is consistent with a model in which dopamine elevation in response to expectancy events positively modulates episodic memory formation.

Formation of a spatial memory trace : a behavioral, cellular and molecular study

THESIS ABSTRACTThis thesis project was aimed at studying the molecular mechanisms underlying learning and memory formation, in particular as they relate to the metabolic coupling between astrocytes and neurons. For that, changes in the metabolic activity of different mice brain regions after 1 or 9 days of training in an eight-arm radial maze were assessed by (14C) 2-deoxyglucose (2DG) autoradiography. Significant differences in the areas engaged during the behavioral task at day 1 (when animals are confronted for the first time to the learning task) and at day 9 (when animals are highly performing) have been identified. These areas include the hippocampus, the fornix, the parietal cortex, the laterodorsal thalamic nucleus and the mammillary bodies at day 1 ; and the anterior cingulate, the retrosplenial cortex and the dorsal striatum at day 9. Two of these cerebral regions (those presenting the greatest changes at day 1 and day 9: the hippocampus and the retrosplenial cortex, respectively) were microdissected by laser capture microscopy and selected genes related to neuron-glia metabolic coupling, glucose metabolism and synaptic plasticity were analyzed by RT-PCR. 2DG and gene expression analysis were performed at three different times: 1) immediately after the end of the behavioral paradigm, 2) 45 minutes and 3) 6 hours after training. The main goal of this study was the identification of the metabolic adaptations following the learning task. Gene expression results demonstrate that the learning task profoundly modulates the pattern of gene expression in time, meaning that these two cerebral regions with high 2DG signal (hippocampus and retrosplenial cortex) have adapted their metabolic molecular machinery in consequence. Almost all studied genes show a higher expression in the hippocampus at day 1 compared to day 9, while an increased expression was found in the retrosplenial cortex at day 9. We can observe these molecular adaptations with a short delay of 45 minutes after the end of the task. However, 6 hours after training a high gene expression was found at day 9 (compared to day 1) in both regions, suggesting that only one day of training is not sufficient to detect transcriptional modifications several hours after the task. Thus, gene expression data match 2DG results indicating a transfer of information in time (from day 1 to day 9) and in space (from the hippocampus to the retrosplenial cortex), and this at a cellular and a molecular level. Moreover, learning seems to modify the neuron-glia metabolic coupling, since several genes involved in this coupling are induced. These results also suggest a role of glia in neuronal plasticity.RESUME DU TRAVAIL DE THESECe projet de thèse a eu pour but l'étude des mécanismes moléculaires qui sont impliqués dans l'apprentissage et la mémoire et, en particulier, à les mettre en rapport avec le couplage métabolique existant entre les astrocytes et les neurones. Pour cela, des changements de l'activité métabolique dans différentes régions du cerveau des souris après 1 ou 9 jours d'entraînement dans un labyrinthe radial à huit-bras ont été évalués par autoradiographie au 2-désoxyglucose (2DG). Des différences significatives dans les régions engagées pendant la tâche comportementale au jour 1 (quand les animaux sont confrontés pour la première fois à la tâche) et au jour 9 (quand les animaux ont déjà appris) ont été identifiés. Ces régions incluent, au jour 1, l'hippocampe, le fornix, le cortex pariétal, le noyau thalamic laterodorsal et les corps mamillaires; et, au jour 9, le cingulaire antérieur, le cortex retrosplenial et le striatum dorsal. Deux de ces régions cérébrales (celles présentant les plus grands changements à jour 1 et à jour 9: l'hippocampe et le cortex retrosplenial, respectivement) ont été découpées par microdissection au laser et quelques gènes liés au couplage métabolique neurone-glie, au métabolisme du glucose et à la plasticité synaptique ont été analysées par RT-PCR. L'étude 2DG et l'analyse de l'expression de gènes ont été exécutés à trois temps différents: 1) juste après entraînement, 2) 45 minutes et 3) 6 heures après la fin de la tâche. L'objectif principal de cette étude était l'identification des adaptations métaboliques suivant la tâche d'apprentissage. Les résultats de l'expression de gènes démontrent que la tâche d'apprentissage module profondément le profile d'expression des gènes dans le temps, signifiant que ces deux régions cérébrales avec un signal 2DG élevé (l'hippocampe et le cortex retrosplenial) ont adapté leurs « machines moléculaires » en conséquence. Presque tous les gènes étudiés montrent une expression plus élevée dans l'hippocampe au jour 1 comparé au jour 9, alors qu'une expression accrue a été trouvée dans le cortex retrosplenial au jour 9. Nous pouvons observer ces adaptations moléculaires avec un retard court de 45 minutes après la fin de la tâche. Cependant, 6 heures après l'entraînement, une expression de gènes élevée a été trouvée au jour 9 (comparé à jour 1) dans les deux régions, suggérant que seulement un jour d'entraînement ne suffit pas pour détecter des modifications transcriptionelles plusieurs heures après la tâche. Ainsi, les données d'expression de gènes corroborent les résultats 2DG indiquant un transfert d'information dans le temps (de jour 1 à jour 9) et dans l'espace (de l'hippocampe au cortex retrosplenial), et ceci à un niveau cellulaire et moléculaire. D'ailleurs, la tâche d'apprentissage semble modifier le couplage métabolique neurone-glie, puisque de nombreux gènes impliqués dans ce couplage sont induits. Ces observations suggèrent un rôle important de la glie dans les mécanismes de plasticité du système nerveux.

TCR signaling requirements for activating T cells and for generating memory.

Over the last two decades the molecular and cellular mechanisms underlying T cell activation, expansion, differentiation, and memory formation have been intensively investigated. These studies revealed that the generation of memory T cells is critically impacted by a number of factors, including the magnitude of the inflammatory response and cytokine production, the type of dendritic cell [DC] that presents the pathogen derived antigen, their maturation status, and the concomitant provision of costimulation. Nevertheless, the primary stimulus leading to T cell activation is generated through the T cell receptor [TCR] following its engagement with a peptide MHC ligand [pMHC]. The purpose of this review is to highlight classical and recent findings on how antigen recognition, the degree of TCR stimulation, and intracellular signal transduction pathways impact the formation of effector and memory T cells.

Dendritic Spine Abnormalities in Hippocampal CA1 Pyramidal Neurons Underlying Memory Deficits in the SAMP8 Mouse Model of Alzheimer's Disease

SAMP8 is a strain of mice with accelerated senescence. These mice have recently been the focus of attention as they show several alterations that have also been described in Alzheimer"s disease (AD) patients. The number of dendritic spines, spine plasticity, and morphology are basic to memory formation. In AD, the density of dendritic spines is severely decreased. We studied memory alterations using the object recognition test. We measured levels of synaptophysin as a marker of neurotransmission and used Golgi staining to quantify and characterize the number and morphology of dendritic spines in SAMP8 mice and in SAMR1 as control animals. While there were no memory differences at 3 months of age, the memory of both 6- and 9-month-old SAMP8 mice was impaired in comparison with age-matched SAMR1 mice or young SAMP8 mice. In addition, synaptophysin levels were not altered in young SAMP8 animals, but SAMP8 aged 6 and 9 months had less synaptophysin than SAMR1 controls and also less than 3-month-old SAMP8 mice. Moreover, while spine density remained stable with age in SAMR1 mice, the number of spines started to decrease in SAMP8 animals at 6 months, only to get worse at 9 months. Our results show that from 6 months onwards SAMP8 mice show impaired memory. This age coincides with that at which the levels of synaptophysin and spine density decrease. Thus, we conclude that together with other studies that describe several alterations at similar ages, SAMP8 mice are a very suitable model for studying AD.

Mammalian Target of Rapamycin Complex 2 Controls CD8 T Cell Memory Differentiation in a Foxo1-Dependent Manner.

Upon infection, antigen-specific naive CD8 T cells are activated and differentiate into short-lived effector cells (SLECs) and memory precursor cells (MPECs). The underlying signaling pathways remain largely unresolved. We show that Rictor, the core component of mammalian target of rapamycin complex 2 (mTORC2), regulates SLEC and MPEC commitment. Rictor deficiency favors memory formation and increases IL-2 secretion capacity without dampening effector functions. Moreover, mTORC2-deficient memory T cells mount more potent recall responses. Enhanced memory formation in the absence of mTORC2 was associated with Eomes and Tcf-1 upregulation, repression of T-bet, enhanced mitochondrial spare respiratory capacity, and fatty acid oxidation. This transcriptional and metabolic reprogramming is mainly driven by nuclear stabilization of Foxo1. Silencing of Foxo1 reversed the increased MPEC differentiation and IL-2 production and led to an impaired recall response of Rictor KO memory T cells. Therefore, mTORC2 is a critical regulator of CD8 T cell differentiation and may be an important target for immunotherapy interventions.

Involvement of hippocampal AMPA glutamate receptor changes and the cAMP/protein kinase A/CREB-P signalling pathway in memory consolidation of an avoidance task in rats

Training in step-down inhibitory avoidance (0.3-mA footshock) is followed by biochemical changes in rat hippocampus that strongly suggest an involvement of quantitative changes in glutamate AMPA receptors, followed by changes in the dopamine D1 receptor/cAMP/protein kinase A (PKA)/CREB-P signalling pathway in memory consolidation. AMPA binding to its receptor and levels of the AMPA receptor-specific subunit GluR1 increase in the hippocampus within the first 3 h after training (20-70%). Binding of the specific D1 receptor ligand, SCH23390, and cAMP levels increase within 3 or 6 h after training (30-100%). PKA activity and CREB-P levels show two peaks: a 35-40% increase 0 h after training, and a second increase 3-6 h later (35-60%). The results correlate with pharmacological findings showing an early post-training involvement of AMPA receptors, and a late involvement of the D1/cAMP/PKA/CREB-P pathway in memory consolidation of this task

Agents that affect cAMP levels or protein kinase A activity modulate memory consolidation when injected into rat hippocampus but not amygdala

Male Wistar rats were trained in one-trial step-down inhibitory avoidance using a 0.4-mA footshock. At various times after training (0, 1.5, 3, 6 and 9 h for the animals implanted into the CA1 region of the hippocampus; 0 and 3 h for those implanted into the amygdala), these animals received microinfusions of SKF38393 (7.5 µg/side), SCH23390 (0.5 µg/side), norepinephrine (0.3 µg/side), timolol (0.3 µg/side), 8-OH-DPAT (2.5 µg/side), NAN-190 (2.5 µg/side), forskolin (0.5 µg/side), KT5720 (0.5 µg/side) or 8-Br-cAMP (1.25 µg/side). Rats were tested for retention 24 h after training. When given into the hippocampus 0 h post-training, norepinephrine enhanced memory whereas KT5720 was amnestic. When given 1.5 h after training, all treatments were ineffective. When given 3 or 6 h post-training, 8-Br-cAMP, forskolin, SKF38393, norepinephrine and NAN-190 caused memory facilitation, while KT5720, SCH23390, timolol and 8-OH-DPAT caused retrograde amnesia. Again, at 9 h after training, all treatments were ineffective. When given into the amygdala, norepinephrine caused retrograde facilitation at 0 h after training. The other drugs infused into the amygdala did not cause any significant effect. These data suggest that in the hippocampus, but not in the amygdala, a cAMP/protein kinase A pathway is involved in memory consolidation at 3 and 6 h after training, which is regulated by D1, ß, and 5HT1A receptors. This correlates with data on increased post-training cAMP levels and a dual peak of protein kinase A activity and CREB-P levels (at 0 and 3-6 h) in rat hippocampus after training in this task. These results suggest that the hippocampus, but not the amygdala, is involved in long-term storage of step-down inhibitory avoidance in the rat.

Emotion strengthens high priority memory traces but weakens low priority memory traces

When people encounter emotional events, their memory for those events is typically enhanced. But it has been unclear how emotionally arousing events influence memory for preceding information. Does emotional arousal induce retrograde amnesia or retrograde enhancement? The current study revealed that this depends on the top-down goal relevance of the preceding information. Across three studies, we found that emotional arousal induced by one image facilitated memory for the preceding neutral item when people prioritized that neutral item. In contrast, an emotionally arousing image impaired memory for the preceding neutral item when people did not prioritize that neutral item. Emotional arousal elicited by both negative and positive pictures showed this pattern of enhancing or impairing memory for the preceding stimulus depending on its priority. These results indicate that emotional arousal amplifies the effects of top-down priority in memory formation.

Hippocampal functional connectivity reflects verbal episodic memory network integrity

Using functional magnetic resonance imaging during a verbal memory task, we investigated correlations of signal fluctuations within the hippocampus and ipsilateral frontal as well as temporal areas in temporal lobe epilepsy patients. Declarative memory abilities were additionally examined before and after temporal lobe epilepsy surgery. A significant difference exists in functional connectivity between patients whose mnemonic functions deteriorated and those who remained stable or improved. Univariate analyses showed significantly higher preoperative coupling between the hippocampus and Brodmann area 22 for the group that decreased in verbal learning. We suggest greater coupling to reflect higher functional network integrity. Postoperatively reduced learning ability in patients with higher preoperative coupling underlines the importance of hippocampal interaction with cortical areas for successful memory formation.

Beta 1-integrins are required for hippocampal AMPA receptor-dependent synaptic transmission, synaptic plasticity, and working memory.

Integrins comprise a large family of cell adhesion receptors that mediate diverse biological events through cell-cell and cell-extracellular matrix interactions. Recent studies have shown that several integrins are localized to synapses with suggested roles in synaptic plasticity and memory formation. We generated a postnatal forebrain and excitatory neuron-specific knock-out of beta1-integrin in the mouse. Electrophysiological studies demonstrated that these mutants have impaired synaptic transmission through AMPA receptors and diminished NMDA receptor-dependent long-term potentiation. Despite the impairment in hippocampal synaptic transmission, the mutants displayed normal hippocampal-dependent spatial and contextual memory but were impaired in a hippocampal-dependent, nonmatching-to-place working memory task. These phenotypes parallel those observed in animals carrying knock-outs of the GluR1 (glutamate receptor subunit 1) subunit of the AMPA receptor. These observations suggest a new function of beta1-integrins as regulators of synaptic glutamate receptor function and working memory.

Bifurcation and singularity analysis of a molecular network for the induction of long-term memory.

Withdrawal reflexes of the mollusk Aplysia exhibit sensitization, a simple form of long-term memory (LTM). Sensitization is due, in part, to long-term facilitation (LTF) of sensorimotor neuron synapses. LTF is induced by the modulatory actions of serotonin (5-HT). Pettigrew et al. developed a computational model of the nonlinear intracellular signaling and gene network that underlies the induction of 5-HT-induced LTF. The model simulated empirical observations that repeated applications of 5-HT induce persistent activation of protein kinase A (PKA) and that this persistent activation requires a suprathreshold exposure of 5-HT. This study extends the analysis of the Pettigrew model by applying bifurcation analysis, singularity theory, and numerical simulation. Using singularity theory, classification diagrams of parameter space were constructed, identifying regions with qualitatively different steady-state behaviors. The graphical representation of these regions illustrates the robustness of these regions to changes in model parameters. Because persistent protein kinase A (PKA) activity correlates with Aplysia LTM, the analysis focuses on a positive feedback loop in the model that tends to maintain PKA activity. In this loop, PKA phosphorylates a transcription factor (TF-1), thereby increasing the expression of an ubiquitin hydrolase (Ap-Uch). Ap-Uch then acts to increase PKA activity, closing the loop. This positive feedback loop manifests multiple, coexisting steady states, or multiplicity, which provides a mechanism for a bistable switch in PKA activity. After the removal of 5-HT, the PKA activity either returns to its basal level (reversible switch) or remains at a high level (irreversible switch). Such an irreversible switch might be a mechanism that contributes to the persistence of LTM. The classification diagrams also identify parameters and processes that might be manipulated, perhaps pharmacologically, to enhance the induction of memory. Rational drug design, to affect complex processes such as memory formation, can benefit from this type of analysis.