Single Training Trial Research Articles

Aversive Bimodal Associations Differently Impact Visual and Olfactory Memory Performance in Drosophila.

Animals form sensory associations and store them as memories to guide behavioral decisions. Although unimodal learning has been studied extensively in insects, it is important to explore sensory cues in combination because most behaviors require multimodal inputs. In our study, we optimized the T-maze to employ both visual and olfactory cues in a classical aversive learning paradigm in Drosophila melanogaster. In contrast to unimodal training, bimodal training evoked a significant short-term visual memory after a single training trial. Interestingly, the same protocol did not enhance short-term olfactory memory and even had a negative impact. However, compromised long-lasting olfactory memory significantly improved after bimodal training. Our study demonstrates that the effect of bimodal integration on learning is not always beneficial and is conditional upon the formed memory strengths. We postulate that flies utilize information on a need-to basis: bimodal training augments weakly formed memories while stronger associations are impacted differently.

The Analysis of Aversive Olfactory-Taste Learning and Memory in Drosophila Larvae.

Drosophila larvae are able to associate an odor stimulus with a temporally overlapping teaching signal encoding reward or punishment. Here, we describe a standardized experimental setup that allows the analysis of larval aversive-odor-taste learning and memory. This is a Pavlovian learning experiment with a single training trial in which larvae are presented with two specific odors in succession, one odor together with salt at a high concentration that is harmful to the larva. In the subsequent test, the trained larvae then show avoidance of the salt-paired odor and spend more time near the unpaired odor. To rule out nonassociative effects (such as naive preferences for odors, exposure, or handling effects), two independent groups of larvae are reciprocally trained. Subsequently, the average of the two individual preference values is determined and quantified as a Performance Index (PI), which assigns a numerical value to the larvae's shown behavioral change.

Learning a non-neutral conditioned stimulus: place preference in the crab Neohelice granulata.

In the wild, being able to recognize and remember specific locations related to food sources and the associated attributes of landmarks is a cognitive trait important for survival. In the present work, we show that the crab Neohelice granulata can be trained to associate a specific environment with an appetitive reward in a conditioned place preference task. After a single training trial, when the crabs were presented with a food pellet in the target quadrant of the training arena, they were able to form a long-term memory related to the event. This memory was evident at least 24 h after training and was protein synthesis dependent. Importantly, the target area of the arena proved to be a non-neutral environment, given that animals initially avoided the target quadrant. In the present work, we introduce for the first time an associative one-trial memory paradigm including a conditioned stimulus with a clear valence performed in a crustacean.

Inhibition of transcription and translation in dorsal hippocampus does not interfere with consolidation of memory of intense training.

Findings of several experiments indicate that many treatments that typically interfere with memory consolidation are ineffective in preventing or attenuating memory induced by intense training. As extensive evidence suggests that the consolidation of newly acquired memories requires gene expression and de novo protein synthesis the present study investigated whether intense training prevents consolidation impairment induced by blockers of mRNA and protein synthesis. Rats were given a single inhibitory training trial using a moderate (1.0 mA) or a relatively intense (2.0 mA) foot-shock. Bilateral hippocampal infusions of the mRNA synthesis blocker DRB (10, 40 or 80 ng/0.5 µL/hemisphere) or the protein synthesis inhibitor anisomycin (ANI), an inhibitor de novo protein synthesis (15.62, 31.25, or 62.50 µg/0.5 µL/hemisphere) were administered 15 min prior to training. Retention was measured at 30 min or 48 h following training. DRB and ANI impaired memory of moderate training in a dose-dependent manner without affecting short-term memory. In contrast, memory consolidation was not impaired in the groups trained with 2.0 mA. The findings showed that: (1) inhibitors of transcription and translation in the hippocampus impair the consolidation of memory of inhibitory avoidance learning induced by moderate levels of aversive stimulation and (2) blocking of mRNA and protein synthesis does not prevent the consolidation of memory induced by relatively high levels of aversive stimulation. These findings do not support the hypothesis that gene expression and de novo protein synthesis are necessary steps for long-term memory formation as memory was not impaired if intense foot-shock was used in training.

Sexually experienced, but not naïve, female rats show a conditioned object preference (COP) for mating after a single training trial

Female rats with mating experience spend more time with the male rat, exhibit shorter contact-return latency to intromission, and display more proceptive behaviors in the male rat's compartment than during the first mating experience. The present study tested 1) whether mating induced conditioned object preference (COP) is possible with a single conditioning trial and 2) whether a preference is induced for an object associated with the first mating encounter or the fifth mating encounter in female rats. Ovariectomized, Long-Evans female rats were primed with estradiol benzoate + progesterone and either exposed to an empty paced mating chamber for 15 min (Naïve) or received a 15 intromission test of paced mating behavior (Experienced) on four separate occasions before undergoing the COP procedure. Experienced, but not Naïve, female rats developed a COP for a single mating bout, indicating that mating is highly rewarding for sexually experienced female rats. The findings raise questions about the effect of sexual experience on reward regions in the brain, the responsiveness of genital tissue, and learning mechanisms.

Impact of 1-day and 4-day MWM training techniques on oxidative and neurochemical profile in rat brain: A comparative study on learning and memory functions

Among multiple behavioral tasks used to assess memory performance, Morris water maze (MWM) is a well-known and reliable conventional behavioral task to monitor spatial memory performance in rodents. Although multiple procedures are employed by researchers for spatial learning training in MWM, but less is known about impact of these training protocol variations on oxidative and neurochemical systems. Therefore, this study aimed to examine whether variations in training protocol will influence spatial memory performance and induce changes in oxidative status and cholinergic and aminergic neurotransmission in rat brain. For this, rats were assigned to four groups; control (unexposed), 1-trial (exposed to single training trial), 1-day (exposed to four training trials for a single day) and 4-day (exposed to four training trials for four days). After conducting training, spatial reference memory performance was determined by performing retention and consolidation probe trials. Rats were then decapitated and their brain and plasma samples were collected for biochemical, oxidative and neurochemical analysis. It was found that spatial reference memory was improved following both 1-day and 4-day training protocols, however, corticosterone levels were raised extensively following 4-day training exposure compared to 1-day training protocol. Similarly, a significant improvement in redox profile and cholinergic and aminergic neurotransmitters was also observed following 1-day training procedure. Thus, 1-day training procedure can be suggested as a better procedure for assessing the spatial memory performance in rats as it has a profound impact on antioxidant status and cholinergic and aminergic neurotransmission in brain. Moreover, use of single-day training procedure can provide a rapid and effective tool for assessing spatial memory in rats compared to prolonged and complicated 4-day training protocol.

Evaluation of operant learning in young foals using target training

The primary purpose of this study was to characterize operant learning performance of young foals. For each of 26 foals, aged 6–20 weeks, learning performance was quantitatively evaluated in a single brief training trial using a standard operant conditioning task and paradigm analogous to those common to training and management of domestic horses, popularly referred to as “target training.” With no human interaction in the interim, retention of the learning was evaluated seven to 26days after the initial training trial. All 26 foals demonstrated learning in this operant paradigm. In this operant paradigm, learning was as efficient in the foals of 6 weeks to 3 months of age (n=14) as foals of 3–5 months of age (n=12). Some evidence was found for more efficient learning in female (n=13) than in male foals (n=13), as well as in certain sire lines. Seventeen of the 26 foals (65%) met the criterion for retention. Differences in the proportions of males and females and of younger and older foals that met this criterion were not significant (Fisher’s Exact Test). This work demonstrates the ability of young foals to efficiently learn and to retain an operant task.

Vibrissal paralysis produces increased corticosterone levels and impairment of spatial memory retrieval

This research was aimed at establishing how the absence of active whisking in rats affects acquisition and recovery of spatial memory. The mystacial vibrissae were irreversibly paralyzed by cutting the facial nerve’s mandibular and buccal branches bilaterally in the facial nerve lesion group (N=14); control animals were submitted to sham-surgery (N=15). Sham-operated (N=11) and facial nerve-lesioned (N=10) animals were trained (one session, eight acquisition trials) and tested 24h later in a circular Barnes maze. It was found that facial nerve lesioned-animals adequately acquired the spatial task, but had impaired recovery of it when tested 24h after training as compared to control ones. Plasma corticosterone levels were measured after memory testing in four randomly chosen animals of each trained group and after a single training trial in the maze in additional facial nerve-lesioned (N=4) and sham-operated animals (N=4). Significant differences respecting the elevation of corticosterone concentration after either a single training trial or memory testing indicated that stress response was enhanced in facial nerve-lesioned animals as compared to control ones. Increased corticosterone levels during training and testing might have elicited the observed whisker paralysis-induced spatial memory retrieval impairment.

Evaluación de la Memoria Tipo Episódica en preescolares empleando una tarea con perspectiva egocéntrica

La Memoria Tipo Episódica (MTE) implica la recuperación integrada de elementos mínimos de la memoria episódica: Qué, Dónde y Cuándo (QDC) de un evento personal. Investigaciones sugieren que una tarea adecuada para niños debe realizarse con una perspectiva egocéntrica y ocurrir en un solo ensayo, porque al realizarse con varios ensayos esto implica el uso de información semántica. Se diseñó una tarea con una perspectiva egocéntrica y realizable en un único ensayo de entrenamiento, que manipuló la magnitud de la consecuencia para dos experiencias, así como el intervalo entre entrenamiento y prueba. Niños preescolares, desde una perspectiva egocéntrica, buscaron monedas grandes o pequeñas en tres contenedores y eligieron el contenido preferido tras dos intervalos de tiempo. La elección de los participantes mostró la recuperación integrada del QDC a partir de un solo ensayo de entrenamiento. Además, las elecciones de los preescolares reflejaron sus estrategias de recuperación: la inmediatez o mayor consecuencia. La tarea se basa en las representaciones del participante más que en la capacidad de reporte verbal, lo que hace a esta preparación sensible a la cognición episódica del recuerdo de la MTE en niños.

Development of a Time-Intensity Evaluation System for Consumers: Measuring Bitterness and Retronasal Aroma of Coffee Beverages in 106 Untrained Panelists.

In order to develop products that are acceptable to consumers, it is necessary to incorporate consumers' intentions into products' characteristics. Therefore, investigation of consumers' perceptions of the taste or smell of common beverages provides information that should be useful in predicting market responses. In this study, we sought to develop a time-intensity evaluation system for consumer panels. Using our system, we performed time-intensity evaluation of flavor attributes (bitterness and retronasal aroma) that consumers perceived after swallowing a coffee beverage. Additionally, we developed quantitative evaluation methods for determining whether consumer panelists can properly perform time-intensity evaluation. In every trial, we fitted an exponential function to measured intensity data for bitterness and retronasal aroma. The correlation coefficients between measured time-intensity data and the fitted exponential curves were greater than 0.8 in about 90% of trials, indicating that we had successfully developed a time-intensity system for use with consumer panelists, even after just a single training trial using a nontrained consumer. We classified participants into two groups based on their consumption of canned coffee beverages. Significant difference was observed in only AUC of sensory modality (bitterness compared with retronasal aroma) among conventional TI parameters using two-way ANOVA. However, three-way ANOVA including a time course revealed significant difference between bitterness and retronasal aroma in the high-consumption group. Moreover, the high-consumption group more easily discriminated between bitterness and retronasal aroma than the low-consumption group. This finding implied that manufacturers should select consumer panelists who are suitable for their concepts of new products.

Dopamine in the Dorsal Hippocampus Impairs the Late Consolidation of Cocaine-Associated Memory

Cocaine is thought to be addictive because it elevates dopamine levels in the striatum, reinforcing drug-seeking habits. Cocaine also elevates dopamine levels in the hippocampus, a structure involved in contextual conditioning as well as in reward function. Hippocampal dopamine promotes the late phase of consolidation of an aversive step-down avoidance memory. Here, we examined the role of hippocampal dopamine function in the persistence of the conditioned increase in preference for a cocaine-associated compartment. Blocking dorsal hippocampal D1-type receptors (D1Rs) but not D2-type receptors (D2Rs) 12 h after a single training trial extended persistence of the normally short-lived memory; conversely, a general and a specific phospholipase C-coupled D1R agonist (but not a D2R or adenylyl cyclase-coupled D1R agonist) decreased the persistence of the normally long-lived memory established by three-trial training. These effects of D1 agents were opposite to those previously established in a step-down avoidance task, and were here also found to be opposite to those in a lithium chloride-conditioned avoidance task. After returning to normal following cocaine injection, dopamine levels in the dorsal hippocampus were found elevated again at the time when dopamine antagonists and agonists were effective: between 13 and 17 h after cocaine injection. These findings confirm that, long after the making of a cocaine-place association, hippocampal activity modulates memory consolidation for that association via a dopamine-dependent mechanism. They suggest a dynamic role for dorsal hippocampal dopamine in this late-phase memory consolidation and, unexpectedly, differential roles for late consolidation of memories for places that induce approach or withdrawal because of a drug association.

Sex- and dose-dependent effects of post-trial calcium channel blockade by magnesium chloride on memory for inhibitory avoidance conditioning

Calcium influx through voltage-dependent Ca2+ channels is critical for many neuronal processes required for learning and memory. Persistent increases in cytosolic intracellular Ca2+ concentrations in aging neurons are associated with learning impairments, while small transient subcellular changes in intracellular calcium concentrations play critical roles in neural plasticity in young neurons. In the present study, young male and female Fisher 344×Brown Norway (FBN) hybrid rats were administered different doses of magnesium chloride (0.0, 100.0, or 200.0mg/kg, i.p.) following a single inhibitory avoidance training trial. Extracellular magnesium ions can non-specifically block voltage-gated calcium channels, and/or reduce the calcium conductance gated via glutamate and serine's activation of neuronal NMDA receptors. In our study, magnesium chloride dose-dependently enhanced memory compared to controls (significantly increased latency to enter a dark compartment previously paired with an aversive stimulus) when tested 48h later as compared to controls. A leftward shift in the dose response curve for memory enhancement by magnesium chloride was observed for male compared to female rats. These findings provide further insights into calcium-dependent modulation of aversive memory, and should be considered when assessing the design of effective treatment options for both male and female patients with dementia or other memory problems.

A metaplasticity-like mechanism supports the selection of fear memories: role of protein kinase a in the amygdala.

How the brain determines which memories are selected for long-term storage is critical for a full understanding of memory. One possibility is that memories are selected based on the history of activity and current state of neurons within a given memory circuit. Many in vitro studies have demonstrated metaplasticity-like effects whereby prior neuronal activity can affect the ability of cells to express synaptic plasticity in the future; however, the significance of these findings to memory is less clear. Here we show in rats that a single pairing of a light with shock, insufficient to support either short- or long-term fear memory, primes future learning such that another trial delivered within a circumscribed time window lasting from ∼60 min to 3 d results in the formation of a long-lasting and robust fear memory. Two adequately spaced training trials support long-term fear memory only if the two trials are signaled by the same cue. Furthermore, although a single training trial does not support formation of an observable fear memory, it does result in the phosphorylation of several targets of protein kinase A (PKA) in the amygdala. Accordingly, blocking PKA signaling in the amygdala before the first training trial completely prevents the ability of that trial to facilitate the formation of long-term fear memory when a second trial is delivered 24 h later. These findings may provide insight into how memories are selected for long-term storage.

PKA increases in the olfactory bulb act as unconditioned stimuli and provide evidence for parallel memory systems: Pairing odor with increased PKA creates intermediate- and long-term, but not short-term, memories

Neonatal odor-preference memory in rat pups is a well-defined associative mammalian memory model dependent on cAMP. Previous work from this laboratory demonstrates three phases of neonatal odor-preference memory: short-term (translation-independent), intermediate-term (translation-dependent), and long-term (transcription- and translation-dependent). Here, we use neonatal odor-preference learning to explore the role of olfactory bulb PKA in these three phases of mammalian memory. PKA activity increased normally in learning animals 10 min after a single training trial. Inhibition of PKA by Rp-cAMPs blocked intermediate-term and long-term memory, with no effect on short-term memory. PKA inhibition also prevented learning-associated CREB phosphorylation, a transcription factor implicated in long-term memory. When long-term memory was rescued through increased β-adrenoceptor activation, CREB phosphorylation was restored. Intermediate-term and long-term, but not short-term odor-preference memories were generated by pairing odor with direct PKA activation using intrabulbar Sp-cAMPs, which bypasses β-adrenoceptor activation. Higher levels of Sp-cAMPs enhanced memory by extending normal 24-h retention to 48-72 h. These results suggest that increased bulbar PKA is necessary and sufficient for the induction of intermediate-term and long-term odor-preference memory, and suggest that PKA activation levels also modulate memory duration. However, short-term memory appears to use molecular mechanisms other than the PKA/CREB pathway. These mechanisms, which are also recruited by β-adrenoceptor activation, must operate in parallel with PKA activation.

Properties of Contextual Memory Formed in the Absence of αCaMKII Autophosphorylation

The alpha-isoform of calcium/calmodulin-dependent kinase II (αCaMKII) is a major synaptic kinase that undergoes autophosphorylation after NMDA receptor activation, switching the kinase into a calcium-independent activity state. This αCaMKII autophosphorylation is essential for NMDA receptor-dependent long-term potentiation (LTP), induced by a single tetanus, in hippocampal area CA1 and in neocortex. Furthermore, the αCaMKII autophosphorylation is essential for contextual long-term memory (LTM) formation after a single training trial but not after a massed training session. Here, we show that in the absence of αCaMKII autophosphorylation contextual fear conditioning is hippocampus dependent and that multi-tetanus-dependent late-LTP cannot be induced in hippocampal area CA1. Furthermore, we show that in the absence of αCaMKII autophosphorylation contextual LTM persists for 30 days, the latest time point tested. Additionally, contextual, but not cued, LTM formation in the absence of αCaMKII autophosphorylation appears to be impaired in 18 month-old mice. Taken together, our findings suggest that αCaMKII autophosphorylation-independent plasticity in the hippocampus is sufficient for contextual LTM formation and that αCaMKII autophosphorylation may be important for delaying age-related impairments in hippocampal memory formation. Furthermore, they propose that NMDA receptor-dependent LTP in hippocampal area CA1 is essential for contextual LTM formation after a single trial but not after massed training. Finally, our results challenge the proposal that NMDA receptor-dependent LTP in neocortex is required for remote contextual LTM.

A Late-Phase, Long-Term Memory Trace Forms in the γ Neurons ofDrosophilaMushroom Bodies after Olfactory Classical Conditioning

Using functional optical imaging in vivo, we demonstrate that the γ mushroom body (MB) neurons of Drosophila melanogaster respond with axonal calcium influx when odors or electric shock stimuli are presented to the fly. Pairing of odor and electric shock stimuli in a single training trial or multiple, massed training trials failed to modify the odor-evoked calcium signal when flies were tested at several different times after training. In contrast, animals that received multiple but spaced odor-shock pairings exhibited a robust increase in calcium influx into the MB axons when tested between 18 and 48 h after training. This time window for the γ neuron memory trace is displaced relative to the modifications that occur between 9 and 24 h after training in the α branch of the α/β MB neurons. The α/β and the γ neuron long-term memory traces were both blocked by expressing a repressor of the transcription factor cAMP response element-binding protein or a calcium/calmodulin-dependent kinase II hairpin RNA. These results demonstrate that behavioral long-term olfactory memory is encoded as modifications of calcium influx into distinct MB neurons during overlapping but different windows of time after training.

Extinction memory in the crab Chasmagnathus: recovery protocols and effects of multi-trial extinction training

A decline in the frequency or intensity of a conditioned behavior following the withdrawal of the reinforcement is called experimental extinction. However, the experimental manipulation necessary to trigger memory reconsolidation or extinction is to expose the animal to the conditioned stimulus in the absence of reinforcement. Recovery protocols were used to reveal which of these two processes was developed. By using the crab contextual memory model (a visual danger stimulus associated with the training context), we investigated the dynamics of extinction memory in Chasmagnathus. Here, we reveal the presence of three recovery protocols that restore the original memory: the old memory comes back 4 days after the extinction training, or when a weak training is administered later, or once the VDS is presented in a novel context 24 h after the extinction session. Another objective was to evaluate whether the administration of multi-trial extinction training could trigger an extinction memory in Chasmagnathus. The results evince that the extinction memory appears only when the total re-exposure time is around 90 min independently of the number of trials employed to accumulate it. Thus, it is feasible that the mechanisms described for the case of the extinction memory acquired through a single training trial are valid for multi-trial extinction protocols. Finally, these results are in agreement with those reports obtained with models phylogenetically far apart from the crab. Behind this attempt is the idea that in the domain of studies on memory, some principles of behavior organization and basic mechanisms have universal validity.

Acute cognitive impairment after lateral fluid percussion brain injury recovers by 1 month: Evaluation by conditioned fear response

Conditioned fear associates a contextual environment and cue stimulus to a foot shock in a single training trial, where fear expressed to the trained context or cue indicates cognitive performance. Lesion, aspiration or inactivation of the hippocampus and amygdala impair conditioned fear to the trained context and cue, respectively. Moreover, only bilateral experimental manipulations, in contrast to unilateral, abolish cognitive performance. In a model of unilateral brain injury, we sought to test whether a single lateral fluid percussion brain injury impairs cognitive performance in conditioned fear. Brain-injured mice were evaluated for anterograde cognitive deficits, with the hypothesis that acute injury-induced impairments improve over time. Male C57BL/6J mice were brain-injured, trained at 5 or 27 days post-injury, and tested 48 h later for recall of the association between the conditioned stimuli (trained context or cue) and the unconditioned stimulus (foot shock) by quantifying fear-associated freezing behavior. A significant anterograde hippocampal-dependent cognitive deficit was observed at 7 days in brain-injured compared to sham. Cued fear conditioning could not detect amygdala-dependent cognitive deficits after injury and stereological estimation of amygdala neuron number corroborated this finding. The absence of injury-related freezing in a novel context substantiated injury-induced hippocampal-dependent cognitive dysfunction, rather than generalized fear. Variations in the training and testing paradigms demonstrated a cognitive deficit in consolidation, rather than acquisition or recall. By 1-month post-injury, cognitive function recovered in brain-injured mice. Hence, the acute injury-induced cognitive impairment may persist while transient pathophysiological sequelae are underway, and improve as global dysfunction subsides.

Aversive Learning under Different Training Conditions: Effects of NMDA Receptor Blockade in Area CA1 of the Hippocampus

Adult male Wistar rats were given either a single training trial or one training trial per day during 3 days followed by a retention test trial in an inhibitory avoidance (IA) task. In animals given a single training trial, pretraining, but not pretest bilateral infusion of the NMDA glutamate receptor antagonist d,l-2-amino-5-phosphonopentanoic acid (AP5) (5.0 microg) into the CA1 hippocampal area blocked IA retention. In animals given three training trials, infusions of AP5 given prior to each of the three training trials severely impaired, but did not block retention. The results indicate that NMDA receptors in the hippocampus are involved in the formation, but not in expression, of aversive memory. In addition, rats given repeated training were able to show a mild improvement of performance across training trials, possibly through mechanisms that do not depend on NMDA receptor activation in the dorsal hippocampus.

Midazolam disrupts fear memory reconsolidation

The current research examines the influence of midazolam (MDZ) on memory reconsolidation using a contextual fear paradigm in rats, based on three context-shock training trials (0.7 mA, 3 s). First, we evaluate the effect of MDZ (1 mg/kg, i.p.) injected shortly after the training procedure. Second, we examined the influence of MDZ after a brief exposure (90 s) either in the training context (reactivation procedure) or in a neutral environment (no reactivation procedure) and one day later, freezing behavior was scored when rats were re-exposed to the training environment. Third, we investigate both the effect of MDZ administered at different times following reactivation on fear memory and the persistence of such effect 10 days after reactivation. Finally, we test whether the MDZ effect could be reverted by a single weak training trial (0.2 mA, 3 s) or by the presentation of the same unconditioned stimulus in the absence of the conditioned stimulus as a reminder which proves to induce significant freezing in rats not previously trained. Results show that MDZ interferes with the formation of a contextual fear memory only when administered after the reactivation procedure but not after the training procedure. This interference was effective up to 60 min after reactivation and not at a later time. No spontaneous recovery of freezing behavior was observed 11 days after MDZ injection which was not reverted by a weak training trial and by the unconditioned stimulus alone. All these data support the idea that stimulating GABA A receptor sites via MDZ selectively disrupts the reconsolidation process of a contextual fear memory.