PKG-mediated MAPK signaling is necessary for long-term operant memory in Aplysia

Signaling pathways necessary for memory formation, such as the mitogen-activated protein kinase (MAPK) pathway, appear highly conserved across species and paradigms. Learning that food is inedible (LFI) represents a robust form of associative, operant learning that induces short- (STM) and long-term memory (LTM) in Aplysia. We investigated the role of MAPK signaling in LFI memory in vivo. Inhibition of MAPK activation in animals prior to training blocked STM and LTM. Discontinuing MAPK signaling immediately after training inhibited LTM with no impact on STM. Therefore, MAPK signaling appears necessary early in memory formation for STM and LTM, with prolonged MAPK activity required for LTM. We found that LFI training significantly increased phospho-MAPK levels in the buccal ganglia. Increased MAPK activation was apparent immediately after training with greater than basal levels persisting for 2 h. We examined the mechanisms underlying training-induced MAPK activation and found that PKG activity was necessary for the prolonged phase of MAPK activation, but not for the early MAPK phase required for STM. Furthermore, we found that neither the immediate nor the prolonged phase of MAPK activation was dependent upon nitric oxide (NO) signaling, although expression of memory was dependent on NO as previously reported. These studies emphasize the role of MAPK and PKG in negatively reinforced operant memory and demonstrate a role for PKG-dependent MAPK signaling in invertebrate associative memory.     

Internal stimuli enhance feeding behavior in the mollusc Aplysia

The hypothesis that subsatiating levels of internal food stimuli can arouse and potentiate feeding behavior was examined in the mollusc Aplysia californica. Animals were fed a small quantity of seaweed and their latencies to show biting responses were determined after food arousal was permitted to partially decay. Control animals were stimulated with food, but were not permitted to ingest it, or were fed nonnutritive glass-fiber filter paper. Compared to controls, animals that were fed showed significantly shorter latencies to respond when tested up to 80 min after previous exposure to food. These results indicate that internal stimuli can function like external stimuli to enhance responsiveness to food and suggest the hypothesis that satiation may be viewed as an interactive process involving the interplay of excitatory and inhibitory influences arising from the alimentary system.     

Effects of food deprivation upon behavioral patterns and time budgeting of Aplysia fasciata

Effects of food deprivation upon time devoted to different behaviors in Aplysia fasciata were investigated. Time invested in mating, egg-laying, swimming, and movement in place was increased, and time spent immobile and crawling was decreased. Time of occurrence of behaviors was highly synchronized, but differed from that seen when food was available. Changes in behavior seem to be adaptive to conditions in which food is scarce: increased motility may lead to finding sites where food is present, while increased egg-laying and mating may ensure survival of progeny. Behavioral changes can be readily accounted for by removal of postingestive stimuli signaling satiation.     

The effect of asymmetrical sample training on retention functions for hedonic samples in rats

Rats were trained in a symbolic delayed matching-to-sample task to discriminate sample stimuli that consisted of the presence of food or the absence of food. Asymmetrical sample training was provided in which one group was initially trained with only the food sample and the other group was initially trained with only the no-food sample. In addition, within each group half of the rats were trained with an illuminated intertrial interval (ITI) and the remaining rats with a dark ITI. While the retention functions did not differ as a function of which sample was trained first, they did differ as a function of the similarity in the illumination conditions during the ITI and the delay interval. Symmetrical retention functions were obtained when the lighting conditions were similar and slightly asymmetrical retention functions were obtained when the lighting conditions were dissimilar. Probe tests confirmed that features of the no-food sample were attended to and used to generate a memory representation for the no-food sample. The results are not consistent with the hypothesis that asymmetrical sample training encourages coding of the sample introduced initially and default responding to the subsequently introduced sample. Rats generate memory representations for both samples when asymmetrical sample training is given with hedonic samples.     

A brief retraining regulates the persistence and lability of a long-term memory

An experience extending the persistence of a memory after training Aplysia californica with inedible food also allows a consolidated memory to become sensitive to consolidation blockers. Long-term (24 h) memory is initiated by 5 min of training and is dependent on protein synthesis during the first few hours after training. By contrast, a more persistent (48 h) memory is dependent on a longer training session and on a later round of protein synthesis. When presented 24 h after training, a 3-min training that produces no memory alone can cause a memory that would have persisted for only 24 h to persist for 48 h. After a 48 h memory has been consolidated, 3 min of training also makes the memory sensitive to a protein-synthesis inhibitor. These findings suggest that a function of allowing a consolidated memory to become sensitive to blockers of protein synthesis may be to allow the memory to become more persistent.Long-term memory of an experience is dependent on a consolidation process that follows the experience. Before the memory is consolidated it is labile and can be disrupted (Lechner et al. 1999; Dudai 2004; Alberini and Taubenfeld 2008), particularly by blocking mRNA and protein synthesis (Alberini and Taubenfeld 2008; Klann and Sweatt 2008), which are needed to produce the changes in synaptic structure and function that underlie long-term memory (Sigurdsson et al. 2007; Bailey and Kandel 2008; De Roo et al. 2008). After long-term memory is established by the initial stages of consolidation, later rounds of consolidation may be needed to extend the persistence of the memory (Wittenberg and Tsien 2002; Bekinschtein et al. 2007). Consolidated memories may also be modified when they are retrieved (Dudai 2006; Alberini and Taubenfeld 2008). Retrieving a memory by exposure to the conditioned stimulus, or by re-experiencing aspects of the original training, can destabilize it and initiate an additional protein-synthesis-dependent process of memory stabilization (Nader 2003; Dudai 2006; Alberini and Taubenfeld 2008). The aim of this communication is to explore whether an experience that alone does not cause long-term memory can make a memory more persistent and can also destabilize a consolidated memory and makes it labile.Training Aplysia californica with inedible food until they stop responding to the food initiates long-term memory that can be measured as a reduction in the time to stop responding. Long-term memory is present 1, 2, and 7 d after a single training session (Schwarz et al. 1991). We examined some of the parameters of training that lead to persistence of memory by determining whether shorter training sessions also produce a persistent long-term memory.As in previous studies (Schwarz et al. 1991; Botzer et al. 1998; Lyons et al. 2005), Aplysia californica were trained with inedible food, the seaweed Ulva wrapped in plastic net. This food induced biting, leading to food entering the mouth. Animals then attempted to swallow the food. The netted food cannot be swallowed and it became lodged in the buccal cavity, producing repetitive failed swallowing responses. Food eventually left the buccal cavity. The experimenter continued to hold the food against the lips, inducing further biting responses, entries into the mouth, and failed swallows. As training proceeded many bites failed to cause entry of food into the mouth. When food did enter the mouth it stayed within for progressively shorter periods, eliciting fewer attempted swallows. In all animals, food was in the mouth eliciting failed attempts to swallow for at least 100 sec of the initial training, since previous experience showed that such animals almost always show long-term memory. Animals in which food was not in the mouth for 100 sec during training were discarded. A full training session until animals stop responding to food requires 10–25 min of training (Fig. 1). Such a training session caused long-term memory measured after 24 h or after 48 h (Fig. 1A). Memory was measured by comparing the time to stop responding to inedible food during training to the time to stop responding when animals were tested 24 or 48 h later, in a procedure identical to that during training. All tests of memory were performed using a blind procedure in which the experimenter was unaware of the previous training procedure.Open in a separate windowFigure 1.Memory 24 and 48 h after different training procedures. The figure shows the time to stop responding in a group of naïve animals trained with a full training session (training continued until animals stop responding to food), as well as the time to stop responding on memory trials 24 and 48 h later (N = 38 naïve animals [naïve animals were run as controls for the various other groups and data from the naïve animals were then combined. There was no significant difference between the various groups of naïve animals: P = 0.405, F_(5,32) = 1.053]); (A) N = 13 animals tested 24 h after a full training; N = 6 animals tested 48 h after a full training; (B) N = 9 animals tested 24 h after a 5-min training; N = 17 animals tested 48 h after a 5-min training; (C) N = 14 animals tested 24 h after a 3-min training; (D) N = 7 animals tested 48 h after a 5-min training, plus an additional 3-min training 24 h later. A one-way analysis of variance showed significant differences between the seven groups shown (P < 0.001, F_(6,97) = 11.95). A post-hoc test (Student-Newman-Keuls, α = 0.05) showed that there were no significant differences between the time to stop in naïve animals and in animals tested 48 h after a 5-min training, or in animals tested 24 h after a 3-min training, indicating that these treatments did not cause memory. By contrast, the times to stop in these three groups were significantly different from that in the other four groups, which were not significantly different from one another. These findings indicate significant memory 24 or 48 h after a full training, as well as 24 h after 5 min of training, and 48 h after 5 min of training with 3 min of reminder training, with no differences in memory after these 4 treatments. Standard errors are shown.We examined whether abbreviated training also caused long-term memory. First, we examined long-term memory when training is stopped after 5 min. During the first 5 min of a full training session, food is in the mouth, and animals are attempting to swallow for a mean of 70% of the time in the mouth during a full training session. Attempts to swallow are an integral part of the training (Katzoff et al. 2006). We confirmed an earlier finding (Botzer et al. 1998) that a 5-min training produced long-term memory measured 24 h after the training (Fig. 1B). However, we have now found that training animals for only 5 min did not cause long-term memory measured 48 h after training (Fig. 1B). A training session that was stopped after 3 min did not give rise to long-term memory measured at 24 h (Fig. 1C), indicating that the last 2 min of a 5-min training are necessary for the production of 24 h memory. These findings allowed us to examine the possible effects of an experience which itself does not cause memory, 3 min of training on memory persistence and memory lability.A 5-min training session gives rise to memory after 24 h, but not after 48 h, whereas additional training gives rise to 48 h memory. Must the additional training take the form of continuing to train animals during the initial training session, or can an additional 3 min of training that itself does not cause 24 h memory enhance the effect of a 5 min of training? To test the ability of a 3-min training to enhance memory, animals were trained with either a full training session, or with a training session that was abbreviated after 5 min. One group of animals trained for 5 min received an additional 3-min training 24 h after the initial training, whereas another group did not. Memory was tested 24 h later, 48 h after the initial training. Animals receiving a full training, and animals receiving a 5-min training plus a reminder consisting of a 3-min additional training, displayed 48 h memory (Fig. 1D; for a fuller presentation of this experiment, see Supplemental material), whereas animals receiving only a 5-min training, with no additional training, showed no 48 h memory. These data show that a 3-min training, which is itself ineffective in producing memory 24 h later, can lead to significant memory when it follows a 5-min training, which itself would not produce 48 h memory.In other learning tasks it has been shown that long-term memory is not a unitary process. An earlier round of protein synthesis is necessary for 24 h memory, but a more persistent memory (>24 h), and the synaptic plasticity underlying it, are dependent on later rounds of protein synthesis (Giustetto et al. 2003; Bekinschtein et al. 2007; Miniaci et al. 2008). We therefore examined whether 24 and 48 h memory differ in their dependence on protein synthesis adjacent to training, or on protein synthesis 6 h after training. The protein-synthesis inhibitor anisomycin was injected into the hemolymph either 10 min before training or 6 h after training. Animals were injected with a 1 cc solution of anisomycin at a concentration that caused a 10 µM concentration within the animals. This concentration blocks protein synthesis in ganglia (Schwartz et al. 1971). Controls were injected with 1 cc of artificial seawater (ASW–NaCl 460 mM, KCl 10 mM, CaCl₂ 11 mM, MgCl₂ 55 mM, and NaHCO₃ 5 mM). Treatment with anisomycin just before training blocked 24 h memory, but anisomycin treatment 6 h after training did not prevent the appearance of 24 h memory, indicating that 24 h memory is consolidated within the first 6 h after training (note that Fig. 2A shows the percent change in time to stop responding during the test of memory, with respect to the time to stop during training). By contrast, 48 h memory was blocked by anisomycin treatment 6 h after training (Fig. 2B), whereas treatment with ASW did not block 48 h memory.Open in a separate windowFigure 2.Protein synthesis dependence of 24 and 48 h memory. Data shows the memory test 24 or 48 h after training as a percent change [−([train-test]/train) × 100] from the initial training session. All tests of memory were after a full training session until animals stopped responding to the food. Memory was tested via two-tailed paired t-tests, in which the time to stop responding in a given animal during the initial training was compared with the time to stop responding 24 or 48 h later, when animals were tested. A significant decrease in the time to stop responding was used as an indicator of memory. Treatments showing such a decrease are marked (*). Standard errors are shown. (A) Anisomycin treatment blocks 24 h memory when applied just before training, but not when applied 6 h after training. Control animals (N = 8) not treated with anisomycin displayed significant 24 h memory (P < 0.001, t₍₇₎ = 10.15). Anisomycin applied 10 min before the training (N = 9) blocked memory, as shown by a lack of significant decrease in the time to stop between the initial training and the test after 24 h (P = 0.96, t₍₈₎ = 0.05). By contrast, application of anisomycin 6 h after the initial training (N = 7) did not block memory measured 24 h after training, as shown by significant savings after 24 h (P = 0.05, t₍₆₎ = 2.45). (B) Forty-eight hour memory is dependent on a later stage of protein synthesis. Anisomycin (N = 26), but not ASW (N = 5) applied 6 h after training blocks 48 h memory, as shown by significant memory after treatment with ASW (P = 0.007, t₍₄₎ = 5.08) but not with anisomycin (P = 0.49, t₍₂₅₎ = 0.69). However, a 3-min reminder training 24 h after the training (N = 12) rescues 48 h memory after it was blocked by anisomycin treatment 6 h after training (P = 0.01, t₍₁₁₎ = 2.93).Memory 48 h after training is dependent on protein synthesis 6 h after training. As shown above, 3 min of training can establish 48 h memory after a 5-min training that alone is too brief to establish 48 h memory. Can 3 min of training also establish 48 h memory after it has been blocked by inhibiting protein synthesis at 6 h? To test this possibility, animals were trained with inedible food until they stopped responding, and were then treated with either ASW or anisomycin 6 h later. One group of animals also received 3 min of training with inedible food 24 h after the initial training. When tested 48 h after training, memory was present in animals treated with ASW and in animals that had received the additional 3-min training, but not in animals receiving only the anisomycin treatment (Fig. 2B). This finding indicates that a 3-min training that is effective in causing 48 h memory after 5 min of training can also rescue 48 h memory after it has been blocked by anisomycin.If 3 min of training saves a blocked 48 h memory, it could also amplify an already-formed 48 h memory. We tested the effect on 48 h memory of a 3-min training 24 h after a full training session, which alone produces 48 h memory. There was no significant difference in memory between animals tested 24 h after training, and animals receiving a 3-min retraining, and then tested 48 h after training, indicating that the 3-min training did not affect the already consolidated 48 h memory (Fig. 3, cf. column 1 and column 3).Open in a separate windowFigure 3.Brief training makes memory labile. Twenty-four hours after training with inedible food, animals were treated with anisomycin, or with ASW. The animals treated with ASW (N = 9), as well as one of the two groups treated with anisomycin (N = 9), were also given a 3-min training session 10 min later. Memory was then tested again 24 h later, 48 h after the training, by comparing the time to stop measured 48 h after training with the time to stop measured during the original training session, using a two-tailed paired t-test. A significant decrease in the time to stop responding was used as an indicator of memory. Data are also shown for 24 h memory after a full training session, to allow a test of whether a 3-min retraining at 24 h improves memory tested at 48 h. Treatment with anisomycin alone (N = 8) did not block 48 h memory (P = 0.002, t₍₇₎ = 4.66). The 3-min reminder training plus treatment with ASW also did not block 48 h memory (P = 0.001, t₍₈₎ = 5.02). By contrast, when the 3-min reminder training was preceded by anisomycin treatment, there was no significant difference between training and the 48 h test (P = 0.27, t₍₈₎ = 1.18), indicating that the 3-min reminder allowed the anisomycin to block 48 h memory. Note that there is also no significant difference in savings after the 3-min training following ASW treatment and 24 h memory following training (P = 0.13, t₍₁₀₎ = 1.63), indicating that the 3-min training on day 2 alone did not affect memory.The 3-min training does not affect an already-formed 48 h memory, but does cause effects 24 h later if the memory is not fully consolidated. Formation of a long-term memory requires protein synthesis, and recalling a consolidated memory can make a memory sensitive to inhibitors of protein synthesis (Nader et al. 2000). Does a 3-min training 24 h after an initial full training initiate a renewed dependence of 48 h memory on protein synthesis? Animals were trained to criterion and then subjected to either anisomycin, or ASW, 24 h later. Both groups then received a 3-min reminder training 10 min later. Memory was then assessed 24 h later (48 h after initial training). There was significant memory 48 h after the training after treatment with ASW, but not after treatment with anisomycin, indicating that the 3-min reminder training restored the ability of anisomycin to block memory (Fig. 3).Our data have shown that 3 min of training with inedible food alone does not produce long-term memory (Fig. 1), and does not amplify an already established 48 h memory (Fig. 3). Nonetheless, 3 min of training induces a protein-synthesis-dependent state in which memory can be modified (Figs. 2, ​,3).3). After a previous training leading to only 24 h memory (Fig. 1), or after a treatment blocking 48 h memory, the 3-min training makes the memory more persistent (Fig. 2). In addition, after a 48 h memory has already been consolidated, 3 min of training makes the memory labile, so that it can be blocked by inhibitors of protein synthesis (Fig. 3).Reconsolidation is a process by which a consolidated memory returns to a protein-synthesis-dependent labile state by retrieving the memory (e.g., Nader et al. 2000; Sangha et al. 2003; Kemenes et al. 2006). In learning that food is inedible 3 min of training makes the memory labile. There has been much speculation on the possible function of memory reconsolidation. It has been suggested that the destabilization of memory by retrieval could function as a means to update and modify the magnitude and the persistence of the original memory trace (Dudai 2002, 2006). There is good evidence that reconsolidation can update (Rodriguez-Ortiz et al. 2005; Morris et al. 2006), strengthen (Frenkel et al. 2005; Tronson et al. 2006; Lee 2008), or modify (Rossato et al. 2006) memories, as well as block them (Nader et al. 2000), but the possible role of reconsolidation in regulating memory persistence has not been examined. Indeed, many of the previous studies on reconsolidation used a conditioned stimulus (CS) alone as a reminder, and thereby caused extinction along with reconsolidation (Eisenberg et al. 2003; Pedreira and Maldonado 2003), rather than causing a strengthening of memory. Previous studies have indicated that after memory is initially consolidated, its persistence is dependent on successive waves of protein synthesis (Giustetto et al. 2003; Bekinschtein et al. 2007; Miniaci et al. 2008) and that reactivation of a memory improves its persistence (Spear 1973). Later waves of protein synthesis affecting persistent memories could be modulated or modified by retrieving a memory.We have taken advantage of a memory task affecting Aplysia feeding to examine the possibility that memory retrieval via a brief additional training affects later waves of protein synthesis, and thereby affects the persistence of a memory. We found that pairing a brief additional training with block of protein synthesis blocks memory (Fig. 3). We also found that a more persistent memory measured at 48 h is dependent on a longer training session (Fig. 1), and a later wave of protein synthesis (Fig. 2) than is 24 h memory. Retrieval of memory by a 3-min retraining can establish 48 h memory even when the latter portion of training is absent (Fig. 1), or when the later wave of protein synthesis is blocked (Fig. 2). This finding suggests that the initial experience required to establish 48 h memory, as well as the wave of protein synthesis elicited by this experience, can be deferred. A later experience, and a later wave of protein synthesis, can substitute for the absence of experience in the initial training, or for the blocked wave of protein synthesis. In addition to creating a more persistent memory, a 3-min retraining also makes a consolidated memory sensitive to blocking by inhibitors of protein synthesis (Fig. 3). Thus, a function of experiences that permit reconsolidation may also be to extend the persistence of a memory.In our study we cannot exclude the possibility that the 3-min retraining does more than just retrieve memory, and thereby make it labile. The increased persistence of memory could be caused by processes initiated by the 3-min training, such as increased consolidation, that are not related to the ability of the added training to make the memory labile. Indeed, previous studies have suggested that memory consolidation and reconsolidation may utilize different molecular mechanisms (Taubenfeld et al. 2001; Lee et al. 2004; Alberini 2005). The different molecular mechanisms activated by consolidation or reconsolidation have been used to classify the process by which retrieval affects memory, although use of the molecular markers to classify a behavioral process is controversial (Nader et al. 2005). Such studies have shown that changing a memory when it is retrieved sometimes utilizes a molecular mechanism for consolidation (Tronel et al. 2005), and sometimes for reconsolidation (Lee 2008). In the learning task utilized we have not examined separate molecular markers for consolidation and reconsolidation, raising the possibility that a 3-min retraining affects persistence by activating molecular mechanisms that are specifically associated with a separate round of consolidation, rather than activating reconsolidation.Memory reconsolidation also affects a number of other molluscan learning paradigms (Sekiguchi et al. 1997; Sangha et al. 2003; Gainutdinova et al. 2005; Kemenes et al. 2006). In some, the memory becomes labile when animals are exposed to a CS alone, which does not itself create memory, whereas in others the memory becomes labile only when the CS is paired with an unconditioned stimulus (US), as in the original training (see Eisenhardt and Stollhoff 2008). In our experiments, memory was made labile by a second training session, which was shorter than that needed to itself create memory. It will be of interest to determine whether components of the experience alone in which animals learn that food is inedible do not give rise to memory, such as lip stimulation alone (Schwarz et al. 1988), can also give rise to reconsolidation, although they would not cause a more persistent memory.Where in the Aplysia nervous system are molecular changes related to reconsolidation and the persistence of memory localized? The earliest molecular changes leading to long-term memory that a food is inedible (e.g., increased expression of C/EBP and sensorin), are localized to the buccal ganglia, specifically to a population of mechanoafferents (Levitan et al. 2008a,b). It is possible that the later molecular changes leading to a persistent memory are also localized to these neurons, similar to the localization to the same sensory neurons of earlier and later molecular events leading to long-term facilitation underlying sensitization in Aplysia (Miniaci et al. 2008). However, it is also possible that the later phases of consolidation, and reconsolidation, may arise by molecular changes localized to other sites with the Aplysia nervous system, similar to the movement of earlier and later phases on long-term declarative memories from the hippocampus to the neocortex (Squire and Kandel 1999). Future studies will need to examine these points.     

Effects of food and mates on time budget in Aplysia fasciata: integration of feeding, reproduction, and locomotion

This study examines the time budgeted to different behaviors in Aplysia fasciata under three conditions: (1) animals have constant access to food and mates: (2) there is access to food, but not to mates; (3) neither food nor mates are present. The data suggest a number of rules underlying behavioral integration: (1) Feeding, reproductive behaviors, and activity seem to be natural categories for behavioral choice. Feeding and reproductive behaviors are controlled in tandem by a common arousal mechanism, while time left over after animals feed and reproduce is distributed in a fixed ratio between locomotion (crawling and swimming) and inactivity (immobility and movement in place). (2) Relative distribution between different forms of locomotion and inactivity is modified by changes in motivational state. More time is spent swimming than crawling when feeding and/or mating is prevented, while more time is spent moving in place than immobile when the arousal level is increased. (3) Feedback control of feeding and reproduction is asymmetric. Satiation of feeding inhibits the common arousal. In the absence of food, time spent on reproductive behaviors increases due to disinhibition of the common arousal. By contrast, positive feedback arising from sexual behavior excites the common arousal. When mating is prevented by removing potential mates, time spent feeding decreases. (4) Generally, animals choose between performing the three main categories of behavior. Although Aplysia simultaneously can feed and mate, or locomote and mate, they do so infrequently. By contrast, different types of reproductive behaviors (male mating, female mating, egg-laying) are commonly performed simultaneously.     

Multiple Memory Processes Following Training That a Food Is Inedible in Aplysia

In many organisms, memory after training can be separated into a number of processes. We now report that separable memory processes are also initiated by a training procedure affecting Aplysia feeding behavior, a model system for examining the neural mechanisms underlying the regulation of a complex behavior. Four distinct memory process were identified: (1) a very short-term memory that declines within 15 min, (2) a short-term memory that persists for 0.5–1.0 hr, (3) an intermediate-term memory, observed 4 hr after training, and (4) a long-term memory that is seen only after a 12- to 24-hr delay. The four memory processes can be distinguished by the different training procedures that are required to elicit them. A single 5-min training session is sufficient to elicit the very short-term memory. However, a longer training session that continues until the animal stops responding to food is needed to elicit short-term memory. Intermediate-term memory is observed only after a spaced training procedure (three 5-min training sessions separated by 30-min intervals). A single 5-min training session that does not cause either short-term or intermediate-term memory is sufficient to induce long-term memory, indicating that short- and long-term memory are independent, parallel processes. Short- and long-term memory can also be separated by the effects of a post-training experience. Long-term, but not short-term, memory can be attenuated by cooling animals immediately after training. Cooling before the training does not affect either the training or the subsequent short- or long-term memory.     

Human Experience Modeler: context-driven cognitive retraining to facilitate transfer of learning.

We describe a cognitive rehabilitation mixed-reality system that allows therapists to explore natural cuing, contextualization, and theoretical aspects of cognitive retraining, including transfer of training. The Human Experience Modeler (HEM) mixed-reality environment allows for a contextualized learning experience with the advantages of controlled stimuli, experience capture and feedback that would not be feasible in a traditional rehabilitation setting. A pilot study for testing the integrated components of the HEM is discussed where the participant presents with working memory impairments due to an aneurysm.     

Context learning and the effect of context on memory retrieval in Lymnaea

Aerial respiratory behavior in Lymnaea was operantly conditioned so that the animals perform aerial respiration significantly less often. Using the standard training procedure (pond water made hypoxic by bubbling N₂ through it) both food-deprived and fed animals learned and exhibited long-term memory (LTM). However, food-deprived animals exhibited neither learning nor memory when trained under a condition in which the hypoxic pond water also contained a food odorant (carrot, the food-odorant procedure). Fed animals, however, learned and exhibited LTM with the food-odorant procedure. Thus, the presence of the food odorant per se did not prevent learning or the establishment of LTM. Further experimentation, however, revealed that the ability of the snails to have recall (i.e., memory) for the learned behavior was dependent on the context in which memory was tested. That is, if animals were trained with the food-odorant procedure they could only exhibit recall if tested in the food-odorant context and vice versa with the standard training procedure. Thus, although fed animals could learn and show LTM with either training and testing procedure, LTM could only be seen when they were tested in the context in which they were trained.     

PKA and PKC are required for long-term but not short-term in vivo operant memory in Aplysia

We investigated the involvement of PKA and PKC signaling in a negatively reinforced operant learning paradigm in Aplysia, learning that food is inedible (LFI). In vivo injection of PKA or PKC inhibitors blocked long-term LFI memory formation. Moreover, a persistent phase of PKA activity, although not PKC activity, was necessary for long-term memory. Surprisingly, neither PKA nor PKC activity was required for associative short-term LFI memory. Additionally, PKA and PKC were not required for the retrieval of short- or long-term memory (STM and LTM, respectively). These studies have identified key differences between the mechanisms underlying nonassociative sensitization, operant reward learning, and LFI memory in Aplysia.     

Common regulation of feeding and mating in Aplysia fasciata: pheromones released by mating and by egg cordons increase feeding behavior.

We examined whether pheromones released by reproductive behaviors (mating and egg-laying) affect feeding behavior. A preliminary experiment demonstrated that the quantity of food eaten can be used to measure the effects of pheromones on feeding. Using this measure, we then showed that Aplysia that were prevented from mating, but that were in the same aquarium as mating conspecifics, eat more food than do Aplysia in a medium lacking mating animals. Mating and feeding were not temporally correlated, indicating that pheromones released by mating probably do not initiate feeding, but rather modulate feeding after it has begun. Aplysia that were in the same aquarium as freshly deposited egg cordons also ate more than did animals in a medium lacking eggs.     

The fate of redundant cues during blocking and a simple discrimination

In each of three experiments animals received blocking, A+ AX+, in which food was always presented after one stimulus, A, that was occasionally accompanied by another stimulus, X. They also received a simple discrimination, AX+ BX-, in which the presence and absence of food was signaled by two compounds that contained one unique cue, A or B, and one common cue, X. In each of these designs, X can be said to be redundant relative to A as a signal for food. Test trials at the end of training revealed that responding during X was stronger after blocking than after the simple discrimination. These results contradict predictions from theories of learning that assume changes in associative strength of a stimulus are determined by a global error term based on the outcome predicted by all the stimuli that are present for a conditioning trial. The results are interpreted, instead, by assuming either that animals store a memory of every trial to which they have been exposed, or that learning is governed by an error term based on the significance of individual stimuli.     

Latent memory for sensitization in Aplysia

In the analysis of memory it is commonly observed that, even after a memory is apparently forgotten, its latent presence can still be revealed in a subsequent learning task. Although well established on a behavioral level, the mechanisms underlying latent memory are not well understood. To begin to explore these mechanisms, we have used Aplysia, a model system that permits the simultaneous study of memory at the behavioral, cellular, and molecular levels. We first demonstrate that robust latent memory is induced by long-term sensitization training of the tail-elicited siphon withdrawal reflex. It is revealed by its ability to facilitate the subsequent induction of three mechanistically distinct temporal domains of sensitization memory: short-term, intermediate-term, and long-term memory. Under our training conditions, the latent memory persists for at least 2 d following the decay of original memory expression but appears to be gone by 4 d. Interestingly, we also find that latent memory is induced even in the absence of overt memory for the original training. These findings now permit the analysis of the cellular and molecular architecture of a common feature of learning and memory.     

Animals may not be stuck in time

Humans have the ability to mentally travel forward and back in time. They can retrieve vivid memories of past events (episodic memories) and can imagine the future (planning). Although it has been suggested that this is a uniquely human ability, the evidence for subjective time travel in humans is typically based on verbal report and elaboration. Such evidence cannot be obtained from animals. However, we may have indirect evidence for episodic memory and planning. For example, we can show that animals can ‘report’ about their recent past experience when they are unexpectedly asked to do so—performance that is analogous to episodic memory. We can also show that animals use the anticipation of a future event as the basis for a present action—analogous to planning. Thus, we have suggestive evidence that animals may not be stuck in time.     

Reactivation-dependent amnesia for appetitive memories is determined by the contingency of stimulus presentation

Previously acquired aversive and appetitive memories are not stable and permanent. The reactivation of such memories by re-exposure to training stimuli renders them vulnerable to disruption by amnestic agents such as the noncompetitive N-methyl-D-aspartate receptor antagonist (+)-5-methyl-10,11-dihydro-SH-dibenzo{a,d}cyclohepten-5,10-imine maleate (MK-801). However, relatively little is known about the parameters that influence the reactivation process. Here, we show that the method of stimulus presentation during memory reactivation is of great importance. Male Lister Hooded rats were trained to acquire a lever press response that delivered a sucrose reward paired with a light conditioned stimulus (CS). The CS-sucrose association was then reactivated through re-exposure to the CS, either contingently upon the lever press response, or noncontingently in the absence of instrumental responding. Systemic administration of MK-801 (0.1 mg/kg) at the time of memory reactivation resulted in amnesia, and hence a reduction in subsequent sucrose seeking induced by, and dependent upon, presentation of the CS, only when the memory was reactivated contingently. Therefore, stimuli may have to be presented in the same manner at memory reactivation as during learning in order to render a previously acquired memory vulnerable to disruption. These results have important implications for the potential translational use of glutamatergic treatments in conjunction with targeted memory reactivation.     

Associative memory in three aplysiids: correlation with heterosynaptic modulation

Much recent research on mechanisms of learning and memory focuses on the role of heterosynaptic neuromodulatory signaling. Such neuromodulation appears to stabilize Hebbian synaptic changes underlying associative learning, thereby extending memory. Previous comparisons of three related sea-hares (Mollusca, Opisthobranchia) uncovered interspecific variation in neuromodulatory signaling: strong in Aplysia californica, immeasureable in Dolabrifera dolabrifera, and intermediate in Phyllaplysia taylori. The present study addressed whether this interspecific variation in neuromodulation is correlated with memory of associative (classical conditioning) learning. We differentially conditioned the tail-mantle withdrawal reflex of each of the three species: Mild touch to one side of the tail was paired with a noxious electrical stimulus to the neck. Mild touch to the other side served as an internal control. Post-training reflex amplitudes were tested 15-30 min after training and compared with pre-test amplitudes. All three species showed conditioning: training increased the paired reflex more than the unpaired reflex. However, the temporal pattern of conditioning varied between species. Aplysia showed modest conditioning that grew across the post-test period. Dolabrifera showed distinctly short-lived conditioning, present only on the first post-test. The time course of memory in Phyllaplysia was intermediate, although not statistically distinguishable from the other two species. Taken together, these experiments suggest that evolutionary changes in nonassociative heterosynaptic modulation may contribute to evolutionary changes in the stability of the memory of classical conditioning.     

Rhesus monkeys (<Emphasis Type="Italic">Macaca mulatta</Emphasis>) show robust evidence for memory awareness across multiple generalization tests

The possibility that memory awareness occurs in nonhuman animals has been evaluated by providing opportunity to decline memory tests. Current evidence suggests that rhesus monkeys (Macaca mulatta) selectively decline tests when memory is weak (Hampton in Proc Natl Acad Sci USA 98:5359–5362, 2001; Smith et al. in Behav Brain Sci 26:317–374, 2003). However, much of the existing research in nonhuman metacognition is subject to the criticism that, after considerable training on one test type, subjects learn to decline difficult trials based on associative learning of external test-specific contingencies rather than by evaluating the private status of memory or other cognitive states. We evaluated whether such test-specific associations could account for performance by presenting monkeys with a series of generalization tests across which no single association with external stimuli was likely to adaptively control use of the decline response. Six monkeys performed a four alternative delayed matching to location task and were significantly more accurate on trials with a decline option available than on trials without it, indicating that subjects selectively declined tests when memory was weak. Monkeys transferred appropriate use of the decline response under three conditions that assessed generalization: two tests that weakened memory and one test that enhanced memory in a novel way. Bidirectional generalization indicates that use of the decline response by monkeys is not controlled by specific external stimuli but is rather a flexible behavior based on a private assessment of memory.     

Potency of food deprivation intensity cues as discriminative stimuli.

Rats were trained to use stimuli arising from 0 and 24 hr without food as discriminative signals for shock. In Experiment 1, one group was shocked under 0-hr food deprivation and not shocked under 24-hr food deprivation. Another group received the reverse contingency. The groups received only 3 training trials under each deprivation level. Learning was revealed in a test phase when greater extinction of freezing was observed under the nonshocked than under the shocked deprivation level for both groups. A similar pattern of results was obtained in Experiment 2 when auditory cues were also relevant throughout training. Furthermore, prior training with food deprivation cues seemed to reduce learning about auditory cues subsequently trained in compound with deprivation stimuli. The results indicate that food deprivation intensity cues can be potent discriminative stimuli. The idea that deprivation cues function as conditioned modulatory stimuli cues is also discussed.