Nitric oxide signals that aplysia have attempted to eat, a necessary component of memory formation after learning that food is inedible

Inhibiting nitric oxide (NO) synthesis during learning that food is inedible in Aplysia blocks subsequent memory formation. To gain insight into the function of NO transmission during learning we tested whether blocking NO synthesis affects aspects of feeding that are expressed both in a nonlearning context and during learning. Inhibiting NO synthesis with L-NAME and blocking guanylyl cyclase with methylene blue decreased the efficacy of ad libitum feeding. D-NAME had no effect. L-NAME also decreased rejection responses frequency, but did not affect rejection amplitude. The effect of L-NAME was explained by a decreased signaling that efforts to swallow are not successful, leading to a decreased rejection rate, and a decreased ability to reposition and subsequently consume food in ad libitum feeding. Signaling that animals have made an effort to swallow is a critical component of learning that food is inedible. Stimulation of the lips with food alone did not produce memory, but stimulation combined with the NO donor SNAP did produce memory. Exogenous NO at a concentration causing memory also excited a key neuron responding to NO, the MCC. Block of the cGMP second-messenger cascade during training by methylene blue also blocked memory formation after learning. Our data indicate that memory arises from the contingency of three events during learning that food is inedible. One of the events is efforts to swallow, which are signaled by NO by cGMP.     

Reversible and Irreversible Stages in the Development of Amnesia after Disruption of the Reactivation of Associative Memory in Snails

Our previous studies on common snails have demonstrated that inhibition of NMDA glutamate receptors during reactivation of a skill consisting of refusal of a defined foodstuff leads to impairment of long-term memory. We report here our studies of the dynamics of the development of amnesia. Snails were trained to refuse a defined foodstuff and were injected 24 h later with the NMDA glutamate receptor antagonist MK-801, and were then presented with the conditioned food stimulus (a reminder). Testing on days 1 and 3 after exposure to MK-801 and the reminder showed gradual decreases in the number of refusals of the conditioned food stimulus. Repeat training of the animals to refuse the same foodstuff performed during these periods led to restoration of the skill seen after the initial training. The number of refusals by snails of the conditioned food stimulus 10 days after MK-801 and the reminder decreased to a minimal level. Repeat training at this time did not lead to the formation of a conditioned reflex to food. Thus, we have provided the first demonstration that impairment of the reactivation of long-term memory induces two stages in the development of amnesia. The first, reversible, stage, which lasted less than 10 days, was characterized by the potential for long-term memory to be restored by repeat training of the snails. The second, irreversible, stage developed 10 days after induction of amnesia and was characterized by disruption of the ability of long-term memory to be restored. These results may have practical value in terms of understanding the mechanisms of acute memory loss due to trauma and neurological diseases.     

Adoptive transfer of gut mucosal antitoxin memory by isolated B cells 1 year after oral immunization with cholera toxin.

A protocol was elaborated for the adoptive transfer of lymphocytes from mice which were orally immunized with cholera toxin (CT) to enable the study of long-term gut mucosal immunological memory at the single-cell level. Mesenteric lymph node (MLN) cells were transferred 1 year after priming immunizations, and recipient animals were challenged perorally on days 1 and 2 with CT before sacrifice on day 6 to 7 following transfer of cells. Strong antitoxin ELISPOT spot-forming cell (SFC) responses were recorded in spleens, MLN, and laminae propriae (LP) of recipient mice. In contrast, no SFC were found in Peyer's patches. The magnitude of the response equaled that of the acute response seen after optimal oral CT immunization and was directly dependent on the number of transferred cells. The memory antitoxin response in MLN and LP required oral challenge with CT as opposed to the spleen SFC response, which could also be triggered by intravenous challenge with antigen. Spleen cells from mice immunized perorally with CT were as effective as MLN cells in transferring immunological memory detectable in the gut immune system. Irrespective of the tissue source of transferring immunological memory detectable in the gut immune system. Irrespective of the tissue source of the memory cells, the isotype distribution of the antitoxin SFC response in recipient mice was similar with predominantly immunoglobulin A (96%) in LP and immunoglobulin G (66%) in MLN and spleen. Transfer of antitoxic memory was completely abrogated by treatment of the cells with J11d monoclonal antibody and complement prior to their injection into recipient mice by was unaffected by treatment with anti-Thy-1.2 antibody and complement, suggesting that long-term gut mucosal memory is carried by B cells. Antitoxin B memory cells might help explain the long-term protection against recurrent disease seen in convalescents from cholera in cholera-endemic areas.     

Immunization of carp (Cyprinus carpio) with a Vibrio anguillarum bacterin: indications for a common mucosal immune system

Uptake and transport of formalin-killed Vibrio anguillarum bacteria were studied in the second gut segment of carp and the resulting reaction of the immune system was investigated. Within a few hours after anal administration antigenic determinants of bacteria were present in intraepithelial macrophages of the second gut segment. In gut and skin mucus and bile, immunoglobulins (Ig's) were detected, but the amount was much lower than found in serum. In mucus of the second gut segment, 4 times more Ig (per mg protein) was found than in the first segment. Upon oral or anal immunization, slightly enhanced antigen-specific Ig titers could be detected in skin mucus and bile, but only after a booster along the same route. The existence of a common mucosal immune system is discussed, with special reference to the significance of the second gut segment. After anal intubation an increase of antigen-specific Ig could also be observed in serum. Following a booster, an enhanced cal memory. After anal boosting equal levels of serum antibody were reached compared with two consecutive intramuscular injections. However, no significant antibody increase occurred in serum after oral immunization, not even when bacteria were administered daily with the food.     

Long-Term Cholera Antitoxin Memory in the Gut can be Triggered to Antibody Formation Associated with Protection within Hours of an Oral Challenge Immunization

A local mucosal immunological memory that could be efficiently triggered to protective antibody formation on renewed antigen exposure might account for the several-year long protection against reinfection and disease seen in individuals after cholera disease. The duration and other functional aspects of gut mucosal immunological memory to the cholera toxin (CT), which is the key pathogenic factor in cholera, were examined in mice. Six months or even 2 years after an initial series of oral immunizations with CT a single repeat oral exposure to CT in submicrogram amounts evoked a brisk IgA antitoxin response in the lamina propria. A three-fold increase in IgA antitoxin-producing cells (SFC) was evident within 16 h, with a further rise in SFC numbers over the next several days. The anamnestic gut mucosal IgA antitoxin response was associated with a substantial increase in protection against challenge of intestinal loops with CT. The rapid increase in IgA antitoxin SFC in the gut is believed to reflect memory cells dispersed in the gut mucosa which can be rapidly triggered into antitoxin formation by antigen encounter in vivo and such cells could clearly be responsible for the long-term immunity seen after cholera disease or oral vaccination.     

Parametric features of inhibition of feeding in Aplysia by associative learning, satiation, and sustained lip stimulation

In order to determine whether different classes of behavioral plasticity affect common or unique neural loci, the effects of three types of processes that inhibit feeding in Aplysia were quantified. Changes in feeding behavior due to an associative learning task in which animals learn that food is inedible were compared with behavioral effects caused by satiation and by sustained lip stimulation. The data indicate that each process modifying feeding can be characterized by differences in time to stop responding to food, by differences in specificity of the decrement to a particular food, and by different patterns of motor output before complete cessation of responsiveness. The data suggest each process inhibiting feeding acts at a different neural site. Learning that food is inedible may be due to facilitation of a specific sensory pathway onto pattern generators producing rejection responses. Sustained lip stimulation seems to inhibit feeding by causing a decrement in all outputs of a particular sensory pathway. Finally, satiation appears to represent inhibition of feeding motor elements.     

The benefits of cognitive training after a coronary artery bypass graft surgery

Cognitive deficits are frequent after coronary artery bypass graft surgery (CABG) in the elderly population. In fact, memory and attention deficits can persist several months after the surgery. Recent studies with healthy older adults have shown that memory and attention can be improved through cognitive training programs. The present study examined whether memory training (method of loci and story generation) and attentional training (dual-task computerized training) could improve cognitive functions in patients aged 65 years and older who underwent CABG surgery. Participants (n = 51) were assigned to one of three groups: (1) control group (tested at 1, 3 and 6 months after the surgery), (2) attention training followed by memory training, (3) memory training followed by attention training (groups 2 and 3: tested at 1, 2, 3 and 6 months after the surgery). The trainings took place between the 6th and 10th week following the surgery. The three groups were compared before and after each training program using attention and memory tests and neuropsychological tests. The results showed that attention and memory trainings lead to significant improvement in the cognitive domain that was trained. It thus seems that cognitive training can be a promising tool to enhance cognitive functions after a CABG surgery.     

Presence of conspecifics facilitates learning that food is inedible in Aplysia fasciata.

The absence of a conspecific, but not of food, interfered with learning and memory of a feeding task in Aplysia fasciata. Interference was shown by a shortened training session and by lack of savings on retraining. The shortened training is not responsible for the lack of savings because brief training in the presence of a conspecific led to savings on retraining. Animals trained in the absence of a conspecific and then tested in its presence did not show signs of having learned, which indicates that the absence of a conspecific interfered with the ability to learn, rather than with the expression of memory. Absence of a conspecific also inhibited other aspects of feeding behavior, such as the latency to respond to food and the length of time that animals respond to food, which indicate that interference with learning was apparently caused by inhibition of feeding behavior, rather than by block of the mechanisms underlying learning.     

Spatial performance in a complex maze is associated with persistent long-term potentiation enhancement in mouse hippocampal slices at early training stages

Long-term potentiation (LTP) and long-term depression (LTD) are principal reflections of synaptic plasticity that have been implicated in learning and memory. We have previously shown that spatial learning in a newly validated complex maze is accompanied by depression of hippocampal CA1 synaptic activity in hippocampal slices of trained mice ("behavioral LTD"). In the present study, we investigated whether behavioral LTD is accompanied by alterations of subsequent LTP induced by high-frequency stimulation (HFS). Moreover, we were interested in the time course of such alterations in relation to training stage. Animals underwent 1, 2, and 8 days of spatial training in the complex maze, respectively. Hippocampal slices were taken 24 h after the last training session. We found a simultaneous decrease of basal synaptic response and increase of HFS induced LTP magnitude compared with slices of untrained animals. Synaptic plasticity was not influenced by repeated running wheel exercise in an additional control group without spatial learning. The mentioned alterations occurred already after day 2 of maze exploration parallel to the most pronounced improvement of behavioral performance but did not change thereafter until day 8 despite further learning progress. They were also found when animals were trained for 2 days and kept at rest for a subsequent 6 days. In conclusion, spatial learning may be reflected by distinct and persistent measurable alterations of synaptic plasticity in hippocampal CA1 neurons at early training stages.     

Temporary amnesia induced by cold anesthesia and hypoxia in Drosophila

Memory consolidation in Drosophila was investigated using cold anesthesia- and hypoxia-induced amnesia. Individual flies were operantly trained to avoid the specific flight orientations with respect to the landmarks surrounding them when paired with heat reinforcement at a flight simulator. Cold anesthesia, introduced immediately after training, exerted a significantly diminishing effect on memory between 15 and 150 min after training. Hypoxia delivered immediately after training had a significantly diminishing effect on memory between 30 and 150 min after training. In addition, cold anesthesia disrupted memory only when introduced within the first 20 min, while hypoxia worked only when delivered within the first 2 min after training. When interpreted in the context of a four-phase model of memory consolidation, the results suggest that 1) cold anesthesia disrupts both short-term memory (STM) and anesthesia-resistant memory (ARM), 2) hypoxia disrupts ARM specifically, 3) both of them leave long-term memory (LTM) intact, and 4) LTM may be independent of availability of STM and ARM in flies.     

NMDA receptor antagonists impair memory for nonspatial,socially transmitted food preference

Rats avoid unfamiliar foods and learn to prefer those that they smell on the breath of conspecifics. Hippocampal lesions produce rapid forgetting of this socially acquired memory. The authors report that NMDA receptor antagonists impair this memory. Rats given CPP were trained in the social transmission of food preference task. Normal rats showed robust memory 72 hr later. CPP-injected rats performed normally 24 hr, but randomly 72 hr, after training. Spatial context was irrelevant: Rats trained and tested in different rooms performed the same as rats trained and tested in 1 room. MK801 and intrahippocampal injections of APV produced amnestic effects similar to CPP. Thus, NMDA receptor activation is crucial for the persistence of socially acquired, hippocampus-dependent, nonspatial memory.     

Role of ACTH in recovery from retrograde amnesia induced by hypothermia in rats

The present investigation assessed whether increased congruency between ACTH state present shortly after training and that at testing contributed to memory recovery. If recovery were related to an increased correspondence between internal state present after training and that at testing, then suppressing ACTH release should block memory recovery. This was the hypothesis that was examined in the present investigation. Specifically, animals were trained on a passive avoidance task, administered hypothermia (the amnestic agent) and, shortly prior to testing, given treatments known to be effective in reversing memory loss induced by hypothermia. Before training (Experiment 1) or testing (Experiment 2) animals were injected with either dexamethasone (an agent that suppresses ACTH release) or saline. Results, in general, indicated that when ACTH release was suppressed, a blunted recovery effect was obtained. This reduction in the extent of memory recovery was observed when ACTH was suppressed either at training or at testing. These data are interpreted as providing support for an ACTH-related, state-dependent retention mechanism contributing to recovery from hypothermia-induced retrograde amnesia in rats.     

Early stages of memory formation in filial imprinting: Fos-like immunoreactivity and behavior in the domestic chick

Early stages of memory formation in filial imprinting were studied in domestic chicks. Chicks trained for 15 min showed strong imprinting, demonstrated by a strong preference for their training stimulus, and the time course of this preference over 2 days after training was similar to that of chicks trained for 60 min. The chicks therefore learned characteristics of the training stimulus very early during training. The intermediate and medial hyperstriatum ventrale (IMHV) is a part of the chick forebrain that is crucial for imprinting. Previous experiments have shown a learning-specific increase in Fos-like immunoreactivity, used as a marker of neuronal activity, in the IMHV after training for 60 min. The time course of Fos expression in the IMHV was measured after training for 15 min and 60 min. The same pattern of expression was found for both training times, showing a peak 120 min after the start of training. The time course of expression was stimulus-dependent. Fos expression in the IMHV, but not the hippocampus, was significantly correlated with strength of imprinting. It is concluded that the learning-specific change in Fos expression in the IMHV is associated with very early components of memory formation.     

Modulation of mucosal immunity in a murine model of food-induced intestinal inflammation

Background Hypersensitivity or uncontrolled responses against dietary antigens can lead to inflammatory disorders like food allergy and current models reflect a variety of causes but do not reveal the detailed modulation of gut immunity in response to food antigens after breakdown in mucosal tolerance. Objective To develop and characterize a murine model for food‐induced intestinal inflammation and to demonstrate the modulation of gut immune response by dietary allergenic antigens. Methods C57BL/6 mice were sensitized with peanut proteins, challenged with peanut seeds and their sera and gut segments were collected for subsequent analyses. Results Sensitization and challenged with peanut seeds led to alterations in gut architecture with inflammatory response characterized by oedema in lamina propria and cell infiltrate composed mainly by eosinophils, mast cells, phagocytes, natural killer and plasma cells, together with low percentage of γδ⁺ and CD4⁺CD25⁺Foxp3⁺ cells in Peyer's patches. These animals also presented high levels of specific IgE and IgG1 in sera and modulation of mucosal immunity was mediated by increased expression of GATA‐3, IL‐4, IL‐13 and TNF‐α in contrast to low IFN‐γ in the gut. Conclusion A murine model for food‐induced intestinal inflammation was characterized in which modulation of gut immunity occurs by peanut antigens in consequence of T‐helper type 2 (Th2) allergic response and failure of regulatory mechanisms necessary for mucosa homeostasis, resembling food allergy. This work shed some light on the understanding of the pathogenesis of gastrointestinal disorders and intolerance in the gut and supports the development of therapies for food‐related enteropathies like food allergy, focusing on gut‐specific immune response.     

Tissue-specific migration pathways by phenotypically distinct subpopulations of memory T cells.

A proportion of T cells recirculate in a tissue-selective manner. Recent studies which showed that the skin-tropic subset of T cells was of memory/activated type, led us to examine whether the preferential homing of T cells to the gut also involved memory T cells, and if so whether these memory T cells were phenotypically distinct from other memory T cells. Lymphocytes migrating through the gut and the skin of sheep was collected by cannulating the lymphatic ducts draining these tissues. Both naive and memory T cells were found to recirculate through the gut, although only memory T cells migrated through the skin. However, when T cells from the gut were labeled with fluorescein isothiocyanate and assessed for their migration back to the gut, it was the memory population which showed a tropism for the gut. Gut-tropic memory T cells migrated poorly through the skin, indicating that these cells were distinct from skin-tropic memory T cells. This was confirmed by phenotypic analysis. Gut memory T cells expressed very low levels of the alpha 6 and beta 1 integrins, in contrast to skin memory T cells which expressed high levels. There was no evidence for heterogeneity within the naive T cell population, which migrated preferentially to lymph nodes. This migration pattern could be explained in part by the high expression of the L-selectin (lymph node homing receptor, LAM-1) on naive T cells, in contrast to memory T cells from gut or skin which were mostly L-selectin negative. These results in sheep indicate that subsets of alpha/beta memory T cells show tissue-selective migration patterns, which probably develop in a particular environment following encounter with antigen.     

Induction of Avian Immunological Responsiveness Following Cloacal Drinking of Immunogen

A solution of human serum albumin which has been cloacally ingested can cause effective development of immunological memory with minimal or no immediate production of antibody in 4 to 6 week old chickens. Such memory production was not accompanied by the generation of circulating rosette-forming ABC (antigen binding cells) and thus the peripheral appearance of such cells as a part of the normal primary response to HSA (human serum albumin) does not accord with the interpretation that they represent memory B cells. The cloacal route may be important for the maturing chick in initiating immunity to the food and micro-organisms of its external environment and gut flora.     

Induction of avian immunological responsiveness following cloacal drinking of immunogen.

A solution of human serum albumin which has been cloacally ingested can cause effective development of immunological memory with minimal or no immediate production of antibody in 4 to 6 week old chickens. Such memory production was not accompanied by the generation of circulating rosette-forming ABC (antigen binding cells) and thus the peripheral appearance of such cells as a part of the normal primary response to HSA (human serum albumin) does not accord with the interpretation that they represent memory B cells. The cloacal route may be important for the maturing chick in initiating immunity to the food and micro-organisms of its external environment and gut flora.