Molecular insights from the crab Neohelice memory model.

Memory acquisition, formation and maintenance depend on synaptic post-translational machinery and regulation of gene expression triggered by several transduction pathways. In turns, these processes lead to stabilization of synaptic modifications in neurons in the activated circuits. In order to study the molecular mechanisms involved in acquisition and memory, we have taken advantage of the context-signal associative learning and, more recently, the place preference task, of the crab Neohelice granulata. In this model organism, we studied several molecular processes, including activation of extracellular signal-regulated kinase (ERK) and the nuclear factor kappa light chain enhancer of activated B cells (NF-κB) transcription factor, involvement of synaptic proteins such as NMDA receptors and neuroepigenetic regulation of gene expression. All these studies allowed description of key plasticity mechanisms involved in memory, including consolidation, reconsolidation and extinction. This article is aimed at review the most salient findings obtained over decades of research in this memory model.

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.

Requirement of NF-kappa B Activation in Different Mice Brain Areas during Long-Term Memory Consolidation in Two Contextual One-Trial Tasks with Opposing Valences.

NF-kappa B is a transcription factor whose activation has been shown to be necessary for long-term memory consolidation in several species. NF-kappa B is activated and translocates to the nucleus of cells in a specific temporal window during consolidation. Our work focuses on a one trial learning tasks associated to the inhibitory avoidance (IA) setting. Mice were trained either receiving or not a footshock when entering a dark compartment (aversive vs. appetitive learning). Regardless of training condition (appetitive or aversive), latencies to step-through during testing were significantly different to those measured during training. Additionally, these testing latencies were also different from those of a control group that only received a shock unrelated to context. Moreover, nuclear NF-kappa B DNA-binding activity was augmented in the aversive and the appetitive tasks when compared with control and naïve animals. NF-kappa B inhibition by Sulfasalazine injected either in the Hippocampus, Amygdala or Nucleus accumbens immediately after training was able to impair retention in both training versions. Our results suggest that NF-kappa B is a critical molecular step, in different brain areas on memory consolidation. This was the case for both the IA task and also the modified version of the same task where the footshock was omitted during training. This work aims to further investigate how appetitive and aversive memories are consolidated.

Surface expression of NMDA receptor changes during memory consolidation in the crab Neohelice granulata.

The aim of the present study was to analyze the surface expression of the NMDA-like receptors during the consolidation of contextual learning in the crab Neohelice granulata Memory storage is based on alterations in the strength of synaptic connections between neurons. The glutamatergic synapses undergo various forms of N-methyl-D aspartate receptor (NMDAR)-dependent changes in strength, a process that affects the abundance of other receptors at the synapse and underlies some forms of learning and memory. Here we propose a direct regulation of the NMDAR. Changes in NMDAR's functionality might be induced by the modification of the subunit's expression or cellular trafficking. This trafficking does not only include NMDAR's movement between synaptic and extra-synaptic localizations but also the cycling between intracellular compartments and the plasma membrane, a process called surface expression. Consolidation of contextual learning affects the surface expression of the receptor without affecting its general expression. The surface expression of the GluN1 subunit of the NMDAR is down-regulated immediately after training, up-regulated 3 h after training and returns to naïve and control levels 24 h after training. The changes in NMDAR surface expression observed in the central brain are not seen in the thoracic ganglion. A similar increment in surface expression of GluN1 in the central brain is observed 3 h after administration of the competitive GABAA receptor antagonist, bicuculline. These consolidation changes are part of a plasticity event that first, during the down-regulation, stabilizes the trace and later, at 3-h post-training, changes the threshold for synapse activation.

NF-κB transcription factor role in consolidation and reconsolidation of persistent memories.

Transcriptional regulation is an important molecular process required for long-term neural plasticity and long-term memory (LTM) formation. Thus, one main interest in molecular neuroscience in the last decades has been the identification of transcription factors that are involved in memory processes. Among them, the nuclear factor κB (NF-κB) family of transcription factors has gained interest due to a significant body of evidence that supports a key role of these proteins in synaptic plasticity and memory. In recent years, the interest was particularly reinforced because NF-κB was characterized as an important regulator of synaptogenesis. This function may be explained by its participation in synapse to nucleus communication, as well as a possible local role at the synapse. This review provides an overview of experimental work obtained in the last years, showing the essential role of this transcription factor in memory processes in different learning tasks in mammals. We focus the review on the consolidation and reconsolidation memory phases as well as on the regulation of immediate-early and late genes by epigenetic mechanisms that determine enduring forms of memories.

Protein degradation by ubiquitin-proteasome system in formation and labilization of contextual conditioning memory.

The ubiquitin-proteasome system (UPS) of protein degradation has been evaluated in different forms of neural plasticity and memory. The role of UPS in such processes is controversial. Several results support the idea that the activation of this system in memory consolidation is necessary to overcome negative constrains for plasticity. In this case, the inhibition of the UPS during consolidation impairs memory. Similar results were reported for memory reconsolidation. However, in other cases, the inhibition of UPS had no effect on memory consolidation and reconsolidation but impedes the amnesic action of protein synthesis inhibition after retrieval. The last finding suggests a specific action of the UPS inhibitor on memory labilization. However, another interpretation is possible in terms of the synthesis/degradation balance of positive and negative elements in neural plasticity, as was found in the case of long-term potentiation. To evaluate these alternative interpretations, other reconsolidation-interfering drugs than translation inhibitors should be tested. Here we analyzed initially the UPS inhibitor effect in contextual conditioning in crabs. We found that UPS inhibition during consolidation impaired long-term memory. In contrast, UPS inhibition did not affect memory reconsolidation after contextual retrieval but, in fact, impeded memory labilization, blocking the action of drugs that does not affect directly the protein synthesis. To extend these finding to vertebrates, we performed similar experiments in contextual fear memory in mice. We found that the UPS inhibitor in hippocampus affected memory consolidation and blocked memory labilization after retrieval. These findings exclude alternative interpretations to the requirement of UPS in memory labilization and give evidence of this mechanism in both vertebrates and invertebrates.

Synaptic NF-kappa B pathway in neuronal plasticity and memory.

Several transcription factors are present at the synapse, and among these are the Rel-NF-kappa B pathway components. NF-kappa B is a constitutive transcription factor, with a strong presence in the brain of which a considerable part is located in the neuropiles. This localization of the transcription factor, plus evidence pointing to different functions, is what gave place to two general hypotheses for synaptic NF-kappa B: (a) The transcription factor plays a role in the synapse to nucleus communication, and it is retrogradely transported from polarized localizations to regulate gene expression; (b) The transcription factor modulates the synaptic function locally. Evidence indicates that both mechanisms can operate simultaneously; here we will present different possibilities of these hypotheses that are supported by an increasing amount of data. We pay special attention to the local role of the transcription factor at the synapse, and based in the described evidence from different animal models, we propose several processes in which the transcription factor may change the synaptic strength.

Characterization of the cardiac ganglion in the crab Neohelice granulata and immunohistochemical evidence of GABA-like extrinsic regulation.

The aim of the present work is to provide an anatomical description of the cardiac system in the crab Neohelice granulata and evidence of the presence of GABA by means of immunohistochemistry. The ganglionic trunk was found lying on the inner surface of the heart's dorsal wall. After dissection, this structure appeared as a Y-shaped figure with its major axis perpendicular to the major axis of the heart. Inside the cardiac ganglion, we identified four large neurons of 63.7 µm ± 3.7 in maximum diameter, which were similar to the motor neurons described in other decapods. All the GABA-like immunoreactivity (GABAi) was observed as processes entering mainly the ganglionic trunk and branching in slender varicose fibers, forming a network around the large neurons suggesting that GABAi processes contact them. Our findings strengthen previous results suggesting that the GABAergic system mediates the cardio-inhibitory response upon sensory stimulation.

NMDA-like receptors in the nervous system of the crab Neohelice granulata: a neuroanatomical description.

N-Methyl-D-aspartate receptors (NMDARs) are involved in learning and memory processes in vertebrates and invertebrates. In Neohelice granulata, NMDARs are involved in the storage of associative memories (see references in text). The aim of this work was to characterize this type of glutamate receptor in Neohelice and to describe its distribution in the central nervous system (CNS). As a first step, a detailed study of the CNS of N. granulata was performed at the neuropil level, with special focus on one of the main structures involved in this type of memory, the supraesophageal ganglion, called central brain. The characterization of the NMDAR was achieved by identifying the essential subunit of these receptors, the NR1-like subunit. The NR1-like signals were found via western blot and immunohistochemistry techniques in each of the major ganglia: the eyestalk ganglia, the central brain, and the thoracic ganglion. Western blots yielded two bands for the crab NR1-like subunit, at ∼88 and ∼84 kDa. This subunit is present in all the major ganglia, and shows a strong localization in synaptosomal membranes. NMDARs are distributed throughout the majority of each ganglion but show prominent signal intensity in some distinguishable neuropils and neurons. This is the first general description of the N. granulata nervous system as a whole and the first study of NMDARs in the CNS of decapods. The preferential localization of the receptor in some neuropils and neurons indicates the presence of possible new targets for memory processing and storage.