Behavioral and Molecular Analysis of Memory in the Dwarf Cuttlefish

Complex memory has evolved because it benefits animals in all areas of life, such as remembering the location of food or conspecifics, and learning to avoid dangerous stimuli. Advances made by studying relatively simple nervous systems, such as those in gastropod mollusks, can now be used to study mechanisms of memory in more complex systems. Cephalopods offer a unique opportunity to study the mechanisms of memory in a complex invertebrates. The dwarf cuttlefish, Sepia bandensis, is a useful memory model because its fast development and small size allows it to be reared and tested in large numbers. However, primary literature regarding the behavior and neurobiology of this species is lacking. This research determined that juvenile S. bandensis exhibited short term memory (STM) and long term memory (LTM). To assess memory in dwarf cuttlefish, a memory test was conducted which utilized the predatory attack in cuttlefish. It was found that 4 week old dwarf cuttlefish retained memory of the experiment up to 4 days. Using an automated tracking software called DanioVision, this research found that cuttlefish selectively inhibit the tentacle striking phase of their predatory behavior, without inhibiting the attention and positioning phases. Determining the molecular mechanisms underlying memory is key to understanding how memory is manifested in the form of altered behavior. At the cellular level, memories are formed by altering the physical and chemical properties within specific neural circuits. The transcription factor, CREB, is responsible for transcribing genes required for initiating these long-term neuronal changes. Using immunohistochemistry, a molecular assay was developed to determine whether CREB is activated in cuttlefish arms during the memory experiment. Trained cuttlefish had a greater number of CREB positive cells in the epithelium of the arm than controls. Trained cuttlefish also had a greater average number of CREB-positive cells in positive suckers than untrained cuttlefish. These results suggest that CREB activation may result from behavioral training in cuttlefish. Lastly, it was found that the distal tip of the arm contained more CREB-positive cells than the proximal part of the arm. Spatial activation of CREB may occur predominantly in the distal portion of the arm. By locating CREB for the first time in a cephalopod, this research presents dwarf cuttlefish as interesting models for studying the molecular mechanisms of memory formation

Molluscan memory of injury: evolutionary insights into chronic pain and neurological disorders.

Molluscan preparations have yielded seminal discoveries in neuroscience, but the experimental advantages of this group have not, until now, been complemented by adequate molecular or genomic information for comparisons to genetically defined model organisms in other phyla. The recent sequencing of the transcriptome and genome of Aplysia californica, however, will enable extensive comparative studies at the molecular level. Among other benefits, this will bring the power of individually identifiable and manipulable neurons to bear upon questions of cellular function for evolutionarily conserved genes associated with clinically important neural dysfunction. Because of the slower rate of gene evolution in this molluscan lineage, more homologs of genes associated with human disease are present in Aplysia than in leading model organisms from Arthropoda (Drosophila) or Nematoda (Caenorhabditis elegans). Research has hardly begun in molluscs on the cellular functions of gene products that in humans are associated with neurological diseases. On the other hand, much is known about molecular and cellular mechanisms of long-term neuronal plasticity. Persistent nociceptive sensitization of nociceptors in Aplysia displays many functional similarities to alterations in mammalian nociceptors associated with the clinical problem of chronic pain. Moreover, in Aplysia and mammals the same cell signaling pathways trigger persistent enhancement of excitability and synaptic transmission following noxious stimulation, and these highly conserved pathways are also used to induce memory traces in neural circuits of diverse species. This functional and molecular overlap in distantly related lineages and neuronal types supports the proposal that fundamental plasticity mechanisms important for memory, chronic pain, and other lasting alterations evolved from adaptive responses to peripheral injury in the earliest neurons. Molluscan preparations should become increasingly useful for comparative studies across phyla that can provide insight into cellular functions of clinically important genes

Psychoneural reduction: a perspective from neural circuits

Abstract: Psychoneural reduction has been debated extensively in the philosophy of neuroscience. In this article I will evaluate metascientific approaches that claim direct molecular and cellular explanations of cognitive functions. I will initially consider the issues involved in linking cellular properties to behaviour from the general perspective of neural circuits. These circuits that integrate the molecular and cellular components underlying cognition and behaviour, making consideration of circuit properties relevant to reductionist debates. I will then apply this general perspective to specific systems where psychoneural reduction has been claimed, namely hippocampal long-term potentiation and the Aplysia gill-withdrawal reflex

Kinase C substrates and synaptic plasticity in Aplysia

Thèse numérisée par la Direction des bibliothèques de l'Université de Montréal

Neurexins and Neuroligins: Recent Insights from Invertebrates

During brain development, each neuron must find and synapse with the correct pre- and postsynaptic partners. The complexity of these connections and the relatively large distances some neurons must send their axons to find the correct partners makes studying brain development one of the most challenging, and yet fascinating disciplines in biology. Furthermore, once the initial connections have been made, the neurons constantly remodel their dendritic and axonal arbours in response to changing demands. Neurexin and neuroligin are two cell adhesion molecules identified as important regulators of this process. The importance of these genes in the development and modulation of synaptic connectivity is emphasised by the observation that mutations in these genes in humans have been associated with cognitive disorders such as Autism spectrum disorders, Tourette syndrome and Schizophrenia. The present review will discuss recent advances in our understanding of the role of these genes in synaptic development and modulation, and in particular, we will focus on recent work in invertebrate models, and how these results relate to studies in mammals

An Investigation of the Neurophysiological Correspondents of Learning and Memory in Two Forebrain Regions of the Day-Old Chick

Spontaneous bursting (5 or more spikes of 200-450mV amplitude at 400Hz) occurs in many areas of chick forebrain. Day-old chicks trained on a one-trial passive avoidance task show a bilateral increase of up to 350% in bursting following training in one of these areas: the intermediate medial hyperstriatum ventrale, or IMHV (Mason & Rose, 1987; 1988). An investigation was carried out into the time course and lateralization of this change in bursting activity following the training of day-old chicks on a passive avoidance task. Chicks were trained to either avoid a bead coated with the bitter-tasting substance methylanthranilate (M-birds) or were trained to peck a water coated bead (W-birds). Bursting was recorded sequentially from the IMHV of both hemispheres at 8 time points over the period 1 to 9 hours post-test. The results indicate that there are significant differences in bursting activity recorded from M-birds only during the period 3-7hr posttest, when compared to W-birds. Between 6-7hr posttest there are significant differences in the burst firing patterns of the right IMHV of M-birds compared to the left. At other time points tested there are no significant differences between hemispheres. No between hemisphere differences are evident in W-birds. Multi-unit recordings were made from the lobus parolfactorius (LPO), another forebrain structure to show changes in biochemistry and morphology following passive avoidance training. M-birds showed a higher incidence of bursting when compared to W-birds over the period 1-10hr posttest. No lateralization of bursting was seen in either group at any time posttest. In a further experiment, chicks trained to avoid the methylanthranilate coated bead were subjected to subconvulsive electroshock 5min posttraining. This procedure was used to test whether the training-induced increase in bursting in the LPO was a direct correlate of memory formation for the task. This electroshock treatment produced two groups of birds: one group that avoided the bead (remembered the task) and another that pecked the bead (forgot the task). Multi-unit recordings from the LPO of these two groups revealed that the group that avoided the bead had a significantly higher mean burst-frequency when compared to the group that pecked the bead, indicating that increased bursting in the LPO following training is directly associated with recall for the task. These results are similar to those of Mason and Rose (1988) who showed that amnesia abolished a training-induced enhancement of bursting in the IMHV. The effects of pretraining bilateral LPO lesions on IMHV bursting activity were examined. The IMHV of four groups of birds was recorded ftom following training: two groups of M-birds, one with LPO lesions the other with sham LPO lesions and two similarly treated groups of W-birds. A significant increase in overall IMHV bursting activity was observed in sham-lesioned M-birds when compared to sham-lesioned W-birds. However, no significant difference in bursting activity was seen between lesioned M-birds and lesioned W-birds. There was a trend towards a higher overall level of bursting in lesioned W-birds, when compared to sham-lesioned W-birds. These results are discussed with reference to previous electrophysiological studies concerning the role of burst-firing patterns in models of learning and memory

Neuronal Activity-Dependent Development of the Nociceptive Circuit in Drosophila

How nature and nurture interact to sculpt the nervous system, which underlies animal behaviors, has fascinated both scientists and the general public for generations. At the level of neural circuit assembly, the answer lies in the interplay between genetic programs and neural activity; the development of a functional nervous system is not just hard-wired by the genome, but depends on sensory experiences and neuronal activities. However, the mechanisms underlying the “activity-dependent development” of the nervous system are poorly understood, mostly due to the lack of a model system that is amenable to efficient gene manipulations and circuit analyses. My dissertation research aims to develop a Drosophila system that is suitable for identifying the mechanisms behind activity-dependent development of neural circuits from the molecular to the circuit and behavioral levels. I have largely achieved this through two projects. First, I discovered that the functional development of Drosophila somatosensory circuits depends on the sensory inputs during animal development. Our behavior analysis demonstrated that larval escaping behavior in response to noxious stimulation is suppressed if a larva experiences enhanced levels of noxious stimulation during development, demonstrating sensory input-induced plasticity. Using imaging-based physiological analyses (calcium and cAMP imaging techniques and optogenetic stimulation of neurons), we found that enhanced noxious stimulation during development reduces the synaptic transmission from nociceptors (i.e., sensory neurons detecting noxious stimuli) to the second-order neurons (SONs) in the pathway. Our study further revealed that this physiological change accounts for the suppressed behavioral outputs. Importantly, we showed that the enhanced noxious experience has no effect on other sensory modalities such as the mechanosensory pathway and elucidated the mechanism that underlies this sensory-pathway-specificity. Second, my work facilitated the discovery that the activity levels of nociceptors regulate their axonal projections in the central nervous system (CNS). Through advanced techniques that combine single-cell labeling and computational analysis, we found that the spatial arrangement of nociceptor axon terminals in the CNS reflects the locations of territories occupied by nociceptor dendrites on the body wall, forming a topographic map. The formation of this map depends on the levels of their activities, and manipulation of neuronal activity at single-cell level disrupts the map formation. This activity-dependent topography in Drosophila is likely established through the interactions of nociceptor presynaptic terminals with their postsynaptic SONs, similar to topography in vertebrates. This work is the first report of an activity-dependent topographic map in Drosophila, and has allowed for mechanistic analyses of the role of neuronal activity in neural circuit wiring. My dissertation research contributes to our understanding of how neural activity interacts with genetic programs to shape the nervous system.PHDCell and Developmental BiologyUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttps://deepblue.lib.umich.edu/bitstream/2027.42/144193/1/tkaneko_1.pd

Novel Small-RNA Mediated Gene Regulatory Mechanisms for Long-Term Memory

Memory storage and memory-related synaptic plasticity rely on precise spatiotemporal regulation of gene expression. To explore the role of small RNAs in memory-related synaptic plasticity we carried out massive parallel sequencing to profile the small RNAs of Aplysia. We identified 170 distinct 21-23 nt sized miRNAs, 13 of which were novel and specific to Aplysia. Nine miRNAs were brain-enriched, and several of these were rapidly down-regulated by transient exposure to serotonin, a modulatory neurotransmitter released during learning. Two abundant, and conserved brain-specific miRNAs, miR-124 and miR-22 were exclusively present pre-synaptically in a sensory-motor synapse where they constrain synaptic facilitation through regulation of the transcriptional factor CREB1 and translation factor CPEB respectively. We therefore provide the first evidence that a modulatory neurotransmitter important for learning can regulate the levels of small RNAs and present a novel role for miR-124 and miR-22 in long-term plasticity of synapses in the mature nervous system. While mining the small RNA libraries for miRNAs, we discovered an unexpected and abundant expression in brain of a 28-nt sized class of piRNAs, which had been thought to be germ-line specific. These piRNAs have unique biogenesis patterns and predominant nuclear localization. Moreover, we find that whereas miRNAs are down-regulated by exposure to serotonin, piRNAs are up-regulated. Importantly, we find that the piwi/piRNA complex facilitates serotonin-dependent methylation of a conserved CpG island in the promoter of CREB2, the major inhibitory constraint of memory in Aplysia, leading to the persistence of long-term synaptic facilitation. Taken together, these findings provide a new serotonin-dependent, bidirectional, small-RNA mediated gene regulatory mechanism during plasticity where miRNAs provide translational control and piRNAs provide long-lasting transcriptional control for the persistence of memory

Activation et inactivation abrupte de sites synaptiques aux synapses sensori-motrices de l'Aplysie

Mémoire numérisé par la Direction des bibliothèques de l'Université de Montréal