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The Journal of Neuroscience : the... Jan 1989We prepared and characterized subcellular membrane fractions from the CNS of Aplysia californica that are enriched in isolated nerve terminals (synaptosomes). Ganglia...
We prepared and characterized subcellular membrane fractions from the CNS of Aplysia californica that are enriched in isolated nerve terminals (synaptosomes). Ganglia were homogenized in 1.1 M sucrose and fractionated on a 2-step sucrose gradient, yielding 50 micrograms protein/animal in the synaptosomal fraction (P3), which was enriched 3-fold in plasma membrane as compared with the initial homogenate. Quantitative morphometry of electron micrographs revealed that P3 contained 25% intact synaptosomes, a 5-fold enrichment over the homogenate. Although fractionation on a 5-step sucrose gradient reduced the yield of protein in the synaptosomal fraction to 40 micrograms/animal, this fraction (the 0.35 M/0.75 M interface) was more enriched in plasma membrane than P3 and was less contaminated by lysosomes and free mitochondria. By electron microscopy, the 0.35 M/0.75 M interface contained up to 50% synaptosomes. Synaptosomal fractions contained cAMP-, Ca2+/calmodulin-, and Ca2+/phospholipid-dependent protein kinase activities and were enriched in a Mr 40,000 pertussis toxin substrate, Gi/o. In the accompanying paper, we show that these synaptosomes retain the ability to release transmitters.
Topics: Animals; Aplysia; Central Nervous System; Centrifugation, Density Gradient; Ganglia; Membranes; Pertussis Toxin; Protein Kinases; Subcellular Fractions; Synaptosomes; Virulence Factors, Bordetella
PubMed: 2913212
DOI: 10.1523/JNEUROSCI.09-01-00038.1989 -
Neuron Mar 1997
Review
Topics: Animals; Aplysia; Gene Expression Regulation; Humans; Long-Term Potentiation; Memory; Models, Neurological; Nerve Tissue Proteins; RNA, Messenger; Serotonin; Synapses
PubMed: 9115727
DOI: 10.1016/s0896-6273(00)81234-9 -
Sleep Dec 2016Insufficient sleep in individuals appears increasingly common due to the demands of modern work schedules and technology use. Consequently, there is a growing need to...
STUDY OBJECTIVES
Insufficient sleep in individuals appears increasingly common due to the demands of modern work schedules and technology use. Consequently, there is a growing need to understand the interactions between sleep deprivation and memory. The current study determined the effects of acute sleep deprivation on short and long-term associative memory using the marine mollusk , a relatively simple model system well known for studies of learning and memory.
METHODS
were sleep deprived for 9 hours using context changes and tactile stimulation either prior to or after training for the operant learning paradigm, learning that food is inedible (LFI). The effects of sleep deprivation on short-term (STM) and long-term memory (LTM) were assessed.
RESULTS
Acute sleep deprivation prior to LFI training impaired the induction of STM and LTM with persistent effects lasting at least 24 h. Sleep deprivation immediately after training blocked the consolidation of LTM. However, sleep deprivation following the period of molecular consolidation did not affect memory recall. Memory impairments were independent of handling-induced stress, as daytime handled control animals demonstrated no memory deficits. Additional training immediately after sleep deprivation failed to rescue the induction of memory, but additional training alleviated the persistent impairment in memory induction when training occurred 24 h following sleep deprivation.
CONCLUSIONS
Acute sleep deprivation inhibited the induction and consolidation, but not the recall of memory. These behavioral studies establish as an effective model system for studying the interactions between sleep and memory formation.
Topics: Animals; Aplysia; Association Learning; Conditioning, Operant; Memory, Long-Term; Memory, Short-Term; Sleep Deprivation
PubMed: 27748243
DOI: 10.5665/sleep.6320 -
Scientific Reports Mar 2021ATP and its ionotropic P2X receptors are components of the most ancient signaling system. However, little is known about the distribution and function of purinergic...
ATP and its ionotropic P2X receptors are components of the most ancient signaling system. However, little is known about the distribution and function of purinergic transmission in invertebrates. Here, we cloned, expressed, and pharmacologically characterized the P2X receptors in the sea slug Aplysia californica-a prominent neuroscience model. AcP2X receptors were successfully expressed in Xenopus oocytes and displayed activation by ATP with two-phased kinetics and Na-dependence. Pharmacologically, they were different from other P2X receptors. The ATP analog, Bz-ATP, was a less effective agonist than ATP, and PPADS was a more potent inhibitor of the AcP2X receptors than the suramin. AcP2X were uniquely expressed within the cerebral F-cluster, the multifunctional integrative neurosecretory center. AcP2X receptors were also detected in the chemosensory structures and the early cleavage stages. Therefore, in molluscs, rapid ATP-dependent signaling can be implicated both in development and diverse homeostatic functions. Furthermore, this study illuminates novel cellular and systemic features of P2X-type ligand-gated ion channels for deciphering the evolution of neurotransmitters.
Topics: Adenosine Triphosphate; Animals; Aplysia; Models, Molecular; Neurons; Phylogeny; Receptors, Purinergic P2X; Signal Transduction; Xenopus
PubMed: 33750901
DOI: 10.1038/s41598-021-84981-5 -
Molecular Brain Jun 2008Whereas the induction of short-term memory involves only covalent modifications of constitutively expressed preexisting proteins, the formation of long-term memory... (Review)
Review
Whereas the induction of short-term memory involves only covalent modifications of constitutively expressed preexisting proteins, the formation of long-term memory requires gene expression, new RNA, and new protein synthesis. On the cellular level, transcriptional regulation is thought to be the starting point for a series of molecular steps necessary for both the initiation and maintenance of long-term synaptic facilitation (LTF). The core molecular features of transcriptional regulation involved in the long-term process are evolutionally conserved in Aplysia, Drosophila, and mouse, and indicate that gene regulation by the cyclic AMP response element binding protein (CREB) acting in conjunction with different combinations of transcriptional factors is critical for the expression of many forms of long-term memory. In the marine snail Aplysia, the molecular mechanisms that underlie the storage of long-term memory have been extensively studied in the monosynaptic connections between identified sensory neuron and motor neurons of the gill-withdrawal reflex. One tail shock or one pulse of serotonin (5-HT), a modulatory transmitter released by tail shocks, produces a transient facilitation mediated by the cAMP-dependent protein kinase leading to covalent modifications in the sensory neurons that results in an enhancement of transmitter release and a strengthening of synaptic connections lasting minutes. By contrast, repeated pulses of 5-hydroxytryptamine (5-HT) induce a transcription- and translation-dependent long-term facilitation (LTF) lasting more than 24 h and trigger the activation of a family of transcription factors in the presynaptic sensory neurons including ApCREB1, ApCREB2 and ApC/EBP. In addition, we have recently identified novel transcription factors that modulate the expression of ApC/EBP and also are critically involved in LTF. In this review, we examine the roles of these transcription factors during consolidation of LTF induced by different stimulation paradigms.
Topics: Animals; Aplysia; Aquatic Organisms; Gene Expression Regulation; Memory, Long-Term; Seawater; Snails; Transcription, Genetic
PubMed: 18803855
DOI: 10.1186/1756-6606-1-3 -
The Journal of Neuroscience : the... Mar 2021Modulators are generally expected to establish a network configuration that is appropriate for the current circumstances. We characterize a situation where the opposite...
Modulators are generally expected to establish a network configuration that is appropriate for the current circumstances. We characterize a situation where the opposite is apparently observed. A network effect of a peptide modulator is counterproductive in that it tends to impede rather than promote the creation of the configuration that is appropriate when the modulator is released. This raises a question: why does release occur? We present data that strongly suggest that it impacts task switching. Our experiments were conducted in an feeding network that generates egestive and ingestive motor programs. Initial experiments focused on egestive activity and the neuron B8. As activity becomes egestive, there is an increase in synaptic drive to B8 and its firing frequency increases (Wang et al., 2019). We show that, as this occurs, there is also a persistent current that develops in B8 that is outward rather than inward. Dynamic clamp introduction of this current decreases excitability. When there is an egestive-ingestive task switch in , negative biasing is observed (i.e., a bout of egestive activity has a negative impact on a subsequent attempt to initiate an ingestive response) (Proekt et al., 2004). Using an analog of negative biasing, we demonstrate that the outward current that develops during egestive priming plays an important role in establishing this phenomenon. Our data suggest that, although the outward current induced as activity becomes egestive is counterproductive at the time, it plays an anticipatory role in that it subsequently impacts task switching. In this study, we identify a peptide-induced circuit modification (induction of an outward current) that does not immediately promote the establishment of a behaviorally appropriate network configuration. We ask why this might occur, and present data that strongly suggest that it plays an important role during task switching. Specifically, our data suggest that the outward current we characterize plays a role in the negative biasing that is seen in the mollusc when there is a transition from egestive to ingestive activity. It is possible that the mechanism that we describe operates in other species. A negative effect of egestion on subsequent ingestion is observed throughout the animal kingdom.
Topics: Action Potentials; Animals; Aplysia; Feeding Behavior; Ganglia, Invertebrate; Motor Neurons
PubMed: 33500278
DOI: 10.1523/JNEUROSCI.2427-20.2021 -
Molecular Biology of the Cell Sep 2015Src tyrosine kinases have been implicated in axonal growth and guidance; however, the underlying cellular mechanisms are not well understood. Specifically, it is unclear...
Src tyrosine kinases have been implicated in axonal growth and guidance; however, the underlying cellular mechanisms are not well understood. Specifically, it is unclear which aspects of actin organization and dynamics are regulated by Src in neuronal growth cones. Here, we investigated the function of Src2 and one of its substrates, cortactin, in lamellipodia and filopodia of Aplysia growth cones. We found that up-regulation of Src2 activation state or cortactin increased lamellipodial length, protrusion time, and actin network density, whereas down-regulation had opposite effects. Furthermore, Src2 or cortactin up-regulation increased filopodial density, length, and protrusion time, whereas down-regulation promoted lateral movements of filopodia. Fluorescent speckle microscopy revealed that rates of actin assembly and retrograde flow were not affected in either case. In summary, our results support a model in which Src and cortactin regulate growth cone motility by increasing actin network density and protrusion persistence of lamellipodia by controlling the state of actin-driven protrusion versus retraction. In addition, both proteins promote the formation and stability of actin bundles in filopodia.
Topics: Actins; Animals; Aplysia; Axons; Cells, Cultured; Cortactin; Down-Regulation; Growth Cones; Pseudopodia; Up-Regulation; src-Family Kinases
PubMed: 26224308
DOI: 10.1091/mbc.E15-03-0142 -
Scientific Reports Mar 2020The formation of appropriate neural connections during development is critical for the proper wiring and functioning of the brain. Although considerable research...
Specificity of synapse formation in Aplysia: paracrine and autocrine signaling regulates bidirectional molecular interactions between sensory and non-target motor neurons.
The formation of appropriate neural connections during development is critical for the proper wiring and functioning of the brain. Although considerable research suggests that the specificity of synapse formation is supported by complex intercellular signaling between potential presynaptic and postsynaptic partners, the extracellular factors and the intracellular signal transduction pathways engaged in this process remain largely unknown. Using the sensory-motor neural circuit that contributes to learning in defensive withdrawal reflexes in Aplysia californica, we investigated the molecular processes governing the interactions between sensory neurons and both target and non-target motor neurons during synapse formation in culture. We found that evolutionarily-conserved intercellular and intracellular signaling mechanisms critical for learning-related plasticity are also engaged during synaptogenesis in this in vitro model system. Our results reveal a surprising bidirectional regulation of molecular signaling between sensory neurons and non-target motor neurons. This regulation is mediated by signaling via both paracrine and autocrine diffusible factors that induce differential effects on transcription and on protein expression/activation in sensory neurons and in target and non-target motor neurons. Collectively, our data reveal novel molecular mechanisms that could underlie the repression of inappropriate synapse formation, and suggest mechanistic similarities between developmental and learning-related plasticity.
Topics: Animals; Aplysia; Autocrine Communication; Coculture Techniques; Cyclic AMP Response Element-Binding Protein; Extracellular Signal-Regulated MAP Kinases; Motor Neurons; Neuronal Plasticity; Neuropeptides; Paracrine Communication; Receptor, trkB; Sensory Receptor Cells; Signal Transduction; Single-Cell Analysis; Synapses
PubMed: 32251363
DOI: 10.1038/s41598-020-62099-4 -
BMC Genomics Jul 2018Large-scale molecular changes occur during aging and have many downstream consequences on whole-organism function, such as motor function, learning, and memory. The...
BACKGROUND
Large-scale molecular changes occur during aging and have many downstream consequences on whole-organism function, such as motor function, learning, and memory. The marine mollusk Aplysia californica can be used to study transcriptional changes that occur with age in identified neurons of the brain, because its simplified nervous system allows for more direct correlations between molecular changes, physiological changes, and their phenotypic outcomes. Behavioral deficits in the tail-withdrawal reflex of aged animals have been correlated with reduced excitation in sensory neurons that control the reflex. RNASeq was used to investigate whole-transcriptome changes in tail-withdrawal sensory neurons of sexually mature and aged Aplysia to correlate transcriptional changes with reduced behavioral and physiological responses.
RESULTS
Paired-end sequencing resulted in 210 million reads used for differential expression analysis. Aging significantly altered expression of 1202 transcripts in sensory neurons underlying the tail-withdrawal reflex, with an approximately equal number of these genes up- and down regulated with age. Despite overall bidirectionality of expression changes, > 80% of ion channel genes that were differentially expressed had decreased expression with age. In particular, several voltage-gated K and Ca channels were down regulated. This marked decrease in ion channel expression may play an important role in previously observed declines in aged sensory neuron excitability. We also observed decreased expression of genes and pathways involved in learning and memory. Genes involved in the stress response showed increased expression in aged Aplysia neurons.
CONCLUSIONS
Significantly altered expression of many genes between sexually mature and aged Aplysia suggests large molecular changes that may impact neuronal function. Decreased ion channel mRNA observed could mean fewer receptors present in aged neurons, resulting in reduced excitability of PVC sensory neurons, ultimately leading to reduced tail-withdrawal reflex observed in aged Aplysia. Significant changes in other genes and pathways, such as stress response and learning and memory, have previously been shown to occur with age in many vertebrate organisms. This suggests that some effects of aging are common across many animal phyla.
Topics: Aging; Animals; Aplysia; Down-Regulation; Gene Ontology; Ion Channels; Principal Component Analysis; RNA; Sensory Receptor Cells; Sequence Analysis, RNA; Signal Transduction; Transcriptome; Up-Regulation
PubMed: 29996779
DOI: 10.1186/s12864-018-4909-1 -
The Journal of Biological Chemistry Oct 2018l- to d-residue isomerization is a post-translational modification (PTM) present in neuropeptides, peptide hormones, and peptide toxins from several animals. In most...
l- to d-residue isomerization is a post-translational modification (PTM) present in neuropeptides, peptide hormones, and peptide toxins from several animals. In most cases, the d-residue is critical for the biological function of the resulting d-amino acid-containing peptide (DAACP). Here, we provide an example in native neuropeptides in which the DAACP and its all-l-amino acid epimer are both active at their newly identified receptor and at a neuronal target associated with feeding behavior. On the basis of sequence similarity to a known DAACP from cone snail venom, we hypothesized that allatotropin-related peptide (ATRP), a neuropeptide from the neuroscience model organism , may form multiple diastereomers in the central nervous system. We determined that ATRP exists as a d-amino acid-containing peptide (d2-ATRP) and identified a specific G protein-coupled receptor as an ATRP receptor. Interestingly, unlike many previously reported DAACPs and their all-l-residue analogs, both l-ATRP and d2-ATRP were potent agonists of this receptor and active in electrophysiological experiments. Finally, d2-ATRP was much more stable than its all-l-residue counterpart in plasma, suggesting that in the case of ATRP, the primary role of the l- to d-residue isomerization may be to protect this peptide from aminopeptidase activity in the extracellular space. Our results indicate that l- to d-residue isomerization can occur even in an all-l-residue peptide with a known biological activity and that in some cases, this PTM may help modulate peptide signal lifetime in the extracellular space rather than activity at the cognate receptor.
Topics: Amino Acids; Animals; Aplysia; Insect Hormones; Neurons; Neuropeptides; Peptide Fragments; Protein Processing, Post-Translational; Receptors, G-Protein-Coupled; Stereoisomerism
PubMed: 30194283
DOI: 10.1074/jbc.RA118.004367