Project description:To explore the role of both Aplysia cell adhesion molecule (ApCAM) and activity of specific protein kinase C (PKC) isoforms in the initial formation of sensory neuron synapses with specific postsynaptic targets (L7 but not L11), we examined presynaptic growth, initial synapse formation, and the expression of the presynaptic neuropeptide sensorin following cell-specific reduction of ApCAM or of a novel PKC activity. Synapse formation between sensory neurons and L7 begins by 3 h after plating and is accompanied by a rapid accumulation of a novel PKC to sites of synaptic interaction. Reducing ApCAM expression specifically from the surface of L7 blocks presynaptic growth and initial synapse formation, target-induced increase of sensorin in sensory neuron cell bodies and the rapid accumulation of the novel PKC to sites of interaction. Selective blockade of the novel PKC activity in L7, but not in sensory neurons, with injection of a dominant negative construct that interferes with the novel PKC activity, produces the same actions as downregulating ApCAM; blockade of presynaptic growth and initial synapse formation, and the target-induced increase of sensorin in sensory neuron cell bodies. The results indicate that signals initiated by postsynaptic cell adhesion molecule ApCAM coupled with the activation of a novel PKC in the appropriate postsynaptic neuron produce the retrograde signals required for presynaptic growth associated with initial synapse formation, and the target-induced expression of a presynaptic neuropeptide critical for synapse maturation.
Project description:To characterize sleep in the marine mollusk, Aplysia californica.Animal behavior and activity were assessed using video recordings to measure activity, resting posture, resting place preference, and behavior after rest deprivation. Latencies for behavioral responses were measured for appetitive and aversive stimuli for animals in the wake and rest states.Circadian research laboratory for Aplysia.A. californica from the Pacific Ocean.N/A.Aplysia rest almost exclusively during the night in a semi-contracted body position with preferential resting locations in the upper corners of their tank. Resting animals demonstrate longer latencies in head orientation and biting in response to a seaweed stimulus and less frequent escape response steps following an aversive salt stimulus applied to the tail compared to awake animals at the same time point. Aplysia exhibit rebound rest the day following rest deprivation during the night, but not after similar handling stimulation during the day.Resting behavior in Aplysia fulfills all invertebrate characteristics of sleep including: (1) a specific sleep body posture, (2) preferred resting location, (3) reversible behavioral quiescence, (4) elevated arousal thresholds for sensory stimuli during sleep, and (5) compensatory sleep rebound after sleep deprivation.
Project description:Neuropeptides are ancient signaling molecules that are involved in many aspects of organism homeostasis and function. Urotensin II (UII), a peptide with a range of hormonal functions, previously has been reported exclusively in vertebrates. Here, we provide the first direct evidence that UII-like peptides are also present in an invertebrate, specifically, the marine mollusk Aplysia californica. The presence of UII in the central nervous system (CNS) of Aplysia implies a more ancient gene lineage than vertebrates. Using representational difference analysis, we identified an mRNA of a protein precursor that encodes a predicted neuropeptide, we named Aplysia urotensin II (apUII), with a sequence and structural similarity to vertebrate UII. With in-situ hybridization and immunohistochemistry, we mapped the expression of apUII mRNA and its prohormone in the CNS and localized apUII-like immunoreactivity to buccal sensory neurons and cerebral A-cluster neurons. Mass spectrometry performed on individual isolated neurons, and tandem mass spectrometry on fractionated peptide extracts, allowed us to define the posttranslational processing of the apUII neuropeptide precursor and confirm the highly conserved cyclic nature of the mature neuropeptide apUII. Electrophysiological analysis of the central effects of a synthetic apUII suggests it plays a role in satiety and/or aversive signaling in feeding behaviors. Finding the homologue of vertebrate UII in the numerically small CNS of an invertebrate animal model is important for gaining insights into the molecular mechanisms and pathways mediating the bioactivity of UII in the higher metazoan.
Project description:The simplified nervous system of Aplysia californica (Aplysia) allows for detailed studies of physiological and molecular changes in small sets of neurons. Sensory neurons of the biting and tail withdrawal reflexes are glutamatergic and show reduced L-Glutamate current density in aged animals, making them a good candidate to study age-related changes in glutamatergic responses. To examine if changes in ionotropic L-Glu receptor (iGluR) transcription underlie reduced physiology, mRNA expression of iGluR was quantified in two sensory neuron clusters of two cohorts of Aplysia at both sexual maturity (~8 months) and advanced age (~12 months). Sensory neuron aging resulted in a significant overall decrease in expression of iGluR subunits in both sensory neuron clusters and cohorts. Although the individual subunits differentially expressed varied between sensory neuron clusters and different cohorts of animals, all differentially expressed subunits were downregulated, with no subunits showing significantly increased expression with age. Overall declines in transcript expression suggest that age-related declines in L-Glu responsiveness in Aplysia sensory neurons could be linked to overall declines in iGluR expression, rather than dysregulation of specific subunits. In both sensory neuron clusters tested the N-methyl-D-aspartate receptor subtype was expressed at significantly greater levels than other iGluR subtypes, suggesting an in vivo role for NMDAR-like receptors in Aplysia sensory neurons.
Project description:The marine mollusc Aplysia is a well established experimental system for cellular and systems neuroscience because of the relatively simple organization of its nervous system and the presence within it of the largest nerve cells in the animal kingdom, many of which are uniquely identifiable in every member of the species. Until now, molecular analyses of Aplysia have been seriously handicapped by lack of adequate genomic information, with only 200 sequences publicly available when this project was initiated in 2003. By sequencing cDNA libraries from the central nervous system, we have identified over 175,000 ESTs (expressed sequence tags), of which 19,814 are unique neuronal gene products and 9469 has been annotated. Through comparison with the complete genomic data available for Drosophila and C. elegans, we estimate that we have sequence information for approximately 50-70% of the total transcriptome of the Aplysia nervous system. We also identified 9,223 unique gene products in a modulatory serotonergic cell and about 1,000 unique gene products from its processes. Using gene expression oligoarrays constructed using the Aplysia EST database we also have characterized the transcript profile of sensory and motor neurons. In addition to increasing the amount of publicly available gene sequences of Aplysia by two orders of magnitude, this collection of transcripts is distinctive from a comparative biology point of view. It represents the largest database available for any member of the Lophotrochozoa clade of the animal kingdom. These molecular resources should allow the detailed study of the genomics of identified cells and circuits and provide in Aplysia a much needed bridge between genes, behavior, and learning. This SuperSeries is composed of the SubSeries listed below. Overall design: Refer to individual Series.
Project description: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 Ca2+ 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.