Invertebrate Neuroscience最新文献

In previous works, we identified a RNA-binding protein in presynaptic terminal of squid neurons, which is likely involved in local mRNA processing. Evidences indicate this strongly basic protein, called p65, is an SDS-stable dimer protein composed of ~ 37 kDa hnRNPA/B-like subunits. The function of p65 in presynaptic regions is not well understood. In this work, we showed p65 and its subunit p37 are concentrated in RNA-enriched regions in synaptosomes. We performed in vitro binding studies with a recombinant protein and showed its propensity to selectively bind actin mRNA at the squid presynaptic terminal. Biochemical analysis using lysed synaptosomes suggested RNA integrity may affect p65 and p37 functions. Mass spectrometry analysis of oligo(dT) pull down indicated squid hnRNPA1, hnRNPA1-like 2, hnRNPA3 and ELAV-like proteins as candidates to interact with p65 and p37 forming a ribonucleoprotein complex, suggesting a role of squid hnRNPA/B-like proteins in site-specific RNA processing.

This study aims to investigate the fine structure of the different cell types in the central brain of Eledone cirrhosa; the organelles in the neurons and the glial cells; the glial hemolymph-brain barrier; the neuro-secretions and the relationships between glial and nerve cells. The brain is surrounded by a non-cellular neurilemma followed by a single layer of perilemmal cells. Ependymal cells, highly prismatic glial cells, astrocytes, oligodendrocytes and epithelial processes were observed. The perikarya of the neurons are filled with slightly oval nuclei with heterochromatin, a strongly tortuous ER, numerous mitochondria and Golgi apparatus with two types of vesicles. In the cellular cortex, glial cells are much less numerous than the neurons and they are located preferably at the border between perikarya and neuropil. Furthermore, they send many branching shoots between the surrounding neuron perikarya and the axons. The glial cytoplasmic matrix appears more electrodense than that of the neurons. Only few ribosomes are attached to the membranes of the ER; the vast majorities are free. In the perikarya of the glial cells, mitochondria, multi-vesicular bodies, various vacuoles and vesicles are present. The essential elements of the hemolymph-brain barrier are the endothelial cells with their tight junctions. The cytoplasm contains various vesicles and mitochondria. However, two other cell types are present, the pericytes and the astrocytes, which are of great importance for the function of the hemolymph-brain barrier. The cell-cell interactions between endothelial cells, pericytes and astrocytes are as close as no other cells.

This meeting report provides an overview of the oral and poster presentations at the first international symposium for invertebrate neuroscience. The contents reflect the contributions of invertebrate neuroscience in addressing fundamental and fascinating challenges in understanding the neural substrates of animal behaviour.

Brugia malayi is a human filarial nematode parasite that causes lymphatic filariasis or 'elephantiasis' a disfiguring neglected tropical disease. This parasite is a more tractable nematode parasite for the experimental study of anthelmintic drugs and has been studied with patch-clamp and RNAi techniques. Unlike in C. elegans however, calcium signaling in B. malayi or other nematode parasites has not been achieved, limiting the studies of the mode of action of anthelmintic drugs. We describe here the development of calcium imaging methods that allow us to characterize changes in cellular calcium in the muscles of B. malayi. This is a powerful technique that can help in elucidating the mode of action of selected anthelmintics. We developed two approaches that allow the recording of calcium signals in the muscles of adult B. malayi: (a) soaking the muscles with Fluo-3AM, promoting large-scale imaging of multiple cells simultaneously and, (b) direct insertion of Fluo-3 using microinjection, providing the possibility of performing dual calcium and electrophysiological recordings. Here, we describe the techniques used to optimize dye entry into the muscle cells and demonstrate that detectable increases in Fluo-3 fluorescence to elevated calcium concentrations can be achieved in B. malayi using both techniques.

Impairment of the dopamine system is the main cause of Parkinson disease (PD). PTEN-induced kinase 1 (PINK1) is possibly involved in pathogenesis of PD. However, its role in dopaminergic neurons has not been fully established yet. In the present investigation, we have used the PINK1 knockout Drosophila model to explore the role of PINK1 in dopaminergic neurons. Electrophysiological and behavioral tests indicated that PINK1 elimination enhances the neural transmission from the presynaptic part of dopaminergic neurons in the protocerebral posterior medial region 3 (PPM3) to PPM3 neurons (which are homologous to those in the substantia nigra in humans). Firing properties of the action potential in PPM3 neurons were also altered in the PINK1 knockout genotypes. Abnormal motor ability was also observed in these PINK1 knockout animals. Our results indicate that knockout of PINK1 could alter both the input and output properties of PPM3 neurons.

Tricaine mesylate, also known as MS-222, was investigated to characterize its effects on sensory neurons, synaptic transmission at the neuromuscular junction, and heart rate in invertebrates. Three species were examined: Drosophila melanogaster, blue crab (Callinectes sapidus), and red swamp crayfish (Procambarus clarkii). Intracellular measures of action potentials in motor neurons of the crayfish demonstrated that MS-222 dampened the amplitude, suggesting that voltage-gated Na + channels are blocked by MS-222. This is likely the mechanism behind the reduced activity measured in sensory neurons and depressed synaptic transmission in all three species as well as reduced cardiac function in the larval Drosophila. To address public access to data, a group effort was used for analysis of given data sets, blind to the experimental design, to gauge analytical accuracy. The determination of a threshold in analysis for measuring extracellular recorded sensory events is critical and is not easily performed with commercial software.

Modelling of human aging, age-related memory loss, and neurodegenerative diseases has developed into a progressive area in invertebrate neuroscience. Gold standard molluscan neuroscience models such as the sea hare (Aplysia californica) and the great pond snail (Lymnaea stagnalis) have proven to be attractive alternatives for studying these processes. Until now, A. californica has been the workhorse due to the enormous set of publicly available transcriptome and genome data. However, with growing sequence data, L. stagnalis has started to catch up with A. californica in this respect. To contribute to this and inspire researchers to use molluscan species for modelling normal biological aging and/or neurodegenerative diseases, we sequenced the whole transcriptome of the central nervous system of L. stagnalis and screened for the evolutionary conserved homolog sequences involved in aging and neurodegenerative/other diseases. Several relevant molecules were identified, including for example gelsolin, presenilin, huntingtin, Parkinson disease protein 7/Protein deglycase DJ-1, and amyloid precursor protein, thus providing a stable genetic background for L. stagnalis in this field. Our study supports the notion that molluscan species are highly suitable for studying molecular, cellular, and circuit mechanisms of the mentioned neurophysiological and neuropathological processes.

Conditioned taste aversion (CTA) in the freshwater pulmonate Lymnaea stagnalis can be formed by presenting ten pairings of sucrose as the conditioned stimulus (CS) and KCl as the unconditioned stimulus (US). The CTA is consolidated to long-term memory (LTM) lasting longer than a month. In the present study, we examined the time course of protein synthesis-dependent period during the consolidation of Lymnaea CTA to LTM by pharmacological inhibition of transcription or translation. The robustness for CTA-LTM was then examined by extinction trials, i.e., repeated presentations of the CS alone. Furthermore, we evaluated the effects of the interstimulus interval (ISI) between the presentation of the CS and US. Our findings indicated that the protein synthesis-dependent period coincides with the CTA training. Repeated presentations of the CS alone after establishment of CTA did not extinguish the CTA, demonstrating the robustness of the CTA-LTM. The ISI ranged from 10 s to a few minutes, and there was no inverted U-shaped function between the ISI and the conditioned response (i.e., suppression of feeding). Thus, CTA still formed even when the presentation of the US was delayed. These features of Lymnaea CTA complement the knowledge for mammalian CTA.

In decapods, dopamine, octopamine, serotonin, and histamine function as locally released/hormonally delivered modulators of physiology/behavior. Although the functional roles played by amines in decapods have been examined extensively, little is known about the identity/diversity of their amine receptors. Recently, a Homarus americanus mixed nervous system transcriptome was used to identify putative neuronal amine receptors in this species. While many receptors were identified, some were fragmentary, and no evidence of splice/other variants was found. Here, the previously predicted proteins were used to search brain- and eyestalk ganglia-specific transcriptomes to assess/compare amine receptor complements in these portions of the lobster nervous system. All previously identified receptors were reidentified from the brain and/or eyestalk ganglia transcriptomes, i.e., dopamine alpha-1, beta-1, and alpha-2 (Homam-DAα2R) receptors, octopamine alpha (Homam-OctαR), beta-1, beta-2, beta-3, beta-4, and octopamine-tyramine (Homam-OTR-I) receptors, serotonin type-1A, type-1B (Homam-5HTR1B), type-2B, and type-7 receptors; and histamine type-1 (Homam-HA1R), type-2, type-3, and type-4 receptors. For many previously partial proteins, full-length receptors were deduced from brain and/or eyestalk ganglia transcripts, i.e., Homam-DAα2R, Homam-OctαR, Homam-OTR-I, and Homam-5HTR1B. In addition, novel dopamine/ecdysteroid, octopamine alpha-2, and OTR receptors were discovered, the latter, Homam-OTR-II, being a putative paralog of Homam-OTR-I. Finally, evidence for splice/other variants was found for many receptors, including evidence for some being assembly-specific, e.g., a brain-specific Homam-OTR-I variant and an eyestalk ganglia-specific Homam-HA1R variant. To increase confidence in the transcriptome-derived sequences, a subset of receptors was cloned using RT-PCR. These data complement/augment those reported previously, providing a more complete picture of amine receptor complement/diversity in the lobster nervous system.

The antenna is a key sensory organ in insects. Factors which pattern its epithelium and the spacing of sensillae will play an important role in shaping its contribution to adaptive behavior. The antenna of the grasshopper S. gregaria has three major articulations: scape, pedicel, and flagellum. During postembryonic development, the flagellum lengthens as segments (so-called meristal annuli) are added at each molt. However, the five most apical annuli do not subdivide; thus, their epithelial domains must already be defined during embryogenesis. We investigated epithelial compartmentalization and its relationship to the developing primordial nervous system of the antenna by simultaneous immunolabeling against the epithelial cell surface molecule Lachesin, against neuron-specific horseradish peroxidase, and against the mitosis marker phospho-histone 3. We found that Lachesin is initially expressed in a highly ordered pattern of "rings" and a "sock" in the apical antennal epithelium of the early embryo. These expression domains appear in a stereotypic order and prefigure later articulations. Proliferative cells segregate into these developing domains and pioneer- and sensory-cell precursors were molecularly identified. Our study allows pioneer neurons, guidepost cells, and the earliest sensory cell clusters of the primordial nervous system to be allocated to their respective epithelial domain. As the apical-most five domains remain stable through subsequent development, lengthening of the flagellum must originate from more basal regions and is likely to be under the control of factors homologous to those which regulate boundary and joint formation in the antenna of Drosophila.