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      Dopamine receptor DOP-4 modulates habituation to repetitive photoactivation of a C. elegans polymodal nociceptor

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          Abstract

          Habituation is a highly conserved phenomenon that remains poorly understood at the molecular level. Invertebrate model systems, like Caenorhabditis elegans, can be a powerful tool for investigating this fundamental process. Here we established a high-throughput learning assay that used real-time computer vision software for behavioral tracking and optogenetics for stimulation of the C. elegans polymodal nociceptor, ASH. Photoactivation of ASH with ChR2 elicited backward locomotion and repetitive stimulation altered aspects of the response in a manner consistent with habituation. Recording photocurrents in ASH, we observed no evidence for light adaptation of ChR2. Furthermore, we ruled out fatigue by demonstrating that sensory input from the touch cells could dishabituate the ASH avoidance circuit. Food and dopamine signaling slowed habituation downstream from ASH excitation via D1-like dopamine receptor, DOP-4. This assay allows for large-scale genetic and drug screens investigating mechanisms of nociception modulation.

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          Most cited references55

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          THE GENETICS OF CAENORHABDITIS ELEGANS

          Methods are described for the isolation, complementation and mapping of mutants of Caenorhabditis elegans, a small free-living nematode worm. About 300 EMS-induced mutants affecting behavior and morphology have been characterized and about one hundred genes have been defined. Mutations in 77 of these alter the movement of the animal. Estimates of the induced mutation frequency of both the visible mutants and X chromosome lethals suggests that, just as in Drosophila, the genetic units in C.elegans are large.
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            Efficient gene transfer in C.elegans: extrachromosomal maintenance and integration of transforming sequences.

            We describe a dominant behavioral marker, rol-6(su-1006), and an efficient microinjection procedure which facilitate the recovery of Caenorhabditis elegans transformants. We use these tools to study the mechanism of C.elegans DNA transformation. By injecting mixtures of genetically marked DNA molecules, we show that large extrachromosomal arrays assemble directly from the injected molecules and that homologous recombination drives array assembly. Appropriately placed double-strand breaks stimulated homologous recombination during array formation. Our data indicate that the size of the assembled transgenic structures determines whether or not they will be maintained extrachromosomally or lost. We show that low copy number extrachromosomal transformation can be achieved by adjusting the relative concentration of DNA molecules in the injection mixture. Integration of the injected DNA, though relatively rare, was reproducibly achieved when single-stranded oligonucleotide was co-injected with the double-stranded DNA.
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              Single-copy insertion of transgenes in Caenorhabditis elegans.

              At present, transgenes in Caenorhabditis elegans are generated by injecting DNA into the germline. The DNA assembles into a semistable extrachromosomal array composed of many copies of injected DNA. These transgenes are typically overexpressed in somatic cells and silenced in the germline. We have developed a method that inserts a single copy of a transgene into a defined site. Mobilization of a Mos1 transposon generates a double-strand break in noncoding DNA. The break is repaired by copying DNA from an extrachromosomal template into the chromosomal site. Homozygous single-copy insertions can be obtained in less than 2 weeks by injecting approximately 20 worms. We have successfully inserted transgenes as long as 9 kb and verified that single copies are inserted at the targeted site. Single-copy transgenes are expressed at endogenous levels and can be expressed in the female and male germlines.
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                Author and article information

                Journal
                Learn Mem
                Learn Mem
                learnmem
                Learning & Memory
                Cold Spring Harbor Laboratory Press
                1072-0502
                1549-5485
                October 2016
                : 23
                : 10
                : 495-503
                Affiliations
                [1 ]DM Centre for Brain Health, University of British Columbia, Vancouver V6T 2B5, Canada
                [2 ]Department of Psychology, University of British Columbia, Vancouver V6T 1Z4, Canada
                [3 ]Institute of Neuroscience, University of Oregon, Eugene, Oregon 97403, USA
                Author notes
                [4]

                These authors contributed equally to this work.

                Corresponding author: crankin@ 123456psych.ubc.ca
                Article
                ArdielLM041830
                10.1101/lm.041830.116
                5026203
                27634141
                13b38225-c21c-48a5-8e79-96d3c2e7b9ef
                © 2016 Ardiel et al.; Published by Cold Spring Harbor Laboratory Press

                This article is distributed exclusively by Cold Spring Harbor Laboratory Press for the first 12 months after the full-issue publication date (see http://learnmem.cshlp.org/site/misc/terms.xhtml). After 12 months, it is available under a Creative Commons License (Attribution-NonCommercial 4.0 International), as described at http://creativecommons.org/licenses/by-nc/4.0/.

                History
                : 26 January 2016
                : 27 April 2016
                Funding
                Funded by: NIH Office of Research Infrastructure Programs
                Award ID: P40 OD010440
                Funded by: NSERC CGSD3
                Funded by: NSERC RGPIN
                Award ID: 1222216-13
                Categories
                Research

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