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The distribution of cholinergic neurons and their co-localization with FMRFamide, in central and peripheral neurons of the spider Cupiennius salei

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Abstract

The spider Cupiennius salei is a well-established model for investigating information processing in arthropod sensory systems. Immunohistochemistry has shown that several neurotransmitters exist in the C. salei nervous system, including GABA, glutamate, histamine, octopamine and FMRFamide, while electrophysiology has found functional roles for some of these transmitters. There is also evidence that acetylcholine (ACh) is present in some C. salei neurons but information about the distribution of cholinergic neurons in spider nervous systems is limited. Here, we identify C. salei genes that encode enzymes essential for cholinergic transmission: choline ACh transferase (ChAT) and vesicular ACh transporter (VAChT). We used in-situ hybridization with an mRNA probe for C. salei ChAT gene to locate somata of cholinergic neurons in the central nervous system and immunohistochemistry with antisera against ChAT and VAChT to locate these proteins in cholinergic neurons. All three markers labeled similar, mostly small neurons, plus a few mid-sized neurons, in most ganglia. In the subesophageal ganglia, labeled neurons are putative efferent, motor or interneurons but the largest motor and interneurons were unlabeled. Groups of anti-ChAT labeled small neurons also connect the optic neuropils in the spider protocerebrum. Differences in individual cell labeling intensities were common, suggesting a range of ACh expression levels. Double-labeling found a subpopulation of anti-VAChT-labeled central and mechanosensory neurons that were also immunoreactive to antiserum against FMRFamide-like peptides. Our findings suggest that ACh is an important neurotransmitter in the C. salei central and peripheral nervous systems.

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Acknowledgements

The authors thank Stephen Whitefield (Dalhousie University) for help with confocal microscopy. We are grateful to Nicole Skaluba and Morgen Livoli for helpful discussions and suggestions. We also thank Drs. Axel Schmid (University of Vienna), Ian Meinertzhagen (Dalhousie University) and Douglas Facey (Saint Michael’s College) for valuable discussions. We would like to acknowledge support through the office of the Vice President of Academic Affairs of SMC and faculty of the Biology Department with the acquisition of essential supplies. Research reported in this paper was supported by an Institutional Development Award (IDeA) from the National Institute of General Medical Sciences of the National Institutes of Health under grant number P20GM103449 (RF-F and CA). Its contents are solely the responsibility of the authors and do not necessarily represent the official views of NIGMS or NIH. Further support was received from the George I. Alden Trust and the John C Hartnett fund (MR) and the Natural Sciences and Engineering Research Council of Canada (NSERC), through discovery grants RGPIN-2014-05565 to PHT and RGPIN/03712 to ASF.

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Correspondence to Ruth Fabian-Fine.

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Fabian-Fine, R., Anderson, C.M., Roush, M.A. et al. The distribution of cholinergic neurons and their co-localization with FMRFamide, in central and peripheral neurons of the spider Cupiennius salei . Cell Tissue Res 370, 71–88 (2017). https://doi.org/10.1007/s00441-017-2652-6

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Keywords

  • Choline acetyltransferase
  • Vesicular acetylcholine transporter
  • Sensory neurons
  • In situ hybridization
  • Central nervous system