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Immunocytochemical localization of a putative strychnine-sensitive glycine receptor in Hydra vulgaris

  • Linda A. HufnagelEmail author
  • Paola Pierobon
  • Gabriele Kass-SimonEmail author
Regular Article
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Abstract

Previous biochemical studies have identified strychnine-sensitive glycine receptors in membrane preparations of Hydra vulgaris (Cnidaria: Hydrozoa). Electrophysiological and behavioral evidence has shown that these receptors play a role in modulating pacemaker activity and feeding behavior. Here, we present our genomic analysis that revealed hydra proteins having strong homology with the strychnine-binding region of the human receptor protein, GlyRα1. We further present immunocytochemical evidence for the specific labeling of cell and tissue preparations of hydra by a commercially available polyclonal anti-GlyRα1 antibody, selected through our genomic analysis. Tissue pieces and cell macerates from the upper and lower thirds of the body and ablated tentacles were double-labeled with this antibody and with an antibody specific for α-tubulin, to identify the glycine receptors and microtubules, respectively. Extensive receptor labeling was evident on the membranes, cell bodies and myonemes of endodermal and ectodermal epithelial cells, cell bodies and neurites of nerve cells, cnidocytes and interstitial cells. Labeling of the membranes of epithelial cells frequently corresponded to conspicuous varicosities (presumptive presynaptic sites) in the associated nerve net. Our findings support the idea that glycine receptors form an integral part of the nerve and effector systems that control hydra behavior.

Keywords

Amino acid transmitters GABA Cnidocytes Nerve net Interstitial cells 

Notes

Acknowledgements

This research is based upon work conducted using the Rhode Island Genomics and Sequencing Center, which was supported in part by National Science Foundation EPSCoR grants. We thank Kathy Su for her help with immunochemical preparations.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Supplementary material

441_2019_3011_Fig18_ESM.png (7.8 mb)
Fig. S1

EMBOSS Needle pair-wise alignment of hydra protein #1 (XP_012560050) and #2 (XP_012560049), using default settings. Proteins #1 and #2 are identical, beginning with amino acid L5 in protein #1. Note that the GY motif necessary for strychnine binding lies far into the sequence of both proteins. Vertical lines = identity,: = strong functional similarity, . = functional similarity (PNG 7980 kb)

441_2019_3011_MOESM1_ESM.tif (6.2 mb)
High Resolution Image (TIF 6383 kb)
441_2019_3011_Fig19_ESM.png (11.5 mb)
Fig. S2

EMBOSS Needle pair-wise alignment of the human GlyRα1 protein (AAH 74980.1) and hydra protein #1, using default settings. Note the many regions of strong homology, including the predicted binding region of the antibody (Q170 to M221 in the hydra protein), containing groups of identical amino acids, together with amino acids of similar functionality. Note the shared presence of the GY sequence (Gly160, Tyr161, in the human protein in literature, and Gly188, Tyr189 in the hydra protein in the figure), requisite in the human protein for strychnine binding, within the beginning of a five-amino acid region of very strong homology (SFGYT in the human protein, overlined). Vertical lines = identity,: = strong functional similarity, . = functional similarity (PNG 11737 kb)

441_2019_3011_MOESM2_ESM.tif (9.6 mb)
High Resolution Image (TIF 9789 kb)
441_2019_3011_Fig20_ESM.png (15.9 mb)
Fig. S3

CLUSTAL Omega multiple sequence alignment of the human GlyRα1 protein (AAH 74980.1) and hydra proteins #1 (XP_012560050), #2 (XP_012560049), #9 (XP_012556838.1) and #18 (XP_012555765.1). Note that all proteins share the GY motif within a large region of strong homology (overlined). * = identity,: = strong functional similarity, . = functional similarity (PNG 16286 kb)

441_2019_3011_MOESM3_ESM.tif (13.7 mb)
High Resolution Image (TIF 14050 kb)

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Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Department of Cell and Molecular Biology & Interdisciplinary Neurosciences ProgramUniversity of Rhode IslandKingstonUSA
  2. 2.Institute of Applied Sciences and Intelligent Systems E. Caianiello, CNRNaplesItaly
  3. 3.Department of Biological Sciences & Interdisciplinary Neurosciences ProgramUniversity of Rhode IslandKingstonUSA

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