title: Positive Modulation of the Glycine Receptor by Means of Glycine Receptor-Binding Aptamers.
abstract: According to the gate control theory of pain, the glycine receptors (GlyRs) are putative targets for development of therapeutic analgesics. A possible approach for novel analgesics is to develop a positive modulator of the glycine-activated Cl(-) channels. Unfortunately, there has been limited success in developing drug-like small molecules to study the impact of agonists or positive modulators on GlyRs. Eight RNA aptamers with low nanomolar affinity to GlyRα1 were generated, and their pharmacological properties analyzed. Cytochemistry using fluorescein-labeled aptamers demonstrated GlyRα1-dependent binding to the plasma membrane but also intracellular binding. Using a fluorescent membrane potential assay, we could identify five aptamers to be positive modulators. The positive modulation of one of the aptamers was confirmed by patch-clamp electrophysiology on L(tk) cells expressing GlyRα1 and/or GlyRα1β. This aptamer potentiated whole-cell Cl(-) currents in the presence of low concentrations of glycine. To our knowledge, this is the first demonstration ever of RNA aptamers acting as positive modulators for an ion channel. We believe that these aptamers are unique and valuable tools for further studies of GlyR biology and possibly also as tools for assay development in identifying small-molecule agonists and positive modulators.
authors.affiliation: Department of Neuroscience, AstraZeneca R&D, Södertälje, Sweden and Division of Protein Technology, School of Biotechnology, Royal Institute of Technology, Stockholm, Sweden ndekki@hotmail.com.
authors.forename: Nancy Dekki
authors.initials: ND
authors.lastname: Shalaly
authors.affiliation: Department of Neuroscience, AstraZeneca R&D, Södertälje, Sweden and Lead Discovery Technologies, Merck Serono SA, Geneva, Switzerland.
authors.forename: Eduardo
authors.initials: E
authors.lastname: Aneiros
authors.affiliation: AptaIT GmbH, Goethestrasse 52, Munich, Germany.
authors.forename: Michael
authors.initials: M
authors.lastname: Blank
authors.affiliation: Department of Structural Chemistry Laboratory, AstraZeneca R&D, Mölndal, Sweden.
authors.forename: Johan
authors.initials: J
authors.lastname: Mueller
authors.affiliation: Department of Neuroscience, AstraZeneca R&D, Södertälje, Sweden.
authors.forename: Eva
authors.initials: E
authors.lastname: Nyman
authors.affiliation: AptaIT GmbH, Goethestrasse 52, Munich, Germany.
authors.forename: Michael
authors.initials: M
authors.lastname: Blind
authors.affiliation: Pelago Bioscience, Nobels väg 3, Solna, Sweden.
authors.forename: Michael A
authors.initials: MA
authors.lastname: Dabrowski
authors.affiliation: Department of Neuroscience, AstraZeneca R&D, Södertälje, Sweden.
authors.forename: Christin V
authors.initials: CV
authors.lastname: Andersson
authors.affiliation: Department of Translational Science, Respiratory, AstraZeneca R&D, Mölndal, Sweden and Department of Medicinal Chemistry, Science for Life Laboratory, Biomedical Center, Uppsala University, Uppsala, Sweden.
authors.forename: Kristian
authors.initials: K
authors.lastname: Sandberg
mesh_terms: D000818:Animals; D052157:Aptamers, Nucleotide / Q000378:metabolism / Q000494:pharmacology*; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D003593:Cytoplasm / Q000378:metabolism; D004353:Drug Evaluation, Preclinical; D007700:Kinetics; D008564:Membrane Potentials / Q000187:drug effects; D051379:Mice; D010843:Pichia; D018009:Receptors, Glycine / Q000819:agonists* / Q000378:metabolism; D052156:SELEX Aptamer Technique; D020349:Surface Plasmon Resonance
chemical_list: D052157:Aptamers, Nucleotide; D018009:Receptors, Glycine
keywords: aptamer; fluorescent membrane potential; glycine receptor; patch-clamp electrophysiology; surface plasmon resonance