title: Functional analysis of Torpedo californica nicotinic acetylcholine receptors in multiple activation states by SSM-based electrophysiology.
abstract: Organophosphorus compounds (OPC), i.e. nerve agents or pesticides, are highly toxic due to their strong inhibition potency against acetylcholinesterase (AChE). Inhibited AChE results in accumulation of acetylcholine in the synaptic cleft and thus the desensitisation of the nicotinic acetylcholine receptor (nAChR) in the postsynaptic membrane is provoked. Direct targeting of nAChR to reduce receptor desensitisation might be an alternative therapeutic approach. For drug discovery, functional properties of potent therapeutic candidates need to be investigated in addition to affinity properties. Solid supported membrane (SSM)-based electrophysiology is useful for functional characterisation of ligand-gated ion channels like nAChRs, as charge translocations via capacitive coupling of the supporting membrane can be measured. By varying the agonist (carbamoylcholine) concentration, different functional states of the nAChR were initiated. Using plasma membrane preparations obtained from Torpedo californica electric organ, functional properties of selected nAChR ligands and non-oxime bispyridinium compounds were investigated. Depending on overall-size, the bispyridinium compounds enhanced or inhibited cholinergic signals induced by 100 μM carbamoylcholine. Applying excessive concentrations of the agonist carbamoylcholine provoked desensitisation of the nAChRs, whereas addition of bispyridinium compounds bearing short alkyl linkers exhibited functional recovery of previously desensitised nAChRs. The results suggest that these non-oxime bispyridinium compounds possibly interacted with nAChR subtypes in a manner of a positive allosteric modulator (PAM). The described newly developed functional assay is a valuable tool for the assessment of functional properties of potential compounds such as nAChR modulating ligands, which might be a promising approach in the therapeutically treatment of OPC-poisonings.
authors: Bundeswehr Institute of Pharmacology and Toxicology, Neuherbergstraße 11, 80937 Munich, Germany; Supervisory Agency for Public Law Tasks of the Bundeswehr Medical Service South, Dachauer Straße 128, 80637 Munich, Germany. Electronic address: KarinNiessen@bundeswehr.org.
authors: K V
authors: KV
authors: Niessen
authors: Bundeswehr Institute of Pharmacology and Toxicology, Neuherbergstraße 11, 80937 Munich, Germany.
authors: S
authors: S
authors: Muschik
authors: Bundeswehr Institute of Pharmacology and Toxicology, Neuherbergstraße 11, 80937 Munich, Germany.
authors: F
authors: F
authors: Langguth
authors: Department of Pharmacy-Center of Drug Research, Ludwig-Maximilians-University Munich, Butenandtstr. 7, 81377 Munich, Germany.
authors: S
authors: S
authors: Rappenglück
authors: Bundeswehr Institute of Pharmacology and Toxicology, Neuherbergstraße 11, 80937 Munich, Germany.
authors: T
authors: T
authors: Seeger
authors: Bundeswehr Institute of Pharmacology and Toxicology, Neuherbergstraße 11, 80937 Munich, Germany.
authors: H
authors: H
authors: Thiermann
authors: Bundeswehr Institute of Pharmacology and Toxicology, Neuherbergstraße 11, 80937 Munich, Germany.
authors: F
authors: F
authors: Worek
mesh_terms: D000110:Acetylcholinesterase / Q000378:metabolism; D000818:Animals; D002462:Cell Membrane / Q000378:metabolism; D004557:Electric Organ / Q000378:metabolism; D055724:Electrophysiological Phenomena; D004594:Electrophysiology / Q000379:methods*; D029941:Fish Proteins / Q000378:metabolism*; D009943:Organophosphorus Compounds / Q000633:toxicity; D011978:Receptors, Nicotinic / Q000378:metabolism*; D014101:Torpedo / Q000378:metabolism*
chemical_list: D029941:Fish Proteins; D009943:Organophosphorus Compounds; D011978:Receptors, Nicotinic; D000110:Acetylcholinesterase
keywords: Activation states; Functional measurement; Nicotinic acetylcholine receptor; Solid supported membranes; Surface electronic event reader (SURFE(2)R); Torpedo californica