Marine Natural Products Acting on the Acetylcholine-Binding Protein and Nicotinic Receptors: From Computer Modeling to Binding Studies and Electrophysiology

Denis Kudryavtsev,Tatyana Makarieva,Christoph Methfessel,Natalia Utkina,Elena Kryukova,Igor Kasheverov,Valentin Stonik,Elena Santalova,Victor Tsetlin

doi:10.3390/md12041859

Abstract

For a small library of natural products from marine sponges and ascidians, in silico docking to the Lymnaea stagnalis acetylcholine-binding protein (AChBP), a model for the ligand-binding domains of nicotinic acetylcholine receptors (nAChRs), was carried out and the possibility of complex formation was revealed. It was further experimentally confirmed via competition with radioiodinated α-bungarotoxin ([125I]-αBgt) for binding to AChBP of the majority of analyzed compounds. Alkaloids pibocin, varacin and makaluvamines С and G had relatively high affinities (Ki 0.5–1.3 μM). With the muscle-type nAChR from Torpedo californica ray and human neuronal α7 nAChR, heterologously expressed in the GH4C1 cell line, no competition with [125I]-αBgt was detected in four compounds, while the rest showed an inhibition. Makaluvamines (Ki ~ 1.5 μM) were the most active compounds, but only makaluvamine G and crambescidine 359 revealed a weak selectivity towards muscle-type nAChR. Rhizochalin, aglycone of rhizochalin, pibocin, makaluvamine G, monanchocidin, crambescidine 359 and aaptamine showed inhibitory activities in electrophysiology experiments on the mouse muscle and human α7 nAChRs, expressed in Xenopus laevis oocytes. Thus, our results confirm the utility of the modeling studies on AChBPs in a search for natural compounds with cholinergic activity and demonstrate the presence of the latter in the analyzed marine biological sources.

Highlights

IntroductionNicotinic acetylcholine receptors (nAChRs) are among the most comprehensively studied ligand-gated ion channels (see reviews [1,2,3,4])
Nicotinic acetylcholine receptors are among the most comprehensively studied ligand-gated ion channels
The most potent competitor of [125I]-αBgt for the ―classical‖ binding site on both tested receptors was makaluvamine C (5): it showed Ki = 1.30 ± 0.05 μM on muscle-type receptor and 3.6 ± 0.3 μM on human α7 nicotinic acetylcholine receptors (nAChRs). Summarizing these results on the activity of the studied compounds from sponges and ascidians, we ascertain that there are substances with sound cholinergic properties in these marine sources acting on both muscle-type and α7 neuronal nAChRs. Their mode of action and affinities are close to those for a number of well-known cholinergic ligands from alkaloids [41], to different α-conotoxins [42,43,44], some snake venoms components [45,46] and native modulators [47] showing similar micromolar potency to the muscle and α7 nAChR subtypes evaluated in competition with [125I]-αBgt

Summary

Introduction

Nicotinic acetylcholine receptors (nAChRs) are among the most comprehensively studied ligand-gated ion channels (see reviews [1,2,3,4]). They are widespread in fish electric organs and mammalian muscles (so-called muscle type of nAChR), central and peripheral nervous systems (neuronal nAChRs) and some non-neuronal tissues (so-called ―non-neuronal‖ nAChRs), where they perform a multitude of functions from conducting nerve-muscle transmission to participation in the different cognitive processes and regulation of inflammatory response. A huge number of compounds of different chemical nature from various taxa (bacteria, plants, mollusks, chordates) are known to interact with nAChRs. Among them, the most well-known and widely used are α-neurotoxins from snake venoms (see reviews [6,7,8]). The conclusions from computer modeling were verified by efficient interaction of the studied compounds with AChBP revealed by radioligand analysis, as well as by their binding to muscle and α7 neuronal nAChRs tested by radioligand analysis and electrophysiology

Isolation of Individual Compounds

Computer Modeling

Radioligand Assay

Electrophysiology Measurements

Conclusions