Old and New Pharmacology: Positive Allosteric Modulation of the α7 Nicotinic Acetylcholine Receptor by the 5-Hydroxytryptamine_2B/C Receptor Antagonist SB-206553 (3,5-Dihydro-5-methyl-N-3-pyridinylbenzo[1,2-b:4,5-b′]di pyrrole-1(2H)-carboxamide)

Abstract

The ␣7 nicotinic acetylcholine receptor (nAChR) has been implicated in Alzheimer's disease and schizophrenia, leading to efforts targeted toward discovering agonists and positive allosteric modulators (PAMs) of this receptor. In a Ca 2ϩ flux fluorometric imaging plate reader assay, SB-206553 (3,5-dihydro -5 -methyl -N -3 -pyridinylbenzo [1, 2 -b : 4, 5 -bЈ] -di pyrrole-1(2H)-carboxamide), a compound known as a 5-hydroxytryptamine 2B/2C receptor antagonist, produced an 8-fold potentiation of the evoked calcium signal in the presence of an EC 20 concentration of nicotine and a corresponding EC 50 of 1.5 M for potentiation of EC 20 nicotine responses in GH4C1 cells expressing the ␣7 receptor. SB-206553 was devoid of direct ␣7 receptor agonist activity and selective against other nicotinic receptors. Confirmation of the PAM activity of SB-206553 on the ␣7 nAChR was obtained in patch-clamp electrophysiological experiments in GH4C1 cells, where it failed to evoke any detectable currents when applied alone, yet dramatically potentiated the currents evoked by an EC 20 (17 M) and EC 100 (124 M) of acetylcholine (ACh). Native nicotinic receptors in CA1 stratum radiatum interneurons of rat hippocampal slices could also be activated by ACh (200 M), an effect that was entirely blocked by the ␣7-selective antagonist methyllycaconitine (MLA). These ACh currents were potentiated by SB-206553, which increased the area of the current response significantly, resulting in a 40-fold enhancement at 100 M. In behavioral experiments in rats, SB-206553 reversed an MK-801 (dizocilpine maleate)-induced deficit in the prepulse inhibition of acoustic startle response, an effect attenuated in the presence of MLA. This latter observation provides further evidence in support of the potential therapeutic utility of ␣7 nAChR PAMs in schizophrenia.Nicotinic acetylcholine receptors (nAChRs) are formed of pentameric combinations of ␣ and non-␣ subunits with a high degree of complexity conferred by 10 different ␣ subunits (␣1-␣10) and seven different non-␣ subunits (␤1-␤4, ␥␦, ε)Article, publication date, and citation information can be found at

Intrinsically Low Open Probability of α7 Nicotinic Acetylcholine Receptors Can Be Overcome by Positive Allosteric Modulation and Serum Factors Leading to the Generation of Excitotoxic Currents at Physiological Temperatures

“…Although PNU-120596 and TQS appear to bind at similar sites in the intrasubunit cavity formed by the four membrane-spanning helices (Young et al, 2008;Gill et al, 2011), there may be alternate protein-PAM interactions, with increased differences in the response to membrane fluidity changes that occur at higher temperature. Although increased temperature had only a modest effect on receptor activation by the allosteric agonist 4-(4-bromophenyl)-3a,4,5,9b-tetrahydro-3H-cyclopenta[c]quinoline-8-sulfonamide (Jindrichova et al, 2012), Sitzia et al, 2011 reported that the potentiating activity of SB-206553, a PAM with properties intermediate to those of the type I and type II PAM classes (Dunlop et al, 2009), was also reduced at 37°C. Although the published data agree that type I PAMs appear to lack in vitro cytotoxicity (at the concentrations of agonists and modulators tested), the data are contradictory regarding the in vitro toxicity of the type II PAM PNU-120596 (Ng et al, 2007;Hu et al, 2009;Dinklo et al, 2011).…”

Understanding the Bases of Function and Modulation of α7 Nicotinic Receptors: Implications for Drug Discovery

“…Thus, when analyzed at the molecular level, potentiation is more complex than initially believed. Moreover, PAMs showing macroscopic intermediate type I/II properties were proposed (Dunlop et al, 2009;Malysz et al, 2009;Dinklo et al, 2011;Sahdeo et al, 2014;Chatzidaki and Millar, 2015). Therefore, the classification of type I and type II appears to be an oversimplification resulting mainly from macroscopic observations, and a more thorough classification may be required.…”

Receptor-Mediated Suppression of Potassium Currents Requires Colocalization within Lipid Rafts

“…Ratiometric determination of intracellular Ca 2ϩ concentration ([Ca 2ϩ ] i ) in individual HEK293 cells was performed using Fura-2-based microfluorimetry. Methods were similar to those described recently by Dunlop et al (2009). In brief, HEK293 cells were plated on 13-mm round glass coverslips and loaded with Fura-2AM (2 M) by incubation for 30 min at 37°C in a standard HEPESbuffered salt solution (HBSS) supplemented with Pluronic acid 0.02% (w/v).…”