In vitro screening strategies for nicotinic receptor ligands

Dunlop, John; Roncarati, Renza; Jow, Brian; Bothmann, Hendrick; Lock, Tim; Kowal, Dianne; Bowlby, Mark R.; Terstappen, Georg C.

doi:10.1016/j.bcp.2007.07.006

Cited by 45 publications

(45 citation statements)

References 44 publications

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“…Ca 2؉ Flux and Membrane Potential Measurements with an FLIPR System. The following recombinant cell lines were used as specific sources of receptors: GH4C1 cells stably transfected with pCEP4/rat ␣7 nAChR as described previously (Dunlop et al, 2007); HEK293 cell lines stably expressing human ␣4␤2 nAChR (obtained from J. Lindstrom, The University of Pennsylvania, Philadelphia, PA) or 5-hydroxytryptamine (5-HT) 3A receptors (Dunlop et al, 2007). Native neuroblastoma SH-SY5Y cells were used as source of human ganglionic nAChRs (␣3), and TE671 rhabdomyosarcoma cells were used as endogenous source of muscle ␣1␤1␦␥ receptors.…”

Section: Methodsmentioning

confidence: 99%

Procognitive and Neuroprotective Activity of a Novel α7 Nicotinic Acetylcholine Receptor Agonist for Treatment of Neurodegenerative and Cognitive Disorders

Roncarati¹,

Scali²,

Comery³

et al. 2009

J Pharmacol Exp Ther

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The ␣7 nicotinic acetylcholine receptor (nAChR) is a promising target for treatment of cognitive dysfunction associated with Alzheimer's disease and schizophrenia. Here, we report the pharmacological properties of 5-morpholin-4-yl-pentanoic acid (4-pyridin-3-yl-phenyl)-amide [SEN12333 (WAY-317538)], a novel selective agonist of ␣7 nAChR. SEN12333 shows high affinity for the rat ␣7 receptor expressed in GH4C1 cells (K i ϭ 260 nM) and acts as full agonist in functional Ca 2ϩ flux studies (EC 50 ϭ 1.6 M). In whole-cell patch-clamp recordings, SEN12333 activated peak currents and maximal total charges similar to acetylcholine (EC 50 ϭ 12 M). The compound did not show agonist activity at other nicotinic receptors tested and acted as a weak antagonist at ␣3-containing receptors. SEN12333 treatment (3 mg/kg i.p.) improved episodic memory in a novel object recognition task in rats in conditions of spontaneous forgetting as well as cognitive disruptions induced via glutamatergic [5H-dibenzo[a,d]cyclohepten-5,10-imine (dizocilpine maleate); MK-801] or cholinergic (scopolamine) mechanisms. This improvement was blocked by the ␣7-selective antagonist methyllycaconitine, indicating that it is mediated by ␣7 activation. SEN12333 also prevented a scopolamine-induced deficit in a passive avoidance task. In models targeting other cognitive domains, including attention and perceptual processing, SEN12333 normalized the apomorphine-induced deficit of prepulse inhibition. Neuroprotection of SEN12333 was demonstrated in quisqualate-lesioned animals in which treatment with SEN12333 (3 mg/kg/day i.p.) resulted in a significant protection of choline acetyltransferase-positive neurons in the lesioned hemisphere. Cumulatively, our results demonstrate that the novel ␣7 nAChR agonist SEN12333 has procognitive and neuroprotective properties, further demonstrating utility of ␣7 agonists for treatment of neurodegenerative and cognitive disorders.The family of nicotinic acetylcholine receptors, which comprises 16 different subunits in human (␣1-7, ␣9 -10, ␤1-4, ␦, ε, and ␥) that can form many functional homo-and heteropentameric receptor ion channel combinations, contributes to cholinergic neurotransmission in the nervous system and at the neuromuscular junction. The ␣7 nicotinic acetylcholine receptors (nAChRs) are rapidly desensitizing ligand-gated ion channels that are abundantly expressed in the cerebral cortex and the hippocampus, a limbic structure intimately linked to attention processing and memory formation (Gotti et al., 2006). In the hippocampus, ␣7 nAChRs are present in interneurons and glutamatergic pyramidal neurons, in which they are localized presynaptically in nerve terminals and postsynaptically in dendritic spines and soma. In line with their localization, ␣7 nAChRs modulate neurotransmitter release and are responsible for direct fast excitatory neuroArticle, publication date, and citation information can be found at

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Section: Methodsmentioning

confidence: 99%

Procognitive and Neuroprotective Activity of a Novel α7 Nicotinic Acetylcholine Receptor Agonist for Treatment of Neurodegenerative and Cognitive Disorders

Roncarati¹,

Scali²,

Comery³

et al. 2009

J Pharmacol Exp Ther

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“…Several automated electrophysiology platforms have now become commercially available for use in higher-throughput drug screening, [3][4][5] with only a handful of publications demonstrating their use for NNR compounds. [6][7][8] These studies have yielded EC 50 data from automated platforms for both α4β2 and α7 nicotinic receptors using the endogenous nicotinic ligand acetylcholine (ACh) that are highly comparable to those obtained with manual patch clamp recordings; however, these platforms cannot truly be considered high throughput.…”

Section: Introductionmentioning

confidence: 99%

“…Furthermore, the allosteric inhibition by noncompetitive antagonists is not dependent on the concentration of agonist, whereas uncompetitive inhibitors become more effective at higher agonist concentrations due to a greater amount of open channels that can be blocked. 10 Because studies using automated electrophysiology platforms have focused only on determining the effects of agonists or competitive antagonists at NNRs, [6][7][8] there have been no published reports demonstrating the effects of open channel blockers using high-throughput electrophysiology-based platforms.…”

Section: Introductionmentioning

confidence: 99%

Validation of a High-Throughput, Automated Electrophysiology Platform for the Screening of Nicotinic Agonists and Antagonists

Graef¹,

Benson²,

Sidach³

et al. 2013

SLAS Discovery

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High-throughput compound screening using electrophysiology-based assays represents an important tool for biomedical research and drug discovery programs. The recent development and availability of devices capable of performing highthroughput electrophysiology-based screening have brought the need to validate these tools by producing data that are consistent with results obtained with conventional electrophysiological methods. In this study, we compared the response properties of hα3β4 and hα4β2 nicotinic receptors to their endogenous ligand acetylcholine (ACh) using three separate electrophysiology platforms: Dynaflow (low-throughput, manual system), PatchXpress 7000A (medium-throughput automated platform), and IonWorks Barracuda (high-throughput automated platform). We found that despite the differences in methodological approaches between these technologies, the EC 50 values from the ACh dose-response curves were consistent between all three platforms. In addition, we have validated the IonWorks Barracuda for both competitive and uncompetitive inhibition assays by using the competitive nicotinic antagonist dihydro-beta-erythroidin (DHβE) and uncompetitive nicotinic antagonist mecamylamine. Furthermore, we have demonstrated the utility of a custom-written algorithm for generating dose-response curves from multiple extrapolated current metrics that allows for discriminating between competitive and uncompetitive inhibition while maintaining high-throughput capacity. This study provides validation of the consistency of results using low-, medium-, and high-throughput electrophysiology platforms and supports their use for screening nicotinic compounds.

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“…The area and peak Y max values were consistent with partial agonism (area/peak: 62%/26% and 73%/28%, respectively). 28 The quinuclidine-containing spiroimidate core was shown to be optimal in earlier series. 18 In order to confirm these observations, several modifications were made to the spirocycle with 7-chloropyrrolo-[2,1-f ][1,2,4]triazine (Table 2).…”

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confidence: 99%

Development of 4-Heteroarylamino-1′-azaspiro[oxazole-5,3′-bicyclo[2.2.2]octanes] as α7 Nicotinic Receptor Agonists

Hill

Fang

King

et al. 2016

ACS Med. Chem. Lett.

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ABSTRACT:We describe the synthesis of quinuclidine-containing spiroimidates and their utility as α7 nicotinic acetylcholine receptor (nAChR) partial agonists. A convergent synthetic route allowed for rapid SAR investigation and provided a diverse set of fused 6,5-heteroaryl analogs. Two potent and selective α7 nAChR partial agonists, (21), were identified. Both agonists improved cognition in a preclinical rodent model of learning and memory. Additionally, 5-HT 3A receptor SAR suggested the presence of a steric site that when engaged led to significant loss of affinity at that receptor. 2.2]octan]-2-amine (20) and (1′S,3′R,4′S)-N-(

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In vitro screening strategies for nicotinic receptor ligands

Cited by 45 publications

References 44 publications

Procognitive and Neuroprotective Activity of a Novel α7 Nicotinic Acetylcholine Receptor Agonist for Treatment of Neurodegenerative and Cognitive Disorders

Procognitive and Neuroprotective Activity of a Novel α7 Nicotinic Acetylcholine Receptor Agonist for Treatment of Neurodegenerative and Cognitive Disorders

Validation of a High-Throughput, Automated Electrophysiology Platform for the Screening of Nicotinic Agonists and Antagonists

Development of 4-Heteroarylamino-1′-azaspiro[oxazole-5,3′-bicyclo[2.2.2]octanes] as α7 Nicotinic Receptor Agonists

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