ARTICLE IN PRESS. Journal of Neuroscience Methods xxx (2008) xxx xxx. Contents lists available at ScienceDirect. Journal of Neuroscience Methods
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1 Journal of Neuroscience Methods xxx (2008) xxx xxx Contents lists available at ScienceDirect Journal of Neuroscience Methods journal homepage: Characterization of compounds on nicotinic acetylcholine receptor alpha7 channels using higher throughput electrophysiology S. Friis a, C. Mathes a,, M. Sunesen a, M.R. Bowlby b, J. Dunlop a a Sophion Bioscience A/S, 154 Baltorpvej, 2750 Ballerup, Denmark b Discovery Neuroscience, Wyeth Research, CN-8000, Princeton, NJ 08543, USA article info abstract Article history: Received 27 March 2008 Received in revised form 29 September 2008 Accepted 6 October 2008 Keywords: Alpha7 nicotinic acetylcholine receptor Automated patch clamp QPatch Ion channels Acetylcholine Alpha7 nicotinic acetylcholine receptor channels are important ligand-gated ion channels that are fast desensitizing, cation selective and have been implicated in the pathophysiology of schizophrenia and Alzheimer s disease. We report here high quality 7 parallel patch clamp recordings using the QPatch automated patch clamp system. The QPatch patch clamps up to 48 cells in parallel with the same high fidelity as conventional patch clamp. EC 50 and IC 50 values were comparable to values obtained with conventional patch clamp. The EC 50 value for acetylcholine (ACh) on the QPatch with area under the curve (AUC) analysis was 26 M compared to a value of 29 M determined from conventional patch clamp experiments. Sequential additions of ACh can be made with minimal decay of the peak amplitude. The competitive 7 antagonist methyllycaconitine (MLA) blocked currents with an IC 50 value of 0.25 nm which is similar to published IC 50 values for MLA. Finally, two different classes of positive allosteric modulators represented by PNU and NS-1738 elicited characteristic responses, thus allowing accurate characterization of modulation and measurements of potency. These results demonstrate that 7 nicotinic acetylcholine receptor channels can be studied reliably in a higher throughput, parallel manner with the QPatch automated patch clamp system Elsevier B.V. All rights reserved. 1. Introduction The alpha7 nicotinic acetylcholine receptor ( 7 nachr) belongs to a class of pentameric ligand-gated ion channels that is highly permeable to calcium and sodium ions (Gotti et al., 2006b; Lindstrom, 1996). Several lines of experimental and clinical evidence support the potential therapeutic utility of 7 nachr agonists in schizophrenia and Alzheimer s disease (Adler et al., 1998; Freedman et al., 1994; Gotti and Clementi, 2004; Gotti et al., 2006a; Jensen et al., 2005). These include pro-cognitive efficacy and normalization of sensory gating deficits with 7 nachr receptor agonists demonstrated in preclinical animal models (Arendash et al., 1995; Hajos et al., 2005; Kitagawa et al., 2003; Levin et al., 1999; Pichat et al., 2007; Simosky et al., 2001; Stevens et al., 1998), and an enriched expression of central 7 nachr in brain regions implicated in these functions (Lindstrom, 1996). Clinically, the high prevalence of smoking in schizophrenics has been postulated as an attempt to self-medicate, and in a recent small clinical trial in schizophrenics the prototype 7 nachr agonist DMXB- A (also known as GTS-21) normalized the P50 auditory-evoked Corresponding author. address: cma@sophion.com (C. Mathes). potential sensory gating deficit, while also increasing attention as assessed in a cognitive test battery (Olincy et al., 2006). A decreased expression of 7 nachr in post-mortem schizophrenic brain has been documented and genetic linkage studies have identified the promoter region of the 7 nachr gene as a susceptibility locus (Freedman et al., 1995, 2001; Freedman and Leonard, 2001; Olincy et al., 2006). Most recently, 7 nachr positive allosteric modulators (PAMs) have been discovered and also reported to improve cognition and normalize sensory gating deficits in rodents (Gronlien et al., 2007; Hurst et al., 2005; Timmermann et al., 2007). Taken together, these data provide a compelling rationale for the development of 7 nachr agonists and PAMs as novel therapeutic agents in schizophrenia and AD. In order to discover 7 nachr agonists and PAMs, a number of groups have developed and successfully implemented robust cell-based calcium flux assays for high-throughput screening and subsequent lead-optimization (Dunlop et al., 2007). While this approach has proven to be very powerful, ion channel targeted drug discovery efforts require confirmation of compound activity using electrophysiological techniques. Electrophysiology overcomes potential limitations inherent to the use of the highthroughput plate-based methods. These include the indirect index of channel function reported in some cell-based assays, e.g., the calcium flux observed upon agonist activation of 7 nachr receptors /$ see front matter 2008 Elsevier B.V. All rights reserved. doi: /j.jneumeth
2 2 S. Friis et al. / Journal of Neuroscience Methods xxx (2008) xxx xxx expressed in GH4-C1 cells, which has been shown to be attributable to secondary activation of L-type calcium channels expressed in these cells (Feuerbach et al., 2005). In addition, the cell-based assays lack mechanistic insight, while the information rich electrophysiological approach supports assessment of compound effects on channel desensitization kinetics and gating. However, conventional manual patch clamp electrophysiology in mammalian cells does not come without its own limitations, being extremely low-throughput in nature and often representing a significant bottleneck in compound screening. To address this bottleneck for the characterization of 7 nachr agonists and PAMs we have explored the use of the QPatch high-throughput and automated electrophysiology platform. The QPatch performs up to 48 independent patch clamp experiments in parallel in a 16- or 48-channel chip array called the QPlate. Briefly, the QPatch performs ligandgated experiments using either four or eight fluidics pipette tips to apply compounds to the patch clamped cells. The compounds flow via integrated microfluidic pathways in the QPlates to the patch clamped cell where the response is recorded via the built-in and parallel patch clamp amplifiers (Mathes, 2006; Willumsen et al., 2003). The present study provides a validation of this approach in comparison to the use of manual patch clamp electrophysiology in conjunction with the Dynaflow rapid perfusion system for the characterization of 7 nachr agonists, antagonists and PAMs. 2. Methods 2.1. Cells, QPatch, and compounds We used GH4-C1 rat pituitary tumor cells expressing the rat 7 nachr. Forty-eight hours prior to experiments, the cells were incubated with 0.5 mm Na butyrate. The cells were detached from the flask with trypsin and kept in serum free culture medium supplied with 25 mm HEPES and 0.04 mg/ml soybean trypsin inhibitor. In this medium, cells could be kept for several hours prior to and during experiments. The application of pressure for forming gigaseals and the wholecell patch clamp configuration (whole-cell) were established using QPatch Assay Software (Sophion Bioscience). Addition of agonists and test compounds was also set up using the QPatch Assay Software. Compounds were added to the cells with the eight pipettes via the QPlate integrated glass microfluidic pathways. Typically 5 L of ligand was added for 2 s followed by washout (four times 5 L of saline). Test compounds were typically preincubated for 150 s prior to agonist addition. The data was acquired at 5 khz for 5 s. During whole-cell recording, the membrane potential was held at 90 mv. All QPatch data were analyzed using Sophion s QPatch Assay software. Results are presented as mean ± S.E.M. The solutions used: Extracellular (mm): 145 NaCl, 4 KCl, 10 HEPES, 2 CaCl 2,1MgCl 2 Intracellular (mm): 120 KF, 20 KCl, 0.1 EGTA, 10 HEPES, 2 MgCl 2 We used QPlates with an integrated waste container holding 250 L (QPlateXL) making longer and more advanced ligand experiments possible Manual recordings Cells were patch clamped at 60 mv, and data were acquired at 1 khz for 2 s episodes (500 ms bath, 500 ms agonist, 1000 ms washout) with 10 s intervals between episodes at room temperature. ACh (124 M) was used as a positive control and also served as a standard for the normalization of responses. Up to 10 different concentrations of test compound were applied in all assays. 7 nachr agonists produce an inward current and the total charge carried by this current as determined by the area under the curve (AUC) was measured and normalized to ACh. EC 50 and Emax values were then determined from this concentration response curve. Agonist and compound applications were achieved using the Dynaflow perfusion system (DF-48). 3. Results 3.1. Assay performance QPatch recordings using GH4-C1 cells were generally not limited by obtaining gigaohm seals onto the cells. Due to their round morphology the sealing rate with the cells was generally good (85 ± 2%). Cell capacitances between 2 and 4 pf were common, and in some experiments apparent single channel activity has been seen (data not shown). The rate of obtaining the whole-cell configuration was 71 ± 2% with an average whole-cell resistance of 1.6 ± 0.1 G measured just after break in (n = 71), however the final success rate of useful experiments was significantly less. This was partially due to the lack of expression of the channel in some of the cells. The first set of experiments determined that measurable 7 current could be recorded with QPatch. Application of ACh produced a clear 7 current in the GH4-C1 cells. An example of a current trace is shown in Fig. 1. An obvious requirement for drug screening with the QPatch or any system is the possibility to obtain stable responses to repeated additions of the same agonist concentration while avoiding cumulative inactivation or down-regulation. This was tested by examining the 7 signal when exposed to repeated application of a maximally effective (1 mm) ACh concentration. Initially there was a decrease in the current response during these experiments (Fig. 1B). The hypothesis that agonist remained bound to or around the cell after the wash had finished was tested by adding an acetylcholine esterase (4 U/mL) to the washing solution. This treatment was found to stabilize the response to repeated application of ACh (Fig. 1B) Agonist concentration response Full agonism Nicotinic receptor channels can be stimulated with several agonists and two of these have been tested on the QPatch. Acetylcholine as the endogenous agonist is considered to define a full agonist response for this receptor, thus the concentration response relationship was determined with eight different concentrations. The raw traces from one of these experiments are shown in Fig. 2. An EC 50 estimation based on the peak values of the agonistevoked responses was estimated to be >1.0 mm (n = 7; data not shown). This value is right-shifted when compared to data obtained with the Dynaflow system, manual patch-clamp and two electrode voltage clamp, where the measured EC 50 value has been found to be between 0.07 and 0.6 mm (Delbono et al., 1997; Dunlop et al., 2007; Papke, 2006; Virginio et al., 2002). This discrepancy is most likely due to the fact that the exchange time is limiting for a correct estimation of the peak value resulting in a concentration response curve without a proper maximum effect. When employing a net charge or AUC measurement, however, there is very good agreement between the two systems: 26 ± 0.6 M(n = 7) determined on
3 S. Friis et al. / Journal of Neuroscience Methods xxx (2008) xxx xxx 3 Fig. 1. A) Signal detection. An acetylcholine induced 7 response in CH4C1 on QPatch. The rise time (10 90) was found to be 5.7 +/ 0.9 ms in responses to 10 mm ACh (n = 7). B) Evaluation of signal reproducibility. Signal reproducibility evaluation with repeated stimulation of 7 with 5 L 1 mm acetylcholine followed by 4 5 L wash with saline on the same cell. shows the normalized I/T-plot +/ SEM from a typical control experiment (n = 7) were it can be seen that the current typically is 50% reduced in the following stimulation. is a normalized I/T-plot +/ SEM from an experiment were the cell again is stimulated with 5 L 1 mm acetylcholine but washed with 4 5 L saline with 4 U/mL acetylcholine esterase (n = 6). Fig. 2. Concentration response experiment with acetylcholine. A) Raw data from an experiment were the cell was stimulated with increasing concentrations of acetylcholine ( mm). The response to some concentrations has been removed for increased clarity. B) Normalized peak responses +/ SEM as a function of acetylcholine concentration (n = 7). C) Normalized AUC responses +/ SEM as a function of acetylcholine concentration and corresponding Hill fit analysis (n = 7). QPatch (Fig. 2) vs. 29 M on Dynaflow (see Dunlop et al., 2007). The rise time of the current was found to be 5.7 ± 0.9 ms in responses to 10 mm ACh (n = 7). The stability of the concentration response curve and peak amplitudes was also measured with repeated agonist experiments. Two five-point acetylcholine concentration response experiments were performed on the same cell to evaluate if the responses were stable at a wide range of concentrations and over an extended period of time. The reproducibility in this assay was analyzed by comparing the peak current-evoked at each of the five concentrations between the 1st and 2nd application. As shown in Table 1, agonist-evoked responses to ACh were found to be highly reproducible between the 1st and 2nd applications. Table 1 Extended signal stability experiment. Signal reproducibility was evaluated by stimulating a cell to increasing concentrations of acetylcholine ( mm) twice. The table summarizes % change +/ SEM that was seen in response 2 compared to response 1. Acetylcholine ( M) (Peak 1/peak 2) 100 (%), average ± S.E.M. (n =5) ± ± ± ± ± Partial agonism In electrophysiological studies, nicotine can appear as a partial 7 nachr agonist compared to ACh (Dunlop et al., 2007; Papke et al., 2007). Testing cells with a broad range of nicotine concentrations clearly shows that the highest AUC occurred at 111 M with an AUCmax = 0.4 compared to ACh (n =5) (Fig. 3B). Analysis of the concentration response elicited with nicotine resulted in an AUC EC 50 of 2.8 ± 0.1 M (Fig. 3B). It is therefore clear that nicotine is only a partial agonist with a higher affinity than acetylcholine but with a lower efficacy Modulator and antagonist dose response assays Antagonist assay Methyllycaconitine (MLA) potently antagonizes 7 nicotinic receptors. MLA was used to confirm the specificity of the currents induced in the GH4-C1/ 7 cell line. A sequence of 1 mm ACh control applications was applied before the cells were preincubated with the test compound. The cells were preincubated for a specified period of time (150 s) before stimulation with acetylcholine plus the test compound. Application of increasing concentrations of MLA antagonized the ACh activated current, as expected, without changing the shape of the signal (Fig. 4A). The IC 50 as determined by analysis of the AUC was 0.25 ± 0.09 nm (n = 6)(Fig. 4B).
4 4 S. Friis et al. / Journal of Neuroscience Methods xxx (2008) xxx xxx Fig. 3. Concentration response experiment with the partial agonist nicotine. A) Raw data from an experiment were the cell was stimulated with 300 um acetylcholine followed by increasing concentrations of (-)-nicotine ( M). B) Concentration response plot of nicotine invoked 7 responses (AUC) expressed as nicotine AUC response/ AUC response to 300 um acetylcholine (n = 5). Fig. 4. Concentration response experiment with MLA. A) Raw data from an experiment were the cell was stimulated with 1 mm acetylcholine and preincubated for 150 sec with increasing concentrations of MLA ( nm). B) Normalized recorded AUC +/ SEM fitted to the Hill Equation (n = 6) Positive allosteric modulators (PAM) A number of compounds modulate the nicotinic 7 receptor. The action of these can be divided into two groups: type I PAMs increase peak amplitudes with little or no effect on the desensitization kinetics and type II PAMs slow desensitization kinetics (Gronlien et al., 2007). Two modulators, one from each group, were tested on the QPatch: one that has a dramatic effect on the desensitization of the channel, PNU , first characterized by Hurst et al. (2005) and another with peak enhancing properties, NS1738, described by Timmermann et al. (2007). Fig. 5. Concentration response experiment with PNU A) Raw data from a typical experiment where a cell was continuously stimulated with 300 M and preincubated for 150 sec. with increasing concentrations of PNU ( M). One control sweep has been magnified due to the dramatic increase in amplitude in the presence of PNU B) Normalized recorded AUC +/ SEM fitted to the Hill Equation (n = 6).
5 S. Friis et al. / Journal of Neuroscience Methods xxx (2008) xxx xxx 5 Fig. 6. Concentration response experiment with NS A) Raw data from a typical experiment were a cell was stimulated with 300 M of acetylcholine and preincubated with increasing concentrations of NS1738 ( M). B) Normalized recorded AUC +/ SEM fitted to the Hill Equation (n = 5). The PAM assay was similar to the antagonist assay, with a number of controls run before the test compound was added. In this case, 300 M ACh was used followed by preincubation with the test compound for 150 s. Current traces with and without PNU are shown in Fig. 5A; note the extreme effect on the current kinetics, where the desensitization was completely removed by the compound at 30 M. The EC 50 for PNU was determined to be 4.9 ± 1.1 M (n =6)(Fig. 5B). NS1738 is a PAM that recently has been developed by NeuroSearch (Ballerup, DK) and has been shown to increase the peak of the acetylcholine induced response (Timmermann et al., 2007). This compound was also tested on the QPatch and the resulting currents show the expected increase in peak current with negligible effect on the kinetics (Fig. 6A). The analysis (Fig. 6B) determined an EC 50 of 12.5 ± 1.5 M (n = 5). 4. Discussion A number of different compounds including agonists, competitive antagonists and modulators were tested against the nicotinic 7 ion channel in this study in order to determine if the QPatch automated patch-clamp system is capable of resolving fast ligandgated ion channels. Testing with a broad range of ACh concentrations showed a right-shift between using the resulting current peak amplitude and charge flux. This is a commonly observed phenomenon that has led to the well-accepted view that a measurement of AUC is a more accurate and relevant measure for analyzing agonists at this receptor. This approach is also validated for the analysis of novel agonists, as the receptor appears to desensitize in an agonist concentrationdependent fashion, in contrast to many other receptors (Papke et al., 2000; Papke and Porter Papke, 2002). Using AUC rather than peak has also been shown previously to be the most effective way of standardizing results between different recording methods (Papke et al., 2000; Papke and Porter Papke, 2002; Papke and Thinschmidt, 1998). The pharmacological data with MLA and modulators was analysed using both peak values and AUC and no significant difference was seen, we therefore chose to present the data as AUC rather than peak analysis. It should be noted that in the case of type II PAMs care should be taken since the charge that passes through the channel at high concentrations is more dependent on the onset of the washing out of agonist and compound. Testing with increasing concentrations of nicotine elicited the maximum AUC response with 111 M nicotine, which gave a response that was found to be 40% of the maximum AUC seen with acetylcholine (300 M) (Fig. 3). Nicotine therefore is an agonist with a higher affinity but a lower efficacy than ACh and we therefore consider this a partial agonist. These results are in very good agreement with data obtained from the Dynaflow system, AUC EC 50 = 9.9 M, Imax = 0.4 compared to ACh (Dunlop et al., 2007) and oocytes, AUC EC 50 = 13.2 M, Imax = 0.6 compared to acetylcholine (Papke et al., 2007). Application of increasing concentrations of MLA antagonized the ACh activated current, as expected (Fig. 4A). The IC 50 as determined by analysis of AUC was 0.25 ± 0.09 nm (n =6)(Fig. 4B). This is in excellent agreement with published values determined in a similar GH4-C1 cell line, oocytes and hippocampal neurons where IC 50 values were reported between 0.02 and 0.6 nm (Alkondon et al., 1992; Briggs and McKenna, 1996; Palma et al., 1996; Virginio et al., 2002). One of the positive allosteric modulators tested was PNU , a compound that exhibits its effect on the desensitizing properties of the channel. The EC 50 was estimated to be 4.5 M which is comparable to the M found by Hurst et al. and Gronlien et al. using a fluorescence-based assay and two electrode voltage clamp (Gronlien et al., 2007; Hurst et al., 2005). The other modulator, NS1738, was found to have an EC 50 of 12 M, which is somewhat right-shifted but still within range of the published value (1.6 M) (Timmermann et al., 2007). The 7 nachr channels were stably expressed in GH4-C1 cells. This cell line worked as well as CHO or HEK293 cells, the more commonly used cell lines for stable ion channel transfections and automated patch-clamp. In QPatch experiments more than 80% of the cells achieved gigaseal formation and 70% of the total remained as tight whole-cell recordings. Approximately 50% of the total cells tested provided usable recordings and this drop was due, in part, to a lack of expression. Initially it was thought that this was due to the small size of the cells which results in a smaller current density per cell, in addition to faulty whole cells that can be detected since these cells are close to the limit of what can be detected reliably. The same problems were encountered when the channel was co-expressed with the putative chaperone ric-3 in the larger HEK293. Getting the conditions right for sufficient expression of functional channels and ric-3 in combination seems therefore to be an important challenge (Williams et al., 2005). Different strategies have been employed by us and others to address these issues e.g. with nicotine or depolarizing agents (Molinari et al., 1998; Quik et al., 1996). Future QPatch studies with 7 nachr channels would benefit from all cells expressing measurable current. However, obtaining five recordings (on average) per 16-channel QPlate and testing 5 10 QPlates per day provides an increase in throughput of 5 10-fold compared to manual patch-clamp experiments. A reduction in current amplitude was initially observed with repeated application of agonist on QPatch. A similar effect has been observed with the PatchXpress platform (MDS Analytical
6 6 S. Friis et al. / Journal of Neuroscience Methods xxx (2008) xxx xxx Technologies), but not with manual patch-clamp with Dynaflow (unpublished observations). Since this effect is removed with the inclusion of acetylcholine esterase in the extracellular solution, this appears to be related to residual ligand remaining on/near the cells. All known 7 agonists have been found to inhibit the channel with a potency that is between 60 and 130 times higher than for activation (Briggs and McKenna, 1998). Exposure to inhibiting amounts of agonist for 100 s has been shown to irreversibly inhibit the response of the channel even after 1 h of constant washing (Olale et al., 1997). It is therefore of high importance that the ligand is washed away completely due to the incubation periods used in this assay (150 s). The amount of acetylcholine esterase added during a wash is 0.08 U/well, which will break down the added acetylcholine (0.005 mole/well when adding 1 mm) into acetate and choline, which is removed by the washing procedure. Choline has been shown to induce little or no desensitization on the receptor and with a 10-fold lower potency any desensitizing effect is minimized (Papke and Porter Papke, 2002). The properties of flow chambers in automated patch-clamp systems may somehow facilitate desensitization, but the detailed mechanism of this is unknown. A constant and more conventional mechanical washing procedure is normally employed by manual patch-clamp, which may more efficiently wash ligand from the chamber or cells. Enzymatic cleaning of the flow channel was developed in a separate P 2 X-assay where it was found that cells in the channel upstream of the patched cell leaked ATP, thus desensitizing the channel. This was avoided by breaking down the ATP using it as a phosphate donor in the enzymatically driven phosphorylation of glucose by having hexokinase and glucose present during experiments. We have not found the need for the enzymatic approach with other ligand-gated assays where it seems that the agonist washes out more easily. The need for enzymatic removal of agonist poses a possible limitation to the use of the assay to efficiently characterize agonists against this channel, since cumulative dose response experiments might be problematic to perform. We have shown that it is possible with nicotine to obtain data comparable with other studies, but other non-commercial full agonists were not amenable to cumulative dose response experiments (data not shown). Of course, treatment of cells with a non-cumulative dose response protocol is a feasible approach in this parallel recording instrument. The QPatch readily dissected the mechanism of action of both type I and II PAMs. This demonstrates the high content information the experimenter gains from using high fidelity parallel patch-clamp, compared to fluorescence-based assays in which the throughput is much higher, but the information content significantly lower. We report that the QPatch can reliably measure fast activating and desensitizing ligand-gated ion channels exemplified by the 7 nachr. The QPatch resolves currents and produces data with a quality that generally agrees with published and internally generated manual patch-clamp data. The platform is suitable for agonist, antagonist and PAM characterization. Assays described in this paper demonstrate that the QPatch system allows higher throughput screening of compounds targeted against 7 nachr channels. Acknowledgements We would like to acknowledge Brian Jow and Flora Jow for providing the Dynaflow data and Jeffrey Webber, Dorthe Nielsen, Nadia Larsen and Angela Kramer for technical assistance. References Adler LE, Olincy A, Waldo M, Harris JG, Griffith J, Stevens K, et al. Schizophrenia, sensory gating, and nicotinic receptors. Schizophr Bull 1998;24: Alkondon M, Pereira EF, Wonnacott S, Albuquerque EX. Blockade of nicotinic currents in hippocampal neurons defines methyllycaconitine as a potent and specific receptor antagonist. Mol Pharmacol 1992;41: Arendash GW, Sengstock GJ, Sanberg PR, Kem WR. Improved learning and memory in aged rats with chronic administration of the nicotinic receptor agonist GTS-21. Brain Res 1995;674: Briggs CA, McKenna DG. Activation and inhibition of the human alpha7 nicotinic acetylcholine receptor by agonists. Neuropharmacology 1998;37: Briggs CA, McKenna DG. Effect of MK-801 at the human alpha7 nicotinic acetylcholine receptor. Neuropharmacology 1996;35: Delbono O, Gopalakrishnan M, Renganathan M, Monteggia LM, Messi ML, Sullivan JP. Activation of the recombinant human alpha7 nicotinic acetylcholine receptor significantly raises intracellular free calcium. J Pharmacol Exp Ther 1997;280: Dunlop J, Roncarati R, Jow B, Bothmann H, Lock T, Kowwal D, et al. In vitro screening strategies for nicotinic receptor ligands. Biochem Pharmacol 2007;74: Feuerbach D, Lingenhohl K, Dobbins P, Mosbacher J, Corbett N, Nozulak J, et al. Coupling of human nicotinic acetylcholine receptors alpha7 to calcium channels in GH3 cells. Neuropharmacology 2005;48: Freedman R, Adler LE, Bickford P, Byerley W, Coon H, Cullum CM, et al. Schizophrenia and nicotinic receptors. Harv Rev Psychiatry 1994;2: Freedman R, Hall M, Adler LE, Leonard S. Evidence in postmortem brain tissue for decreased numbers of hippocampal nicotinic receptors in schizophrenia. Biol Psychiatry 1995;38: Freedman R, Leonard S. Genetic linkage to schizophrenia at chromosome 15q14. Am J Med Genet 2001;105: Freedman R, Leonard S, Gault JM, Hopkins J, Cloninger CR, Kaufmann CA, et al. Linkage disequilibrium for schizophrenia at the chromosome 15q13 14 locus of the alpha7-nicotinic acetylcholine receptor subunit gene (CHRNA7). Am J Med Genet 2001;105:20 2. Gotti C, Clementi F. Neuronal nicotinic receptors: from structure to pathology. Prog Neurobiol 2004;74: Gotti C, Riganti L, Vailati S, Clementi F. Brain neuronal nicotinic receptors as new targets for drug discovery. Curr Pharm Des 2006a;12: Gotti C, Zoli M, Clementi F. Brain nicotinic acetylcholine receptors: native subtypes and their relevance. Trends Pharmacol Sci 2006b;27: Gronlien JH, Hakerud M, Ween H, Thorin-Hagene K, Briggs CA, Gopalakrishnan M, et al. Distinct profiles of alpha7 nachr positive allosteric modulators revealed by structurally diverse chemotypes. Mol Pharmacol 2007;72: Hajos M, Hurst RS, Hoffmann WE, Krause M, Wall TM, Higdon NR, et al. The selective alpha7 nicotinic acetylcholine receptor agonist PNU [N-[(3R)-1-Azabicyclo[2.2.2]oct-3-yl]-4-chlorobenzamide hydrochloride] enhances GABAergic synaptic activity in brain slices and restores auditory gating deficits in anesthetized rats. J Pharmacol Exp Ther 2005;312: Hurst RS, Hajos M, Raggenbass M, Wall TM, Higdon NR, Lawson JA, et al. A novel positive allosteric modulator of the alpha7 neuronal nicotinic acetylcholine receptor: in vitro and in vivo characterization. J Neurosci 2005;25: Jensen A, Frolund B, Liljefors T, Krogsgaard-Larsen P. Neuronal nicotinic acetylcholine receptors: structural revelations, target identifications and therapeutic inspirations. J Med Chem 2005;48: Kitagawa H, Takenouchi T, Azuma R, Wesnes KA, Kramer WG, Clody DE, et al. Safety, pharmacokinetics, and effects on cognitive function of multiple doses of GTS-21 in healthy, male volunteers. Neuropsychopharmacology 2003;28: Levin ED, Bettegowda C, Blosser J, Gordon J. AR-R17779, and alpha7 nicotinic agonist, improves learning and memory in rats. Behav Pharmacol 1999;10: Lindstrom J. Neuronal nicotinic acetylcholine receptors. Ion Channels 1996;4: Mathes C. QPatch: the past, present and future of automated patch clamp. Expert Opin Ther Targets 2006;10: Molinari EJ, Delbono O, Messi ML, Renganathan M, Arneric SP, Sullivan JP, et al. Up-regulation of human alpha7 nicotinic receptors by chronic treatment with activator and antagonist ligands. Eur J Pharmacol 1998;347: Olale F, Gerzanich V, Kuryatov A, Wang F, Lindstrom J. Chronic nicotine exposure differentially affects the function of human alpha3, alpha4, and alpha7 neuronal nicotinic receptor subtypes. J Pharmacol Exp Ther 1997;283: Olincy A, Harris JG, Johnson LL, Pender V, Kongs S, Allensworth D, et al. Proof-ofconcept trial of an alpha7 nicotinic agonist in schizophrenia. Arch Gen Psychiatry 2006;63: Palma E, Bertrand S, Binzoni T, Bertrand D. Neuronal nicotinic alpha7 receptor expressed in Xenopus oocytes presents five putative binding sites for methyllycaconitine. J Physiol 1996;491(Pt 1): Papke RL. Estimation of both the potency and efficacy of alpha7 nachr agonists from single-concentration responses. Life Sci 2006;78: Papke RL, Dwoskin LP, Crooks PA. The pharmacological activity of nicotine and nornicotine on nachrs subtypes: relevance to nicotine dependence and drug discovery. J Neurochem 2007;101: Papke RL, Meyer E, Nutter T, Uteshev VV. Alpha7 receptor-selective agonists and modes of alpha7 receptor activation. Eur J Pharmacol 2000;393: Papke RL, Porter Papke JK. Comparative pharmacology of rat and human alpha7 nachr conducted with net charge analysis. Br J Pharmacol 2002;137:49 61.
7 S. Friis et al. / Journal of Neuroscience Methods xxx (2008) xxx xxx 7 Papke RL, Thinschmidt JS. The correction of alpha7 nicotinic acetylcholine receptor concentration response relationships in Xenopus oocytes. Neurosci Lett 1998;256: Pichat P, Bergis OE, Terranova JP, Urani A, Duarte C, Santucci V, et al. SSR180711, a novel selective alpha7 nicotinic receptor partial agonist. II. Efficacy in experimental models predictive of activity against cognitive symptoms of schizophrenia. Neuropsychopharmacology 2007;32: Quik M, Choremis J, Komourian J, Lukas RJ, Puchacz E. Similarity between rat brain nicotinic alpha-bungarotoxin receptors and stably expressed alphabungarotoxin binding sites. J Neurochem 1996;67: Simosky JK, Stevens KE, Kem WR, Freedman R. Intragastric DMXB-A, an alpha7 nicotinic agonist, improves deficient sensory inhibition in DBA/2 mice. Biol Psychiatry 2001;50: Stevens KE, Kem WR, Mahnir VM, Freedman R. Selective alpha7-nicotinic agonists normalize inhibition of auditory response in DBA mice. Psychopharmacology (Berl) 1998;136: Timmermann GB, Gronlien JH, Kohlhaas KL, Nielsen EO, Dam E, Jorgensen TD, et al. An allosteric modulator of the alpha7 nicotinic acetylcholine receptor possesses cognition-enhancing properties in vivo. J Pharmacol Exp Ther 2007;323: Virginio C, Giacometti A, Aldegheri L, Rimland JM, Terstappen GC. Pharmacological properties of rat alpha7 nicotinic receptors expressed in native and recombinant cell systems. Eur J Pharmacol 2002;445: Williams ME, Burton B, Urrutia A, Shcherbatko A, Chavez-Noriega LE, Cohen CJ, et al. Ric-3 promotes functional expression of the nicotinic acetylcholine receptor alpha7 subunit in mammalian cells. J Biol Chem 2005;280: Willumsen NJ, Bech M, Olesen SP, Jensen BS, Korsgaard MP, Christophersen P. High throughput electrophysiology: new perspectives for ion channel drug discovery. Recept Channels 2003;9:3 12.
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