doi: 10.1016/j.neuropharm.2010.07.006. Epub 2010 Jul 13.
In vitro and in vivo characterization of a novel negative allosteric modulator of neuronal nAChRs
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- PMID:20633568
- PMCID: PMC2998338
- DOI: 10.1016/j.neuropharm.2010.07.006
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In vitro and in vivo characterization of a novel negative allosteric modulator of neuronal nAChRs
Galya R Abdrakhmanova et al. Neuropharmacology.2010 Nov.
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Abstract
In this study, we compared the in vitro and in vivo neuronal nicotinic acetylcholine receptor (nAChR) properties of 1,2,3,3a,4,8b-hexahydro-2-benzyl-6-N,N-dimethylamino-1-methylindeno[1,2,-b]pyrrole (HDMP, 4) to that of negative allosteric modulator (NAM), PCP. Patch-clamp experiments showed that HDMP exhibited an inhibitory functional activity at α7 nAChRs with an IC(50) of 0.07 μM, and was 357- and 414-fold less potent at α4β2 and α3β4 nAChRs, with IC(50)s of 25.1 and 29.0 μM, respectively. Control patch-clamp experiments showed that PCP inhibited α7, α4β2 and α3β4 nAChRs with IC(50)s of to 1.3, 29.0 and 6.4 μM, respectively. Further, HDMP did not exhibit any appreciable binding affinity to either α7 or α4β2 nAChRs, suggesting its action via a non-competitive mechanism at these neuronal nAChR subtypes. The in vivo study showed that HDMP was a potent antagonist of nicotine-induced analgesia in the tail-flick (AD(50)=0.008 mg/kg), but not in the hot-plate test. All together, our in vitro and in vivo data suggest that HDMP is a novel NAM of neuronal nAChRs with potent inhibitory activity at α7 nAChR subtype at concentrations ≤ 1μM that are not effective for α4β2 and α3β4 nAChRs.
Copyright © 2010 Elsevier Ltd. All rights reserved.
Figures
Structures of nicotine (1), epibatidine (2), dihydro-β-erythroidine (3), PCP, HDMP (4), and5.
Analysis of the antagonist activity of HDMP in three neuronal nAChR subtypes. A, Concentration-response relationships for HDMP constructed in α3β4 (■), α4β2 (●) and α7 (□) nAChR subtypes. The peak amplitude of ACh (EC50)-evoked currents was taken in each cell to normalize the peak amplitude of the currents that were evoked in the presence of HDMP at different concentrations. The curves were fitted to Hill equation. Symbols and bars represent the mean ± S.E.M. B, Examples of the effect of HDMP at 0.1 μM concentration illustrate a pronounced (55%) inhibition of whole-cell currents in α7, but no effect in α4β2 or α3β4 nAChR expressing cells. Holding potential in A and B was −80 mV.
Comparison of PCP-induced inhibitory effect on three different neuronal nAChR subtypes. A, Concentration-response relationships for PCP constructed in α3β4 (■), α7 (◆) and α4β2 (◆) nAChRs. The peak amplitude of ACh (EC50)-evoked currents was taken in each cell to normalize the peak amplitude of the currents that were evoked in the presence of PCP at different concentrations. The curves were fitted to Hill equation. Symbols and bars represent the mean ± S.E.M. B, Examples of inhibitory effect of PCP at concentrations close to determined IC50 values in three representative cells expressing α4β2 (top), α3β4 (middle) and α7 (bottom) nAChRs. Holding potential in A and B was −80 mV.
Voltage-dependence of inhibitory effect of HDMP on neuronal nAChRs.Each panel shows data obtained from a single representative cell expressing α7 (A, triangles), α4β2 (B, circles) or α3β4 (C, rhombs) nAChRs. ACh(EC50)-induced currents were evoked at various holding potentials in the range from −100 to +60 mV, quantified at its maximal amplitude in the absence (closed symbols) and presence (open symbols) of HDMP at the concentration close to its IC50 value determined for each receptor subtype, and plotted versus the corresponding holding potential. The inserts in each panel represent superimposed traces of the ACh-induced currents in the absence and presence of HDMP.
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