Abstract
The effects of the endocannabinoid anandamide (arachidonylethanolamide, AEA) on the function of α4β2 nicotinic acetylcholine receptors (nAChR) stably expressed in SH-EP1 cells were investigated using the whole-cell patch-clamp technique. In the concentration range of 200 nM to 2 μM, AEA significantly reduced the maximal amplitudes and increased the desensitization of acetylcholine (ACh)-induced currents. The effects of AEA could be neither replicated by the exogenous cannabinoid Δ9-tetrahydrocannabinol (1 μM) nor reversed by the selective CB1 receptor antagonist 5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methyl-N-(piperidin-1-yl)-1H-pyrazole-3-carboxamide (SR-141716A) (1 μM). The actions of AEA were apparent when applied extracellularly but not during intracellular dialysis. Furthermore, the effects of AEA ACh currents were not altered by the calcium chelator 1,2-bis(2-aminophenoxy)ethane-N,N,N′,N′-tetraacetic acid. The onset and washout of the AEA effects required several minutes (10–30 min), but the latter was significantly decreased in the presence of lipid-free bovine serum albumin (BSA). Moreover, BSA alone increased peak ACh current amplitudes and diminished desensitization rates in naive cells, suggesting a tonic modulation of α4β2 nAChR function by an endogenous AEA-like lipid. Further analysis of AEA effects on α4β2 nAChR-mediated currents, using a two-stage desensitization model, indicated that the first forward rate constant leading to desensitization, k1, increased nearly 30-fold as a linear function of the AEA concentration. In contrast, the observation that the other three rate constants were unaltered by AEA suggested that AEA raised the energy of the activated state. These results indicate that AEA directly inhibits the function of α4β2 nAChRs in a CB1 receptor-independent manner.
Footnotes
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This study was supported by funds from the Intramural Research Program of the National Institute on Drug Abuse, U.S. Department of Health and Human Services.
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Article, publication date, and citation information can be found at http://molpharm.aspetjournals.org.
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doi:10.1124/mol.107.036939.
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ABBREVIATIONS: Δ9-THC, Δ9-tetrahydrocannabinol; ACh, acetylcholine; AEA, arachidonylethanolamide, anandamide, N-(2-hydroxyethyl)-5,8,11,14-eicosatetraenamide (all-Z)-; BAPTA, 1,2-bis(2-aminophenoxy)ethane-N,N,N′,N′-tetraacetic acid; BSA, bovine serum albumin; CNS, central nervous system; CB1, cannabinoid type 1 receptor; nAChR, nicotinic acetylcholine receptor; SR-141716A, 5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methyl-N-(piperidin-1-yl)-1H-pyrazole-3-carboxamide; URB 597, cyclohexyl carbamic acid 3′-carbamoylbiphenyl-3-yl ester.
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↵ The online version of this article (available at http://molpharm.aspetjournals.org) contains supplemental material.
- Received April 11, 2007.
- Accepted July 12, 2007.
- The American Society for Pharmacology and Experimental Therapeutics
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