Dual Action of n-Butanol on Neuronal Nicotinic 4 2 Acetylcholine Receptors

n-Alcohols exert a dual action on neuronal nicotinic acetylcholine (ACh) receptors with short-chain alcohols exhibiting potentiating action and long-chain alcohols exhibiting inhibitory action. n-Butanol lies at the transition point from potentiation to inhibition. To elucidate the mechanism of dual action of alcohols, the effects of n-butanol on the human 4 2 ACh receptors expressed in the HEK293 cell line were analyzed in detail by the whole-cell patch-clamp technique. Prolonged applications of n-butanol evoked small currents with an EC50 value of 230 90 mM and a Hill coefficient of 1.8 0.4. This current was blocked by either the ACh channel blocker mecamylamine or the receptor blocker dihydro-erythroidine, indicating that butanol activated receptors as a partial agonist. As expected from its partial agonist action, n-butanol also modulated AChinduced currents in a concentration-dependent manner. Butanol at 300 mM potentiated currents induced by low concentrations of ACh ( 30 M), while inhibiting the currents induced by high concentrations of ACh (100–3000 M). In addition, butanol at a low concentration (10 mM) suppressed the currents evoked by 10 to 3000 M ACh, a result consistent with a channel-blocking action. Most features of n-butanol effects were satisfactorily simulated by a model in which butanol acts as a partial agonist and as a channel blocker. The importance of neuronal nicotinic acetylcholine receptors (nnAChRs) in brain function and drug action is increasingly recognized. Because nnAChRs are located on soma, preterminal, and presynaptic regions of GABAergic and other interneurons in the cortex and hippocampus, their modulation caused by various drugs could lead to a cascade of synaptic events involving multiple neurotransmitters, resulting in complex behavioral changes. n-Alcohols have been shown to exert a dual action on nAChRs, depending on the carbon chain length. In muscle nAChRs, ethanol and other short-chain alcohols prolong the decay phase of the miniature end-plated currents (EPCs) (Gage et al., 1975), increase the peak EPC amplitude, and prolong the channel lifetime (Bradley et al., 1980; Linder et al., 1984). In contrast, longer chain alcohols (n-butanol, nhexanol, and n-octanol) accelerate the EPC decay and reduce the peak EPC amplitude (Bradley et al., 1984). At the singlechannel level, butanol and pentanol increase the burst frequency, resulting from an increase in the opening rate of the ACh receptor channel (Dilger et al., 1994; Liu et al., 1994). This effect would account for the observation that ethanol increased the saturating response induced by high concentrations of ACh (Aistrup et al., 1999; Zuo et al., 2001). In addition, n-alcohols reduce single-channel conductance and shorten the mean channel open time in muscle nAChRs, both of which are thought to be the basis for the inhibitory action of alcohols (Murrell et al., 1991; Forman and Zhou, 1999). Experiments performed with nnAChRs of native neurons have shown that ethanol potentiates ACh-induced currents in 4 2-type nnAChRs in rat cortical neurons in primary culture (Aistrup et al., 1999). Using the 4 2 nnAChRs stably expressed in human embryonic kidney (HEK) cell line, we have previously found that shorter chain alcohols from methanol to n-propanol potentiate the currents induced by ACh, whereas longer chain alcohols from n-pentanol to n-dodecanol (C5–C12) inhibit the currents (Zuo et al., 2001). n-Butanol (C4) is at the transition position from potentiation to inhibition and exerts a biphasic effect, either potentiating or inhibiting the currents depending on the concentrations of ACh and butanol. At low ACh concentrations, butanol exhibits a biphasic inhibition being accentuated by increasing butanol concentration from 1 to 100 mM, but becoming less pronounced or even being converted to potentiation at concentrations higher than 100 mM (Zuo et al., 2001). Because n-butanol is at the transition point from potentiation to inhibition in the action of a series of aliphatic chain n-alcohols, and also because n-butanol itself exhibits a dual This work was supported by National Institutes of Health Grant AA07836. Article, publication date, and citation information can be found at http://jpet.aspetjournals.org. DOI: 10.1124/jpet.102.044537. ABBREVIATIONS: nAChR, nicotinic acetylcholine receptor; EPC, end-plated current; ACh, acetylcholine; nnAChR, neuronal nAChR; HEK, human embryonic kidney; DH E, dihydro-erythroidine. 0022-3565/03/3043-1143–1152$7.00 THE JOURNAL OF PHARMACOLOGY AND EXPERIMENTAL THERAPEUTICS Vol. 304, No. 3 Copyright © 2003 by The American Society for Pharmacology and Experimental Therapeutics 44537/1044503 JPET 304:1143–1152, 2003 Printed in U.S.A.