Differential Central NOS-NO Signaling Underlies Clonidine Exacerbation of Ethanol-Evoked Behavioral Impairment
Article first published online: 17 DEC 2009
Copyright © 2009 by the Research Society on Alcoholism
Alcoholism: Clinical and Experimental Research
Volume 34, Issue 3, pages 555–566, March 2010
How to Cite
Bender, T. S. and Abdel-Rahman, A. A. (2010), Differential Central NOS-NO Signaling Underlies Clonidine Exacerbation of Ethanol-Evoked Behavioral Impairment. Alcoholism: Clinical and Experimental Research, 34: 555–566. doi: 10.1111/j.1530-0277.2009.01121.x
- Issue published online: 2 MAR 2010
- Article first published online: 17 DEC 2009
- Received for publication May 29, 2009; accepted October 24, 2009.
- Neuronal NOS;
- Endothelial NOS;
- Locus Coeruleus
Background: The molecular mechanisms that underlie clonidine exacerbation of behavioral impairment caused by ethanol are not fully known. We tested the hypothesis that nitric oxide synthase (NOS)-derived nitric oxide (NO) signaling in the locus coeruleus (LC) is implicated in this phenomenon.
Methods: Male Sprague–Dawley rats with intracisternal (i.c.) and jugular vein cannulae implanted 6 days earlier were tested for drug-induced behavioral impairment. The latter was assessed as the duration of loss of righting reflex (LORR) and rotorod performance every 15 minutes until the rat recovered to the baseline walk criterion (180 seconds). In a separate cohort, we measured p-neuronal NOS (nNOS), p-endothelial NOS (eNOS), and p-ERK1/2 in the LC following drug treatment, vehicle, or NOS inhibitor.
Results: Rats that received clonidine [60 Ig/kg, i.v. (intravenous)] followed by ethanol (1 or 1.5 g/kg, i.v.) exhibited synergistic impairment of rotorod performance. Intracisternal pretreatment with nonselective NOS inhibitor Nω-nitro-l-arginine methyl ester (l-NAME, 0.5 mg) or selective nNOS inhibitor N-propyl-l-arginine (1 μg) exacerbated the impairment of rotorod performance caused by clonidine–ethanol combination. Exacerbation of behavioral impairment was caused by l-NAME enhancement of the effect of ethanol, not clonidine. l-NAME did not influence blood ethanol levels; thus, the interaction was pharmacodynamic. LORR caused by clonidine (60 μg/kg, i.v.)–ethanol (1 g/kg, i.v.) combination was abolished by selective inhibition of central eNOS (l-NIO, 10 μg i.c.) but not by nNOS inhibition under the same conditions. Western blot analyses complemented the pharmacological evidence by demonstrating that clonidine–ethanol combination inhibits phosphorylation (activation) of nNOS (p-nNOS) and increases the level of phosphorylated eNOS (p-eNOS) in the LC; the change in p-nNOS was paralleled by similar change in LC p-ERK1/2. NOS inhibitors alone did not affect the level of nitrate/nitrite, p-nNOS, p-eNOS, or p-ERK1/2 in the LC.
Conclusions: Alterations in NOS-derived NO in the LC underlie clonidine–ethanol induced behavioral impairment. A decrease in nNOS activity, due at least partly to a reduction in nNOS phosphorylation, mediates rotorod impairment, while enhanced eNOS activity contributes to LORR, elicited by clonidine–ethanol combination.