In the present study, we demonstrate that choline, a precursor of acetylcholine (ACh) and a product of acetylcholine hydrolysis by acetylcholinesterase (AChE), acts as an efficient and relatively selective agonist of alpha7-containing nicotinic acetylcholine receptors (nAChR) in neurons cultured from the rat hippocampus, olfactory bulb and thalamus as well as in PC12 cells. Choline was able to activate postsynaptic and presynaptic alpha7 nAChRs, with the latter action resulting in the release of other neurotransmitters. Although choline was approximately one order of magnitude less potent than ACh (EC50 of 1.6 mM for choline and 0.13 mM for ACh), it acted as a full agonist at alpha7 nAChRs. In contrast, choline did not activate alpha4beta2 agonist-bearing nAChRs on hippocampal neurons, and acted as a partial agonist at alpha3beta4-containing nAChRs on PC12 cells. The ethyl alcohol moiety of choline is required for the selective action on alpha7 nAChR. Exposure of cultured hippocampal neurons for 10 min to choline (10-100 microM) resulted in desensitization of the native alpha7 nAChRs. Moreover, chronic exposure (10 days) of the cultured hippocampal neurons to a desensitizing concentration of choline (approximately 30 microM) decreased their responsiveness to ACh. The selective action of choline on native alpha7 nAChRs suggests that this naturally occurring compound may act in vivo as an endogenous ligand for these receptors. Putative physiological actions of choline include retrograde messenger activity during the development of the mammalian central nervous system and during periods of elevated synaptic activity that leads to long-term potentiation.

Galantamine (Reminyl), an approved treatment for Alzheimer's disease (AD), is a potent allosteric potentiating ligand (APL) of human ␣3␤4, ␣4␤2, and ␣6␤4 nicotinic receptors (nAChRs), and of the chicken/mouse chimeric ␣7/5-hydroxytryptamine 3 receptor, as was shown by whole-cell patch-clamp studies of human embryonic kidney-293 cells stably expressing a single nAChR subtype. Galantamine potentiates agonist responses of the four nAChR subtypes studied in the same window of concentrations (i.e., 0.1-1 M), which correlates with the cerebrospinal fluid concentration of the drug at the recommended daily dosage of 16 to 24 mg. At concentrations Ͼ10 M, galantamine acts as an nAChR inhibitor. The other presently approved AD drugs, donepezil and rivastigmine, are devoid of the nicotinic APL action; at micromolar concentrations they also block nAChR activity. Using five CHO-SRE-Luci cell lines, each of them expressing a different human muscarinic receptor, and a reporter gene assay, we show that galantamine does not alter the activity of M1-M5 receptors, thereby confirming that galantamine modulates selectively the activity of nAChRs. These studies support our previous proposal that the therapeutic action of galantamine is mainly produced by its sensitizing action on nAChRs rather than by general cholinergic enhancement due to cholinesterase inhibition. Galantamine's APL action directly addresses the nicotinic deficit in AD.

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Nicotinic acetylcholine receptors play important roles in numerous cognitive processes as well as in several debilitating central nervous system (CNS) disorders. In order to fully elucidate the diverse roles of nicotinic acetylcholine receptors in CNS function and dysfunction, a detailed knowledge of their cellular and subcellular localizations is essential. To date, methods to precisely localize nicotinic acetylcholine receptors in the CNS have predominantly relied on the use of antireceptor subunit antibodies. Although data obtained by immunohistology and immunoblotting are generally in accordance with ligand binding studies, some discrepancies remain, in particular with electrophysiological findings. In this context, nicotinic acetylcholine receptor subunit-deficient mice should be ideal tools for testing the specificity of subunitdirected antibodies. Here, we used standard protocols for immunohistochemistry and western blotting to examine the antibodies raised against the a3-, a4-, a7-, b2-, and b4-nicotinic acetylcholine receptor subunits on brain tissues of the respective knock-out mice. Unexpectedly, for each of the antibodies tested, immunoreactivity was the same in wild-type and knock-out mice. These data imply that, under commonly

ABSTRACT:Evidence gathered from epidemiologic and behavioral studies have indicated that neuronal nicotinic receptors (nAChRs) are intimately involved in the pathogenesis of a number of neurologic disorders, including Alzheimer's disease, Parkinson's disease, and schizophrenia. In the mammalian brain, neuronal nAChRs, in addition to mediating fast synaptic transmission, modulate fast synaptic transmission mediated by the major excitatory and inhibitory neurotransmitters glutamate and GABA, respectively. Of major interest, however, is the fact that the activity of the different subtypes of neuronal nAChR is also subject to modulation by substances of endogenous origin such as choline, the tryptophan metabolite kynurenic acid, neurosteroids, and ␤-amyloid peptides and by exogenous substances, including the so-called nicotinic allosteric potentiating ligands, of which galantamine is the prototype, and psychotomimetic drugs such as phencyclidine and ketamine. The present article reviews and discusses the effects of unconventional ligands on nAChR activity and briefly describes the potential benefits of using some of these compounds in the treatment of neuropathologic conditions in which nAChR function/expression is known to be altered.