The identity of nicotinic receptor subtypes sufficient to elicit both the acute and chronic effects of nicotine dependence is unknown. We engineered mutant mice with a4 nicotinic subunits containing a single point mutation, Leu9' --> Ala9' in the pore-forming M2 domain, rendering a4* receptors hypersensitive to nicotine. Selective activation of a4* nicotinic acetylcholine receptors with low doses of agonist recapitulates nicotine effects thought to be important in dependence, including reinforcement in response to acute nicotine administration, as well as tolerance and sensitization elicited by chronic nicotine administration. These data indicate that activation of a4* receptors is sufficient for nicotine-induced reward, tolerance, and sensitization.

Summary α6-containing (α6*) nicotinic ACh receptors (nAChRs) are selectively expressed in dopamine (DA) neurons and participate in cholinergic transmission. We generated and studied mice with gain-of-function α6* nAChRs, which isolate and amplify cholinergic control of DA transmission. In contrast to gene knockouts or pharmacological blockers, which show necessity, we show that activating α6* nAChRs and DA neurons is sufficient to cause locomotor hyperactivity. α6L9’S mice are hyperactive in their home cage and fail to habituate to a novel environment. Selective activation of α6* nAChRs with low doses of nicotine, by stimulating DA but not GABA neurons, exaggerates these phenotypes and produces a hyperdopaminergic state in vivo. Experiments with additional nicotinic drugs show that altering agonist efficacy at α6* provides fine-tuning of DA release and locomotor responses. α6*-specific agonists or antagonists may, by targeting endogenous cholinergic mechanisms, provide a new method for manipulating DA transmission in Parkinson’s disease, nicotine dependence, or attention deficit hyperactivity disorder.

Understanding effects of chronic nicotine requires identifying the neurons and synapses whose responses to nicotine itself, and to endogenous acetylcholine, are altered by continued exposure to the drug. To address this problem, we developed mice whose ␣4 nicotinic receptor subunits are replaced by normally functioning fluorescently tagged subunits, providing quantitative studies of receptor regulation at micrometer resolution. Chronic nicotine increased ␣4 fluorescence in several regions; among these, midbrain and hippocampus were assessed functionally. Although the midbrain dopaminergic system dominates reward pathways, chronic nicotine does not change ␣4* receptor levels in dopaminergic neurons of ventral tegmental area (VTA) or substantia nigra pars compacta. Instead, upregulated, functional ␣4* receptors localize to the GABAergic neurons of the VTA and substantia nigra pars reticulata. In consequence, GABAergic neurons from chronically nicotine-treated mice have a higher basal firing rate and respond more strongly to nicotine; because of the resulting increased inhibition, dopaminergic neurons have lower basal firing and decreased response to nicotine. In hippocampus, chronic exposure to nicotine also increases ␣4* fluorescence on glutamatergic axons of the medial perforant path. In hippocampal slices from chronically treated animals, acute exposure to nicotine during tetanic stimuli enhances induction of long-term potentiation in the medial perforant path, showing that the upregulated ␣4* receptors in this pathway are also functional. The pattern of cell-specific upregulation of functional ␣4* receptors therefore provides a possible explanation for two effects of chronic nicotine: sensitization of synaptic transmission in forebrain and tolerance of dopaminergic neuron firing in midbrain.