Characterization of Nicotinic Agonist-Induced [ 3 H]Dopamine Release from Synaptosomes Prepared from Four Mouse Brain Regions SHARON R. GRADY, KAREN L. MURPHY, JIAN CAO, MICHAEL J. MARKS, J. MICHAEL MCINTOSH, and ALLAN C. COLLINS Institute for Behavioral Genetics, University of Colorado, Boulder, Colorado (S.R.G., K.L.M., J.C., M.J.M., A.C.C.); and Departments of Biology and Psychiatry, University of Utah, Salt Lake City, Utah (M.M.) Received September 24, 2001; accepted January 18, 2002 This article is available online at http://jpet.aspetjournals.org ABSTRACT The inhibition of uptake of [ 3 H]dopamine into synaptosomes prepared from four mouse brain regions was investigated. The inhibition curves demonstrated that in three regions, striatum, nucleus accumbens, and olfactory tubercle, [ 3 H]dopamine was taken up exclusively by dopaminergic terminals. In frontal cor- tex, however, only a portion of the uptake was into dopaminer- gic terminals, with a larger amount taken up by noradrenergic terminals, and another small portion by serotonergic terminals. Release studies in frontal cortex indicated that in this region only dopaminergic and noradrenergic terminals are capable of packaging [ 3 H]dopamine in a form allowing vesicle-mediated release; additionally, only the dopaminergic terminals have functional presynaptic nAChRs that, when stimulated by nico- tinic agonists, evoke [ 3 H]dopamine release. Agonist-stimulated [ 3 H]dopamine release was characterized from synaptosomes prepared from four mouse brain regions. -Conotoxin MII was a partial inhibitor of dopamine release in all of the brain regions, which suggests that a minimum of two nicotinic cholinergic receptors (nAChRs) are expressed in the nerve terminals of all four brain regions. No nicotine-induced [ 3 H]dopamine release was detected in any brain region when the synaptosomes were prepared from 2 null mutant mice, which indicates that the 2 subunit is required for all nAChRs mediating this release. Dose- response curves were constructed for seven agonists in each of the brain regions. The pharmacological properties of synap- tosomal [ 3 H]dopamine release appear similar across the four brain regions. The results suggest that all four regions express the same nAChRs, although subtle regional differences may exist. Major dopaminergic pathways originate in the substantia nigra (SN) and the ventral tegmental area (VTA) of the midbrain and project to several forebrain regions via the medial forebrain bundle. The nigrostriatal and mesocortical projections have a distinct topography, both mediolateral and rostrocaudal and, in the reverse orientation, dorsoventral (Moore and Bloom, 1978; Fuxe at al., 1985; Cragg et al., 1997). The density and intraneuronal distribution of varicos- ities also vary across brain region. Extensive axonal collat- eralization and dense varicosities are seen in the striatum, nucleus accumbens, and olfactory tubercle, whereas in the frontal cortex infrequent varicosities are seen on single axons (Moore and Bloom, 1978). There are also measurable differ- ences in levels of mRNA and proteins involved in dopamine metabolism (Ciliax et al., 1995; Cragg et al., 1997), as well as in rates of dopamine uptake and release in the various ter- minal fields (Garris and Wightman, 1994). It has been suggested that there are two functionally re- lated components of dopamine release: phasic and tonic (Grace, 1991). Phasic release is mediated by the somatoden- dritic activation and firing of dopaminergic neurons, result- ing in dopamine release in the terminal fields. Tonic release is a modulatory release mediated by presynaptic stimulation of dopaminergic terminals that may be important in regulat- ing the responsiveness of the dopaminergic system. In sup- port of this idea, the dopamine transporter null mutant mice, which exhibit chronic elevation of extracellular dopamine at the terminal field, also show decreased responsiveness of the synthesis- and release-regulating autoreceptors (Jones et al., 1999). Nicotinic cholinergic receptors (nAChRs) are found on do- paminergic neurons at somatodendritic sites as well as pre- This work was supported by Grants DA-03194, DA-12242, and MH-53631, and a Research Scientist Award, DA-00197 (to A.C.C.). Portions of this work have been presented previously in Grady SR, Clark AL, and Collins AC (1996) Comparison of L-nicotine-stimulated [ 3 H]dopamine release from four mouse brain regions. Soc Neurosci 22:1270. ABBREVIATIONS: SN, substantia nigra; VTA, ventral tegmental area; nAChR, nicotinic cholinergic receptor; DHE, dihydro--erythroidine; ACh, acetylcholine; CtxMII, -conotoxin MII; NA, nucleus accumbens; FC, frontal cortex; OT, olfactory tubercle; ST, striatum; GBR12909, 1-(2-[bis(4- fluorophenyl)methoxy]ethyl]-4-[3-phenylpropyl]piperazine dihydrochloride. 0022-3565/02/3012-651–660$7.00 THE JOURNAL OF PHARMACOLOGY AND EXPERIMENTAL THERAPEUTICS Vol. 301, No. 2 Copyright © 2002 by The American Society for Pharmacology and Experimental Therapeutics 4573/981653 JPET 301:651–660, 2002 Printed in U.S.A. 651 at ASPET Journals on January 17, 2017 jpet.aspetjournals.org Downloaded from