Previous studies have revealed the existence of a gene family that encodes a group of neuronal nicotinic acetylcholine receptor (nAChR) subunits. Four members of this family have been characterized thus far; three of these subunits (alpha 2, alpha 3, and alpha 4) are structurally related to the ligand binding subunit expressed in muscle and form functional nAChRs when combined with the beta 2 gene product in Xenopus oocytes. In addition, the alpha 4 gene appears to encode two different products (alpha 4-1 and alpha 4-2) that have been proposed to arise by alternative mRNA splicing. Nine different [35S]-complementary ribonucleic acid (cRNA) probes were used in the present study to map the distribution of these nAChR subunit mRNAs throughout the central nervous system (CNS) of the rat. It was found that the beta 2 gene is expressed in most regions of the CNS, as are the alpha subunit genes as a group. However, each alpha gene is expressed in a unique, although partly overlapping, set of neuronal structures. Alpha 4 is the most widely expressed alpha gene, and the evidence suggests that mRNAs for the alpha 4-1 and alpha 4-2 products are virtually always found in the same regions, in approximately the same ratios (alpha 4-2 greater than alpha 4-1). In addition, there are several examples of cell groups that express beta 2 but none of the alpha subunit mRNAs examined here (particularly in the hypothalamus), as well as all groups that express the converse, thus suggesting that additional neuronal nAChR subunits remain to be characterized. Finally, the extensive expression of multiple alpha subunits in certain regions, particularly for alpha 3 and alpha 4 in the thalamus, suggests that there is microheterogeneity in a small population of cells or that some neurons may express more than one alpha subunit. This problem needs to be examined directly with double labeling methods but raises the possibility that some neuronal nAChRs may be composed of more than one kind of alpha subunit. The wide expression of these receptor genes suggests that nAChRs constitute major excitatory systems in the CNS.

DBA mice were chronically treated with nicotine by continuous intravenous infusion of 4.0 mg/kg/hr for 10 d. Drug-treated mice were tolerant to the acute effects of nicotine on locomotor activity and body temperature. The effect of chronic treatment on the amount of L-3H-nicotine binding and RNA encoding for alpha 4, the most widely expressed nicotinic alpha-subunit, was measured in three brain regions. Chronic treatment increased L-3H-nicotine binding in cortex and midbrain but had no effect in cerebellum. In contrast, chronic treatment had no effect on the levels of mRNA encoding for alpha 4 in any of the three brain regions. Subsequently brains were sectioned and L-3H-nicotine binding was measured using quantitative autoradiographic methods. In addition, the relative amounts of mRNA for the major nicotinic receptor subunits (alpha 4 and beta 2), as well as for three additional minor subunits (alpha 2, alpha 3, and alpha 5), were determined by in situ hybridization histochemistry followed by quantitation of image intensity. Chronic nicotine treatment resulted in increases in the amount of L-3H-nicotine binding in many but not all brain areas measured. In contrast, chronic treatment had little effect on the intensity of the hybridization signal for the nicotinic subunit mRNA. The results suggest that chronic treatment with nicotine under conditions resulting in maximal steady-state increases in L-3H-nicotine binding has little effect on RNA levels encoding any of four nicotinic alpha-subunits and the beta 2-subunit.

Rasmussen's encephalitis is a progressive childhood disease of unknown cause characterized by severe epilepsy, hemiplegia, dementia, and inflammation of the brain. During efforts to raise antibodies to recombinant glutamate receptors (GluRs), behaviors typical of seizures and histopathologic features mimicking Rasmussen's encephalitis were found in two rabbits immunized with GluR3 protein. A correlation was found between the presence of Rasmussen's encephalitis and serum antibodies to GluR3 detected by protein immunoblot analysis and by immunoreactivity to transfected cells expressing GluR3. Repeated plasma exchanges in one seriously ill child transiently reduced serum titers of GluR3 antibodies, decreased seizure frequency, and improved neurologic function. Thus, GluR3 is an autoantigen in Rasmussen's encephalitis, and an autoimmune process may underlie this disease.

conducting cations, and are in desensitized or inactive states while unresponsive to agonist. The likelihood of *Division of Neuroscience Baylor College of Medicine being in a particular state depends on many factors, including the nAChR subtype, the agonist concentra-Houston, Texas 77030-3498 † Molecular Neurobiology Laboratory tion, and the rate of agonist application. A rapid pulse of agonist causes synchronized activation of nAChRs, The Salk Institute San Diego, California 92186-5800 but long-term exposure to an agonist causes desensitization. A slow application of a low agonist concentration Tobacco use in developed countries has been estimated can cause some desensitization without activation because the desensitized receptor has a higher affinity for to cause nearly 20% of all deaths, making it the largest single cause of premature death (Peto et al., 1992). The agonist than the resting or open receptor. In addition, there is evidence that neuronal nAChRs can exist on drive for tobacco by humans is clear. The majority of smokers have tried repeatedly to quit and failed. In about the cell surface as nonfunctional receptors (Margiotta et al., 1987) or can enter long-lived inactivated states 80% of the attempts to quit, smokers return to tobacco in less than 2 years (Schelling, 1992). Although the un- (Lester and Dani, 1994). The higher affinity of the desensitized receptor for derlying mechanisms that cause tobacco abuse are not well understood, the accumulation of evidence indicates agonist and the changing distribution of nAChRs among the various functional states must be considered to un-that nicotine is the primary component of tobacco that motivates continued use despite harmful effects (Schel-derstand what takes place during sustained nicotine use. A knowledge of long-term forms of inactivation may ling, 1992; Stolerman and Shoaib, 1991).Nicotine alone, free of smoke or associated factors, be especially important for understanding the phases of withdrawal symptoms and the development of toler-can elicit drug-seeking behavior in animal studies as demonstrated by self-administration and place prefer-ance to nicotine. Aspects of tolerance and withdrawal could be explained by nicotinic receptors slowly recov-ence experiments (Stolerman and Shoaib, 1991;Corrigall and Coen, 1989). Intravenous self-administration of ering to functional states from various levels of desensitization and inactivation. nicotine is best demonstrated under conditions of limited availability; rats have higher lever-pressing rates Pathways of Reward in Nicotine AbuseA multiplicity of psychopharmacological effects contrib-when nicotine is delivered intermittently rather than continuously (Goldberg and Henningfield, 1988). The re-ute to the reinforcing actions of drugs. A widely accepted hypothesis is that drugs of abuse commandeer sponding rate to nicotine is dose dependent, falling off at both lower and higher concentrations. Responding existing reward pathways that are normally essential for survival. The mesolimbic dopaminergic sys...

Ionotropic glutamate receptors are composed of homomeric or heteromeric configurations of glutamate receptor subunits. We have cloned a member of a novel class of the rat ionotropic glutamate receptor family, termed chi-1. This subunit exhibits an average identity of 27% to NMDA subunits and 23% to non-NMDA subunits. Regional transcript levels of chi-1 are elevated just prior to and during the first postnatal week, with the highest levels present in the spinal cord, brainstem, hypothalamus, thalamus, CA1 field of the hippocampus, and amygdala. The spatial distribution of chi-1 expression is similar from postnatal day 1 (P1) to adulthood. However, transcript levels decline sharply between P7 and P14 and remain attenuated into adulthood. Functional expression studies in Xenopus oocytes injected with in vitro transcribed chi-1 RNA did not demonstrate agonist-activated currents. Pairwise expression of chi-1 with members of the AMPA, KA, or delta class of glutamate recepto subunits either failed to generate agonist-activated currents or failed to alter the underlying current generated by the coexpressed subunit. However, coexpression of chi-1 with subunits forming otherwise functional NMDA receptors resulted in an inhibition of current responses. Since chi-1 did not alter the currents generated by non-NMDA subunits, this suggests that chi-1 may specifically interact with NMDA receptor subunits. Further characterization will be required to establish the precise role of this glutamate receptor subunit in neuronal signaling.

Kainate receptors are abundantly expressed in the hippocampus. Mice with disruption of kainate receptor subunits allow the genetic dissection of the role of each kainate receptor subunits in the synaptic physiology of the hippocampus, as well as in excitotoxic processes. We have compared the action of domoate and kainate on CA1 pyramidal neurons in slices from wild-type and GluR6-/- mice. The difference in the amplitude of inward currents evoked by domoate and kainate between wild-type and GluR6-/- mice demonstrates the presence of functional kainate receptors in CA1 pyramidal neurons. Block of domoate-activated inward currents by the AMPA receptor antagonists 2,3-dihydroxy-6-nitro-7-sulfonyl-benzo(F)quinoxaline (1 microM) and 1-(4-aminophenyl)-3-methylcarbamyl-4-methyl7, 8-methylenedioxy-3,4-dihydro-5H-2,3-benzodiazepine) (GYKI 53655) (50 microM) is complete in GluR6-/- mice but only partial in wild-type mice. In the presence of GYKI 53655, kainate receptor activation dramatically increases the frequency of spontaneous IPSCs in CA1 pyramidal cells from wild-type, as well as GluR6-/-, mice. This results from the kainate receptor-mediated activation of a sustained inward current and an increased action potential firing in afferent GABAergic interneurons of the CA1 field. These effects are observed in wild-type, as well as GluR6-/-, mice. Kainate receptors also decrease the amplitude of evoked IPSCs in CA1 pyramidal cells by increasing synaptic failures in wild-type and GluR6-/- mice. These results indicate that in CA1 pyramidal cells, distinct subtypes of kainate receptors mediate several functionally antagonistic effects.

The regional, cellular, and subcellular distributions of N-methyl-D-aspartate (NMDA) receptor subunit 1, NMDAR-1, were investigated in monkey hippocampus by using a monoclonal antibody directed against a fusion protein corresponding to aa 660-811 of NMDAR-1. The data indicate that many neurons in each subfield of the hippocampus contain NMDAR-1 protein, although the intensity and distribution of immunoreactivity varied across regions, strata, and cellular compartments. In stratum lucidum of CA3, mossy fiber axons were immunoreactive for NMDAR-1, which may correspond to previously hypothesized presynaptic receptors. NMDAR-1-labeled postsynaptic profiles were present in stratum radiatum of CA3 but were largely absent from stratum lucidum. Such intraneuronal segregation of glutamate receptor subunits or classes may be spatially correlated with afferent systems that exhibit laminar segregation and terminate in different portions of the postsynaptic dendritic tree. For example, in CA3 pyramidal cells, NMDA receptors are postsynaptic in distal apical dendrites (stratum radiatum), where NMDA-dependent longterm potentiation in rats is mediated by associational/commissural afferents, and are absent from proximal apical dendrites (stratum lucidum), where NMDA-independent longterm potentiation is mediated by the mossy fiber input.Excitatory neurotransmission in the central nervous system is thought to be largely mediated by the excitatory amino acids glutamate and aspartate through their effects at pre-and postsynaptic receptors (1). The molecular, physiological, and structural characteristics of various excitatory amino acid receptors have been recently reviewed (2-4) and therefore will not be comprehensively described here.The function and distribution of putative postsynaptic N-methyl-D-aspartate (NMDA) receptors have been investigated in numerous physiological, pharmacological, and in situ-hybridization studies (for review, see refs. 2 and 5); however, presynaptic NMDA receptors are also proposed to exist (6). Although, several immunohistochemical studies have recently been published on the distribution of non-NMDA-receptor subunit proteins (7-12), the cellular and synaptic distribution of the NMDA subunit proteins has yet to be characterized in detail. Such data will be crucial to the development of a precise anatomical framework for the role of NMDA receptors in synaptic transmission, long-term potentiation (LTP), and the synaptic interactions between NMDA and non-NMDA-receptor classes.LTP, a potential early step in the cellular mechanism for learning (13), has been induced in various subfields of the hippocampus in rats-including region CAl via stimulation of Schaffer collaterals, region CA3 via mossy fiber and associational/commissural fiber stimulation, and in dentate gyrus granule cells after stimulation of the perforant path (14).The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §17...

No abstract