α7-Nicotinic receptor expression and the anatomical organization of hippocampal interneurons

C3H and DBA/2 mice differ in their hippocampal inhibitory function, as measured by the inhibitory gating of pyramidal neuron response to repeated auditory stimulation. This functional difference appears to be related to differences in expression of the α7 nicotinic cholinergic receptor, which may be generally expressed by interneurons. This study examines the relationship between genetic variation in α7 receptor subunit expression and GABAergic interneuron distribution in various regions and layers of the hippocampus in the two mouse strains. Subpopulations of hippocampal interneurons in both mouse strains were found to bind [¹²⁵I]α-bungarotoxin. However, the distribution of the [¹²⁵I]α-bungarotoxin-positive hippocampal interneurons was significantly different between C3H and DBA/2 mice. In region CA1, and to a lesser extent in region CA3, DBA/2 mice had increased numbers of [¹²⁵I]α-bungarotoxin-positive neurons in stratum lacunosum-moleculare and decreased numbers in stratum oriens. Similar differences in GABAergic neuron distribution were observed in region CA1 in the two strains. C3H/DBA/2 F1 animals were backcrossed to the C3H parental strain for six generations, with selection for either the DBA/2 or C3H allelic variant of the α7 receptor gene. The distribution of [¹²⁵I]α-bungarotoxin labeling closely resembled the DBA/2 parental phenotype in animals retaining the DBA/2 allele of the α7 gene. These data suggest that the α7 receptor gene locus may influence the anatomical organization of at least a subset of hippocampal interneurons by an as yet unidentified mechanism. This difference in interneuron anatomy may also contribute to functional differences in inhibitory sensory gating between the two strains.     

Comparison of alpha7 nicotinic acetylcholine receptor development in the hippocampal formation of C3H and DBA/2 mice

A strain-specific restriction fragment length polymorphism in the alpha7 receptor gene locus has been reported to significantly affect the expression of the alpha7 subtype of nicotinic receptor in adult mouse hippocampus. The goal of the present study was to characterize the development of the alpha7 receptor in hippocampus from two mouse strains (C3H and DBA/2) with different alleles of the alpha7 receptor gene locus by using alpha-bungarotoxin (alpha-BTX) autoradiography. Binding of alpha-BTX was initially detected in fetal C3H mice on embryonic day 13 (E13) in the dorsal portion of the hippocampal anlage. In contrast, alpha-BTX binding was initially detected primarily in hippocampal area CA3 in the DBA/2 strain on E16. Binding of alpha-BTX was absent from the neuroepithelium in both strains. A marked increase in alpha-BTX binding was observed in hippocampal area CA1 and to a lesser extent in area CA3 between E18 and postnatal day 5 (P5) in neonatal C3H mice, an increase that was not observed in the DBA/2 mice. By the end of the first postnatal month, hippocampal alpha-BTX binding appeared adult-like in each strain. These data suggest that variations in the alpha7 receptor gene locus differentially influence the developmental expression of the alpha7 receptor in murine hippocampus. Therefore, the potential influence of the alpha7 receptor on developmental processes such as cell migration, dendritic elaboration and/or axonal connectivity may exhibit strain-selective differences because of the dissimilar time courses of alpha7 receptor expression in C3H and DBA/2 mice.     

Distribution and development of nicotinic acetylcholine receptor subtypes in the optic tectum of Rana pipiens

Acetylcholine allows the elicitation of visually evoked behaviors mediated by the frog optic tectum, but the mechanisms behind its effects are unknown. Although nicotinic acetylcholine receptors (nAChRs) exist in the tectum, their subtype has not been assessed. By using quantitative autoradiography, we examined the binding of [(3)H]cytisine and [(125)I]alpha-bungarotoxin in the laminated tectum. In mammalian systems, these radioligands bind with high affinity to alpha4 nAChR subunits and alpha7 nAChR subunits, respectively. [(3)H]Cytisine demonstrated high specific binding in adult frogs in retinorecipient layer 9, intermediate densities in layer 8, and low binding in layers 1-7 of the tectum. [(3)H]Cytisine binding was significantly higher in the tecta of adults than in those of tadpoles. Lesioning the optic nerve for 6 weeks decreased [(3)H]cytisine binding in layers 8/9 by 70+/-1%, whereas 6-month lesions decreased binding by 76+/-3%. Specific binding of [(125)I]alpha-bungarotoxin in adults was present only at intermediate levels in tectal layers 8 and 9, and undetectable in the deeper tectal layers. However, the nucleus isthmi, a midbrain structure reciprocally connected to the tectum, exhibited high levels of binding. There were no significant differences in tectal [(125)I]alpha-bungarotoxin binding between tadpoles and adults. Six-week lesions of the optic nerve decreased tectal [(125)I]alpha-bungarotoxin binding by 33+/-10%, but 6-month lesions had no effect. The pharmacokinetic characteristics of [(3)H]cytisine and [(125)I]alpha-bungarotoxin binding in the frog brain were similar to those demonstrated in several mammalian species. These results indicate that [(3)H]cytisine and [(125)I]alpha-bungarotoxin identify distinct nAChR subtypes in the tectum that likely contain non-alpha7 and alpha7 subunits, respectively. The majority of non-alpha7 receptors are likely associated with retinal ganglion cell terminals, whereas alpha7-containing receptors appear to have a different localization.     

Reduced expression of neuronal nicotinic acetylcholine receptors during the early stages of damage by oxidative stress in PC12 cells.

The mechanism for a large loss of neuronal nicotinic acetylcholine receptors (nAChRs) in brains with neurodegenerative diseases remains unclear. Based on our previous results of [(3)H]epibatidine binding influenced by lipid peroxidation, we suggest that nAChR deficit in neurodegenerative diseases might be related to the neurons attacked by free radicals. To further understand how free radicals influence the expression of nAChRs, we detected [(125)I]alpha-bungarotoxin binding, nAChR subunit protein and mRNA during the early stage of damage by oxidative stress in PC12 cells in the present study. The results showed that free radical insult (FeSO(4)) within the concentration range (1 -100 microM) used in the study induced dose-dependent increases in lipid peroxidation and toxicity to PC12 cells, but did not result in apoptosis or necrosis. Significant reductions in [(125)I]alpha-bungarotoxin binding site, protein level for the alpha3 and alpha7 subunits, and mRNA level for the alpha7 subunit were observed in PC12 cells treated by FeSO(4) at the concentrations without inducing cell death compared to control. Pretreatment of cultural cells with antioxidant such as Vitamin E and reduced glutathione prevented the inhibiting effect of free radicals on [(125)I]alpha-bungarotoxin and [(3)H]epibatidine bindings. The present results further demonstrate that oxidative stress might reduce the number of [(125)I]alpha-bungarotoxin binding site and selectively suppress the expression of the nAChR subunits at protein and mRNA levels during the early stages of damage in PC12 cells.     

Loss of zolpidem efficacy in the hippocampus of mice with the GABAA receptor gamma2 F77I point mutation

Zolpidem is a hypnotic benzodiazepine site agonist with some gamma-aminobutyric acid (GABA)(A) receptor subtype selectivity. Here, we have tested the effects of zolpidem on the hippocampus of gamma2 subunit (gamma2F77I) point mutant mice. Analysis of forebrain GABA(A) receptor expression with immunocytochemistry, quantitative [(3)H]muscimol and [(35)S] t-butylbicyclophosphorothionate (TBPS) autoradiography, membrane binding with [(3)H]flunitrazepam and [(3)H]muscimol, and comparison of miniature inhibitory postsynaptic current (mIPSC) parameters did not reveal any differences between homozygous gamma2I77/I77 and gamma2F77/F77 mice. However, quantitative immunoblot analysis of gamma2I77/I77 hippocampi showed some increased levels of gamma2, alpha1, alpha4 and delta subunits, suggesting that differences between strains may exist in unassembled subunit levels, but not in assembled receptors. Zolpidem (1 microm) enhanced the decay of mIPSCs in CA1 pyramidal cells of control (C57BL/6J, gamma2F77/F77) mice by approximately 60%, and peak amplitude by approximately 20% at 33-34 degrees C in vitro. The actions of zolpidem (100 nm or 1 microm) were substantially reduced in gamma2I77/I77 mice, although residual effects included a 9% increase in decay and 5% decrease in peak amplitude. Similar results were observed in CA1 stratum oriens/alveus interneurons. At network level, the effect of zolpidem (10 microm) on carbachol-induced oscillations in the CA3 area of gamma2I77/I77 mice was significantly different compared with controls. Thus, the gamma2F77I point mutation virtually abolished the actions of zolpidem on GABA(A) receptors in the hippocampus. However, some residual effects of zolpidem may involve receptors that do not contain the gamma2 subunit.     

Hippocampal CA3 pyramidal cells selectively innervate aspiny interneurons

The specific connectivity among principal cells and interneurons determines the flow of activity in neuronal networks. To elucidate the connections between hippocampal principal cells and various classes of interneurons, CA3 pyramidal cells were intracellularly labelled with biocytin in anaesthetized rats and the three-dimensional distribution of their axon collaterals was reconstructed. The sections were double-stained for substance P receptor (SPR)- or metabotropic glutamate receptor 1alpha (mGluR-1alpha)-immunoreactivity to investigate interneuron targets of the CA3 pyramidal cells. SPR-containing interneurons represent a large portion of the GABAergic population, including spiny and aspiny classes. Axon terminals of CA3 pyramidal cells contacted SPR-positive interneuron dendrites in the hilus and in all hippocampal strata in both CA3 and CA1 regions (7.16% of all boutons). The majority of axons formed single contacts (87.5%), but multiple contacts (up to six) on single target neurons were also found. CA3 pyramidal cell axon collaterals innervated several types of morphologically different aspiny SPR-positive interneurons. In contrast, spiny SPR-interneurons or mGluR-1alpha-positive interneurons in the hilus, CA3 and CA1 regions were rarely contacted by the filled pyramidal cells. These findings indicate a strong target selection of CA3 pyramidal cells favouring the activation of aspiny classes of interneurons.     

Implication of neuronal Ca2+ -sensor protein VILIP-1 in the glutamate hypothesis of schizophrenia

Post mortem studies in the hippocampus of schizophrenia patients revealed increased expression of neuronal Ca(2+)-sensor VILIP-1 (visinin-like protein) and enhanced co-localization with alpha4beta2 nAChR in interneurons. To study the pathological role of VILIP-1, particularly in interneurons, in the context of the glutamate hypothesis of schizophrenia, we have used ketamine-treated rats, a NMDA receptor hypofunction model, and hippocampal cultures as model systems for schizophrenia. Treatment with ketamine leads to enhanced VILIP-1 expression in interneurons in rat hippocampal CA1 region. In cultures glutamate treatment led to an increase in VILIP-1-positive interneurons, which is not dependent on NMDA receptor but metabotropic glutamate receptor activation. VILIP-1 mainly co-localizes with the interneuron marker calretinin, mGluR1alpha and the VILIP-1 interaction partner alpha4beta2 nAChR in hippocampal slices. Overexpression of VILIP-1 leads to enhanced nAChR-dependent inhibitory postsynaptic current (IPSC) generation by interneurons. This novel molecular link between the pathological role of mGluRs, VILIP-1 and its interaction partner alpha4beta2 nAChR by converging pathological glutamatergic and nicotinergic transmission may underlie cognitive impairments in schizophrenia.     

Immunolocalization of metabotropic glutamate receptor 1alpha (mGluR1alpha) in distinct classes of interneuron in the CA1 region of the rat hippocampus

In the hippocampal CA1 region, metabotropic glutamate subtype 1 (mGluR1) receptors have been implicated in a variety of physiological responses to glutamate, which include modulation of synaptic transmission and plasticity, as well as neuronal excitability and synchronization. The mGluR1alpha isoform is characteristically expressed only by nonprincipal cells, and it is particularly enriched in somatostatin (SS)-containing interneurons in stratum oriensalveus. Anatomical and physiological data have indicated the presence of mGluR1alpha in several distinct classes of interneurons with their somata located also in strata pyramidale, radiatum, and lacunosum moleculare. Each different interneuron subtype, as defined by functionally relevant criteria, including input/ output characteristics and expression of selective molecular markers, subserves distinct functions in local hippocampal circuits. We have investigated which of the different CA1 interneuron classes express mGluR1alpha by immunofluorescent labeling, combining antibodies to mGluR1alpha, calcium-binding proteins, and neuropeptides, and by intracellular labeling in vitro. Several types of interneuron that are immunopositive for mGluR1alpha each targeted different domains of pyramidal cells and included (1) O-LM inter-neurons, found to coexpress both SS and parvalbumin (PV); (2) interneurons with target selectivity for other interneurons, expressing vasoactive intestinal polypeptide (VIP) and/or the calcium-binding protein calretinin; (3) procholecystokinin-immunopositive interneurons probably non-basket and dendrite-targeting; and (4) an as-yet unidentified SS-immunoreactive but PV-immunonegative interneuron class, possibly corresponding to oriens-bistratified cells. Estimation of the relative proportion of mGluR1alpha-positive interneurons showed 43%, 46%, and 30% co-labeling with SS, VIP, or PV, respectively. The identification of the specific subclasses of CA1 interneurons expressing mGluR1alpha provides the network basis for assessing the contribution of this receptor to the excitability of the hippocampus.     

An autoradiographic study of the distribution of binding sites for the novel alpha7-selective nicotinic radioligand [3H]-methyllycaconitine in the mouse brain.

[3H]-Methyllycaconitine ([3H]-MLA) is a new radioligand with selectivity for alpha7-type neuronal nicotinic acetylcholine receptors (nAChRs). In our previous study [Davies, A.R.L., Hardick, D.J., Blagbrough, I.S., Potter, B.V.L., Wolstenholme, A.J. & Wonnacott, S. (1999) Neuropharmacology, 38, 679-690], this radioligand labelled a single class of site in rat brain membranes; its pharmacology and distribution in crudely dissected brain regions closely paralleled that of the well-established alpha7-ligand [125I]-alpha-bungarotoxin. However, a small population of [3H]-MLA binding sites was apparently insensitive to alpha-bungarotoxin. Here we have extended the study to mouse brain, using autoradiography to examine the distribution of [3H]-MLA and [125I]-alpha-bungarotoxin binding sites. [3H]-MLA labelled a single class of site in mouse brain membranes with a KD of 2.2 nM and a Bmax of 45.6 fmol/mg protein. Specific binding, defined by unlabelled MLA (Ki = 0.69 nM), was completely inhibited by (-)-nicotine (Ki = 1.62 microM), whereas alpha-bungarotoxin inhibited only 85% of specific binding (Ki = 3.5 nM). The distributions of [125I]-alpha-bungarotoxin and [3H]-MLA binding sites were compared by autoradiography, and binding was quantitated in 72 brain regions. Binding of both radioligands was highly correlated, with highest densities in the dorsal tegmental nucleus of the pons, colliculi and hippocampus. Serial sections labelled with [3H]-MLA in the absence or presence of unlabelled MLA or alpha-bungarotoxin provided no evidence for any alpha-bungarotoxin-resistant binding. The results are discussed in terms of binding sites that are inaccessible to alpha-bungarotoxin in membrane preparations. This study demonstrates the utility of [3H]-MLA for characterization of alpha7-type nicotinic receptors in mammalian brain, and suggests that it labels a population identical to that defined by [125I]-alpha-bungarotoxin.     

Higher expression of alpha7 nicotinic acetylcholine receptors in human fetal compared to adult brain

Neuronal nicotinic acetylcholine receptors are thought to be involved in regulation of several processes during neurogenesis of the brain. In this study the expression of the alpha7 nicotinic acetylcholine receptor subtype was investigated in human fetal (9-11 weeks of gestation), middle-aged (28-51 years) and aged (68-94 years) medulla oblongata, pons, frontal cortex, and cerebellum. The specific binding of the alpha7 receptor antagonist [(125)I]alpha-bungarotoxin was significantly higher in fetal than in both middle-aged and aged medulla oblongata and aged pons. No significant decrease in [(125)I]alpha-bungarotoxin binding sites was observed from fetal to adult cortex and cerebellum. The alpha7 mRNA expression was significantly higher in all fetal brain regions investigated, except for aged cortex, than in corresponding middle-aged and aged tissue. The high expression of alpha7 nicotinic acetylcholine receptors in fetal compared to adult brain supports the view that these receptors play an important role during brain development.     

Effects of Abeta1-42 fibrils and of the tetrapeptide Pr-IIGL on the phosphorylation state of the tau-protein and on the alpha7 nicotinic acetylcholine receptor in vitro

In order to investigate the possible links connecting beta-amyloid (Abeta) accumulation, tau-hyperphosphorylation and nicotinic receptor expression, rat embryonic primary hippocampal cultures were incubated with amyloidogenic peptides. Exposure to 0.5 microm fibrillar Abeta(1-42) for 3 days caused retraction of dendrites, shrinkage of cell bodies and a decrease in the expression of microtubule-associated proteins 2b (MAP2b), without affecting the total number of neurons and their viability. No impact on the tau-phosphorylation sites Ser-202, Thr231/Ser235, Ser262 and Ser396/Ser404 was found. The total number of homomeric alpha7-nicotinic receptors (alpha7-nAChRs) and their affinity for [(125)I]alpha-bungarotoxin remained unaltered. Upon incubation with the putatively protective tetrapeptide propionyl-isoleucine-isoleucine-glycine-leucine (Pr-IIGL), an analogue of the region [31-34] of Abeta, cell bodies were swollen in the region of the apical dendrite. These morphological alterations, different from those elicited by Abeta(1-42), did not involve MAP2 expression changes. In contrast to Abeta(1-42), Pr-IIGL caused a massive hyperphosphorylation of the tau-protein at Ser-202 and at Ser396/Ser404. The total number of homomeric alpha7-nAChRs and their affinity for [(125)I]alpha-bungarotoxin were unaffected. In conclusion, the present results show a toxic effect of Abeta(1-42) on the cytoskeletal structure at concentrations normally present in the brains of Alzheimer's disease patients, but raise some doubts about the role of Abeta(1-42) fibrils as a direct trigger of tau-hyperphosphorylation. The tetrapeptide Pr-IIGL cannot be considered protective with regard to cell morphology. Although it prevents the Abeta(1-42)-induced retraction of dendrites, it exhibits other toxic properties. The homomeric alpha7-nAChRs were not affected either by Abeta(1-42) incubation or by Pr-IIGL-induced tau-hyperphosphorylation.     

Hippocampal mossy fiber distribution and long-term potentiation in two inbred mouse strains

We studied long-term potentiation in the inbred mouse strains DBA/2 and C3H/He known to be different in both hippocampal mossy fiber distribution and several aspects of learning. Tetanic stimulation of mossy fibers resulted in a significantly stronger increase of the population spike amplitude in the CA3 pyramidal cell layer of C3H mice. This result suggests that the extent of the CA3 hippocampal mossy fiber projection correlates with synaptic efficacy in mice.     

Changes in hippocampal GABAA receptor subunit composition in bipolar 1 disorder

Postmortem CNS studies have suggested an uncoupling of the gamma-aminobutyric acid (GABA) and benzodiazepine binding sites on the hippocampal GABA(A) receptor in schizophrenia. The GABA(A) receptor is an assembly of discrete subunits that form a ligand-gated ion channel, the binding characteristics of which are defined by receptor subunit composition. Thus, a likely explanation for an uncoupling between the GABA and benzodiazepine binding sites on the GABA(A) receptor would be a change in receptor subunit composition. To test this hypothesis we measured the density of GABA ([(3)H]muscimol) and benzodiazepine ([(3)H]flumazenil) binding sites on the GABA(A) receptor in hippocampi, obtained postmortem, from schizophrenic, bipolar I disorder and control subjects. In addition, we measured the amount of [(3)H]flumazenil binding that could be displaced with zolpidem and clonazepam. Levels of both [(3)H]muscimol and [(3)H]flumazenil binding were significantly decreased in part of the CA2 from subjects with schizophrenia; the decrease in [(3)H]flumazenil being due to decreases in both zolpidem-sensitive and -insensitive radioligand binding. There were complex regionally specific changes in [(3)H]muscimol binding in the hippocampus from subjects with bipolar I disorder but there were no significant changes in the overall levels of [(3)H]flumazenil binding. There were significant decreases in zolpidem-sensitive and increases in zolpidem-insensitive [(3)H]flumazenil binding in most regions of the sections of the hippocampal formation studied in bipolar I disorder. Unlike [(3)H]flumazenil, zolpidem does not bind to the alpha5 subunit of the GABA(A) receptor; therefore, we postulate that there is an increase in GABA(A) receptors containing alpha5 subunit in the hippocampus from subjects with bipolar I disorder.     

Multistep expression and assembly of neuronal nicotinic receptors is both host-cell- and receptor-subtype-dependent

We tested the hypothesis that the folding, assembly and insertion of neuronal nicotinic receptors are critically dependent on the host cell line. We used recombinant adenoviruses encoding either the rat alpha7, alpha4 or beta2 subunits in which expression of the subunit is controlled by a tetracycline-dependent promoter to screen five cell lines (GH4C1, SH-EP1, CV1, SN-56, and CHO-CAR). All five lines do not express detectable nicotinic receptor but do express receptor for human adenovirus, and all expressed mRNA for alpha7, alpha4 and beta2 subunits when infected with viruses. Each cell line expressed varying levels of alpha4beta2 receptors that bound [3H]cytisine, but only the GH4C1 and SH-EP1 cell lines expressed either surface or internal alpha7 receptors that bound [125I]alpha-bungarotoxin ([125I]alpha-BGT). All five cell lines expressed a 60 kDa protein immunoblotted by anti-alpha7 antibodies when infected with the alpha7 virus, presumably representing unassembled alpha7 subunits. In addition, GH4C1 cells expressed over 10-fold more surface alpha7 receptor than SH-EP1 cells, even though the total alpha7 receptor in the two cell lines was similar. Sedimentation experiments indicate that SH-EP1 cells only partially assemble alpha7 receptors compared with GH4C1 cells and control alpha7 from rat brain. These data suggest that not only is surface alpha7 receptor expression a multistep process, but that each step may involve cell-specific assembly factors.     

Nicotinic receptor abnormalities in Alzheimer's disease.

Loss of cortical nicotinic acetylcholine receptors with high affinity for agonists (20-50%) in patients with Alzheimer's disease is a common finding. Recent immunochemical analyses indicate that this deficit is predominantly associated with the loss of alpha4 subunits (30-50%), although modest reductions of alpha3 may occur in some individuals (25-29%). No reduction of beta2 subunit protein expression or levels of alpha3 and alpha4 messenger RNA has been reported. Decline in cortical [(125)I]alpha-bungarotoxin binding and alpha7 protein expression does not appear to be as extensive or widespread as the loss of alpha4 (0-40%), with no reduction in messenger RNA expression. In the thalamus, there was a trend for reduced [(3)H]nicotine binding in the majority of nuclei (0-20%) in Alzheimer's disease; however, there was a significant decline in [(125)I]alpha-bungarotoxin binding in the reticular nucleus. In the striatum [(3)H]nicotine binding was reduced in Alzheimer's disease, and although neuroleptic medication accentuated this change, it occurred in those free of neuroleptics. Changes in nicotinic acetylcholine receptors in Alzheimer's disease are distinct from those in normal aging and are likely to contribute to clinical features and possibly neuropathology.     

125I]Iodomethyllycaconitine binds to alpha7 nicotinic acetylcholine receptors in monkey brain

We examined the binding of the novel nicotinic acetylcholine receptor (nAChR) ligand [125I]iodomethyllycaconitine (iodoMLA) in the brains of M. cynomologous (macaque) monkeys. [125I]iodoMLA bound throughout the brain with the greatest density in the thalamus and moderate intensity in the basal ganglia and cortical regions. The Kd and Bmax in whole brain tissue were similar whether 1 mM nicotine (Kd 33.25 +/- 15.17 nM, Bmax 5.80 +/- 1.06 fmol/mg) or 2 microM of the alpha7-selective antagonist alpha-bungarotoxin (Kd 46.12 +/- 18.45 nM, Bmax 6.30 +/- 1.06 fmol/mg) was used for nonspecific binding. The subtype-selectivity of this ligand was further studied with competition binding studies using nicotine, alpha-bungarotoxin and noniodinated MLA. Each ligand completely inhibited [125I]iodoMLA binding throughout the monkey brain, with Ki values of 2.23 +/- 0.85 microM for nicotine, 2.72 +/- 1.71 nM for alpha-bungarotoxin and 1.83 +/- 0.35 nM MLA in the caudate and 2.03 +/- 1.14 microM, 2.65 +/- 0.86 nM and 3.32 +/- 0.71 nM, respectively, in the putamen. The alpha3beta2/alpha6*-selective antagonist alpha-conotoxin MII failed to inhibit [125I]iodoMLA binding in any brain region. In monkeys with cognitive deficits resulting from 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine administration, [125I]iodoMLA binding was significantly increased in the striatum, similar to results previously observed for [125I]alpha-bungarotoxin. These results suggest that, under the present experimental conditions, [125I]iodoMLA was selective for alpha7-containing nAChRs and did not bind to alpha6-containing nAChRs. This radioligand may be a useful tool for selectively imaging alpha7-containing nAChRs in vivo.     

Glutamic acid decarboxylase-67-positive hippocampal interneurons undergo a permanent reduction in number following kainic acid-induced degeneration of ca3 pyramidal neurons

Kainic acid (KA)-induced degeneration of CA3 pyramidal neurons leads to synaptic reorganization and hyperexcitability in both dentate gyrus and CA1 region of the hippocampus. We hypothesize that the substrate for hippocampal inhibitory circuitry incurs significant and permanent alterations following degeneration of CA3 pyramidal neurons. We quantified changes in interneuron density (N(v)) in all strata of the dentate gyrus and the CA1 and CA3 subfields of adult rats at 1, 4, and 6 months following intracerebroventricular (icv) KA administration, using glutamic acid decarboxylase-67 (GAD-67) immunocytochemistry. At 1 month postlesion, GAD-67-positive interneuron density was significantly reduced in all strata of every hippocampal region except stratum pyramidale of CA1. The reduction in GAD-67-positive interneuron density either persisted or exacerbated at 4 and 6 months postlesion in every stratum of all hippocampal regions. Further, the soma of remaining GAD-67-positive interneurons in dentate gyrus and CA3 subfield showed significant hypertrophy. Thus, both permanent reductions in the density of GAD-67-positive interneurons in all hippocampal regions and somatic hypertrophy of remaining GAD-67-positive interneurons in dentate gyrus and CA3 subfield occur following icv KA. In contrast, the density of interneurons visualized with Nissl in CA1 and CA3 regions was nearly equivalent to that in the intact hippocampus at all postlesion time points. Collectively, these results suggest that persistent reductions in GAD-67-positive interneuron density observed throughout the hippocampus following CA3 lesion are largely due to a permanent loss of GAD-67 expression in a significant fraction of interneurons, rather than widespread degeneration of interneurons. Nevertheless, a persistent decrease in interneuron activity, as evidenced by permanent down-regulation of GAD-67 in a major fraction of interneurons, would likely enhance the degree of hyperexcitability in the CA3-lesioned hippocampus.     

Nicotine and clozapine selectively reverse a PCP-induced deficit of PPI in BALB/cByJ but not NMRI mice: comparison with risperidone

Schizophrenic patients have deficits in prepulse inhibition (PPI) that may be alleviated by smoking/nicotine. The effect of nicotinic agents on PPI in rodents is equivocal and few studies in mice have been reported. Thus, we assessed nicotine's (0.03-1mg/kg) effect on PPI in five mouse strains with no effects. We next determined if nicotine would reverse a phencyclidine (PCP)-induced deficit of PPI in BALB/cByJ and NMRI mice. BALB/cByJ mice have a low density of [(125)I]alpha-bungaratoxin binding in the hippocampus and poor inhibitory gating of auditory evoked potentials (AEPs), a model related to PPI. At 1mg/kg, nicotine selectively reversed the PCP-induced deficit of PPI in BALB/cByJ mice. The pharmacokinetic profile of nicotine (T(1/2), C(max), T(max) and AUC) was identical in both strains, obviating this as a factor for the strain-dependent effect observed. Moreover, 1mg/kg nicotine inhibited in vivo [(3)H]epibatidine binding with the same time-course in both strains, indicating no difference in brain "kinetics". Since high doses of nicotine were effective in BALB/cByJ mice a role for low-affinity nicotinic receptors, e.g. alpha(7) receptors, is plausible. Clozapine, but not risperidone, also only reversed the PCP deficit of PPI in BALB/cByJ. Clozapine and nicotine also enhance inhibitory gating of AEPs in DBA/2 mice, and clozapine's effect is antagonized by an alpha(7) antagonist. Our data and previous evidence possibly suggest a role for low-affinity nicotinic receptors in the effects of clozapine and nicotine. Furthermore, BALB/cByJ mice may represent a model to test the effects of nicotinic agents acting at low-affinity nicotinic receptors.