Gilles Tamagnan

Human Tobacco Smokers in Early Abstinence Have Higher Levels of β2* Nicotinic Acetylcholine Receptors than Nonsmokers

Nicotine, the addictive chemical in tobacco smoke, initiates its actions in brain through nicotinic acetylcholine receptors (nAChRs). In particular, nAChRs containing β2-subunits (β2*-nAChRs) the most prevalent subtype, mediate the reinforcing properties of nicotine. We hypothesized that abnormal numbers of β2*-nAChRs during early abstinence contribute to the perpetuation of addiction to tobacco smoking. Using molecular imaging, specifically single-photon emission computed tomography with the nAChR agonist radiotracer [123I]5-IA-85380 ([123I]5-IA), we imaged β2*-nAChR availability in human smokers. First, using nonhuman primates treated chronically with nicotine, we estimated the time interval necessary for smokers to abstain from smoking so that residual nicotine would not interfere with [123I]5-IA binding to the β2*-nAChR as ∼7 d. Thus, we imaged human smokers at 6.8 ± 1.9 d (mean ± SD) of abstinence. Abstinence was confirmed by daily assessments of urinary cotinine and expired carbon monoxide levels. In smokers, [123I]5-IA uptake was significantly higher throughout the cerebral cortex (26–36%) and in the striatum (27%) than in nonsmokers, suggesting higher β2*-nAChR in recently abstinent smokers. β2*-nAChR availability in recently abstinent smokers correlated with the days since last cigarette and the urge to smoke to relieve withdrawal symptoms but not the severity of nicotine dependence, severity of nicotine withdrawal, or the desire to smoke. Higher brain β2*-nAChR during early abstinence indicates that, when smokers quit smoking, they do so in the face of a significant increase in the receptors normally activated by nicotine. Greater β2*-nAChR availability during early abstinence may impact the ability of smokers to maintain abstinence.

β2-Nicotinic Acetylcholine Receptor Availability During Acute and Prolonged Abstinence From Tobacco Smoking

CONTEXT Available levels of nicotinic acetylcholine receptors containing the beta(2) subunit (beta(2)*-nAChR) are higher in recently abstinent tobacco smokers compared with participants who never smoked. Variations in beta(2)*-nAChR availability during the course of abstinence may be related to the urge to smoke, the extent of nicotine withdrawal, and successful abstinence. OBJECTIVE To examine changes in beta(2)*-nAChR availability during acute and prolonged abstinence from tobacco smoking and to determine how changes in beta(2)*-nAChR availability were related to clinical features of tobacco smoking. DESIGN Tobacco smokers participated in up to 4 iodide 123-labeled 5-iodo-A-85380 ([(123)I]5-IA) single-photon emission computed tomography (SPECT) scans during abstinence at 1 day (n = 7) and 1 (n = 17), 2 (n = 7), 4 (n = 11), and 6 to 12 (n = 6) weeks. Age-matched nonsmokers participated in a single [(123)I]5-IA SPECT scan. All participants completed 1 magnetic resonance imaging study. SETTING Academic imaging center. PARTICIPANTS Tobacco smokers (n = 19) and an age-matched nonsmoker comparison group (n = 20). Main Outcome Measure The [(123)I]5-IA SPECT images were converted to distribution volume and were analyzed using regions of interest. RESULTS Compared with nonsmokers, beta(2)*-nAChR availability in the striatum, cortex, and cerebellum of smokers was not different at 1 day of abstinence, was significantly higher at 1 week of abstinence, and was not different at 4 or at 6 to 12 weeks of abstinence. In smokers, beta(2)*-nAChR availability was significantly lower in the cortex and cerebellum at 6 to 12 weeks compared with 1 week of abstinence. In addition, cerebellar beta(2)*-nAChR availability at 4 weeks of abstinence was positively correlated with craving on the day of the SPECT scan. CONCLUSIONS These data suggest that higher beta(2)*-nAChR availability persists up to 1 month of abstinence and normalizes to nonsmoker levels by 6 to 12 weeks of abstinence from tobacco smoking. These marked and persistent changes in beta(2)*-nAChR availability may contribute to difficulties with tobacco cessation.

Positive Allosteric Modulation of Metabotropic Glutamate 5 (mGlu5) Receptors Reverses N-Methyl-D-Aspartate Antagonist-Induced Alteration of Neuronal Firing in Prefrontal Cortex

BACKGROUND Several lines of evidence suggest that N-methyl-D-aspartate (NMDA) receptor hypofunction may be associated with schizophrenia. Activation of metabotropic glutamate 5 (mGlu5) receptors enhances NMDA receptor mediated currents in vitro, implying that allosteric modulation of mGlu5 receptors may have therapeutic efficacy for schizophrenia. The aim of this study was to determine if positive allosteric modulators of mGlu5 receptors are effective in reversing two cellular effects of NMDA receptor antagonists that are relevant to schizophrenia: increases in corticolimbic dopamine neurotransmission and disruption of neuronal activity in the prefrontal cortex (PFC). METHODS In freely moving rats, we measured the effects of the positive modulator of mGlu5 receptor 3-cyano-N-(1,3-diphenyl-1H-pyrazol-5-yl)benzamide (CDPPB) alone or in combination with the NMDA antagonist MK801 on 1) spontaneous firing and bursting of medial PFC (mPFC) neurons, and 2) dopamine release as measured by microdialysis in the mPFC and nucleus accumbens (NAc). RESULTS The predominant effect of CDPPB on mPFC neurons was excitatory, leading to an overall excitatory population response. Pretreatment with CDPPB prevented MK801-induced excessive firing and reduced spontaneous bursting. In contrast, CDPPB had no significant effect on basal dopamine release as compared with control rats and did not alter MK801-induced activation of dopamine release in the mPFC and NAc. CONCLUSIONS These results show that positive modulation of mGlu5 receptors reverses the effects of noncompetitive NMDA antagonists on cortical neuronal firing without affecting dopamine neurotransmission. Thus, these compounds may be effective in ameliorating PFC mediated behavioral abnormalities that results from NMDA receptor hypofunction.

Sex Differences in Diencephalon Serotonin Transporter Availability in Major Depression

BACKGROUND Major depression is more prevalent in women than men. The present study evaluated if previous findings that demonstrated decreased 5-hydroxytryptamine (5-HT) transporter availability in depressed patients would be confirmed in a larger sample and also evaluated sex differences. METHODS Depressed (n = 32) and healthy subjects (n = 32), including 16 pairs of women and men, participated in an iodine-123-2 beta-carbomethoxy-3beta-(4-iodophenyltropane) ([(123)I]beta-CIT) single photon emission computed tomography (SPECT) and a magnetic resonance imaging (MRI) scan. Participants were administered [(123)I]beta-CIT (225.7 +/- 3.7 MBq) and imaged 23.0 +/- 1.6 hours later. Statistical analyses included analysis of variance and a regression analysis of the main and interactive effects of age, sex, and depression. RESULTS Overall, depressed patients demonstrated 12% lower diencephalon and no change in striatal or brainstem [(123)I]beta-CIT uptake. Significant age by sex, sex by depression, and age by sex by depression interactions were noted due to 22% lower diencephalon [(123)I]beta-CIT uptake in depressed women compared with less than a 1% decrease in depressed men. CONCLUSIONS As observed previously, diencephalon 5-HT transporter availability is decreased in depressed patients. However, the decrease appears to be sex-specific and age-dependent. These findings suggest that serotonergic mechanisms mediating depressed mood differ between men and women in an age-dependent manner and may explain why young women respond better to treatment with selective serotonin reuptake inhibitor (SSRI) antidepressants.

A Novel Family of Potent Negative Allosteric Modulators of Group II Metabotropic Glutamate Receptors

Group II metabotropic glutamate receptors (mGluRs), mGluR2 and mGluR3, play a number of important roles in mammalian brain and represent exciting new targets for certain central nervous system disorders. We now report synthesis and characterization of a novel family of derivatives of dihydrobenzo[1,4]diazepin-2-one that are selective negative allosteric modulators for group II mGluRs. These compounds inhibit both mGluR2 and mGluR3 but have no activity at group I and III mGluRs. The novel mGluR2/3 antagonists also potently block mGluR2/3-mediated inhibition of the field excitatory postsynaptic potentials at the perforant path synapse in hippocampal slices. These compounds induce a rightward shift and decrease the maximal response in the glutamate concentration-response relationship, consistent with a noncompetitive antagonist mechanism of action. Furthermore, radioligand binding studies revealed no effect on binding of the orthosteric antagonist [3H]LY341495 [2S-2-amino-2-(1S,2S-2-carboxycyclopropan-1-yl)-3-(xanth-9-yl)propionic acid]. Site-directed mutagenesis revealed that a single point mutation in transmembrane V (N735D), previously shown to be an important residue for potentiation activity of the mGluR2 allosteric potentiator LY487379 [N-(4-(2-methoxyphenoxy)phenyl)-N-(2,2,2-trifluoroethylsulfonyl)pyrid-3-ylmethylamine], is not critical for the inhibitory activity of negative allosteric modulators of group II mGluRs. However, this single mutation in human GluR2 almost completely blocked the enhancing activity of biphenyl-indanone A, a novel allosteric potentiator of mGluR2. Our data suggest that these two positive allosteric modulators of mGluR2 may share a common binding site and that this site may be distinct from the binding site for the new negative allosteric modulators of group II mGluRs.

Persistent β2*-Nicotinic Acetylcholinergic Receptor Dysfunction in Major Depressive Disorder

BACKGROUND Modulation of nicotinic acetylcholine receptors (nAChRs), specifically those containing the β2 subunit, may be effective in treating patients with major depressive disorder. Using [123I]5-I-A-85380 single photon emission computed tomography (SPECT), the authors studied the availability of β2-subunit-containing nAChRs (β2*-nAChRs) in depressed patients. To understand its molecular basis, the authors also studied β2*-nAChR binding in postmortem brain samples from depressed subjects. METHOD The participants were 23 medication-free, nonsmoking subjects with familial, early-onset depression (eight acutely ill and 15 recovered) and 23 age- and gender-matched nonsmoking comparison subjects. Each received one [123I]5-I-A-85380 SPECT scan and an MRI scan. The availability of β2*-nAChRs was quantified as VT/fP. Postmortem analysis of β2*-nAChR binding was conducted with [123I]5-I-A-85380 on prefrontal cortex samples from 14 depressed subjects and 14 age-matched comparison subjects. RESULTS The β2*-nAChR availability in both the acutely ill and recovered depressed subjects was significantly lower across all brain regions than in the respective comparison subjects, and it was lower in the acutely ill subjects than in those who were recovered. In the depressed patients, β2*-nAChR availability was significantly correlated with lifetime number of depressive episodes, trauma score, and anxiety score. There were no differences in β2*-nAChR number between groups in the postmortem study. CONCLUSIONS Depressed patients have lower β2*-nAChR availability than do healthy subjects. The difference between β2*-nAChR availability in vivo and in post-mortem samples may be analogous to data with dopaminergic PET ligands and dopamine receptor availability; lower receptor availability for the SPECT ligand could be caused by greater endogenous acetylcholine.

Radiosynthesis of [18F] N-3-fluoropropyl-2-β-carbomethoxy-3-β-(4-iodophenyl) nortropane and the first human study with positron emission tomography☆

A procedure for the routine preparation of [18F]FP-CIT has been developed. Purification of the final product was achieved by preparative HPLC using phenethyl column without decomposition or epimerization. [18F] labeled-N-fluoropropyl-2 beta-carbomethoxy-3 beta-(4-iodophenyl) nortropane was prepared and PET imaging was performed on human subjects. A high uptake into striatal regions was observed. HPLC plasma analysis using [18F]FP-CIT indicated the presence of only one metabolite. By directly comparing the behavior of these three radiotracers ([18F]DOPA, [123I]FP-CIT, and [18F]FP-CIT) in the same subjects, we can enhance our understanding of the dopaminergic system as well as the relative potential of these techniques in a clinical research setting.

Metabotropic glutamate receptor 5 antagonist protects dopaminergic and noradrenergic neurons from degeneration in MPTP-treated monkeys.

Degeneration of the dopaminergic nigrostriatal system and of noradrenergic neurons in the locus coeruleus are important pathological features of Parkinsons disease. There is an urgent need to develop therapies that slow down the progression of neurodegeneration in Parkinsons disease. In the present study, we tested whether the highly specific metabotropic glutamate receptor 5 antagonist, 3-[(2-methyl-1,3-thiazol-4-yl) ethynyl] pyridine, reduces dopaminergic and noradrenergic neuronal loss in monkeys rendered parkinsonian by chronic treatment with low doses of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine. Weekly intramuscular 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine injections (0.2-0.5 mg/kg body weight), in combination with daily administration of 3-[(2-methyl-1,3-thiazol-4-yl) ethynyl] pyridine or vehicle, were performed until the development of parkinsonian motor symptoms in either of the two experimental groups (1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine/3-[(2-methyl-1,3-thiazol-4-yl) ethynyl] pyridine versus 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine/vehicle). After 21 weeks of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine treatment, all 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine/vehicle-treated animals displayed parkinsonian symptoms, whereas none of the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine/3-[(2-methyl-1,3-thiazol-4-yl) ethynyl] pyridine-treated monkeys were significantly affected. These behavioural observations were consistent with in vivo positron emission tomography dopamine transporter imaging data, and with post-mortem stereological counts of midbrain dopaminergic neurons, as well as striatal intensity measurements of dopamine transporter and tyrosine hydroxylase immunoreactivity, which were all significantly higher in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine/3-[(2-methyl-1,3-thiazol-4-yl) ethynyl] pyridine-treated animals than in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine/vehicle-treated monkeys. The 3-[(2-methyl-1,3-thiazol-4-yl) ethynyl] pyridine treatment also had a significant effect on the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-induced loss of norepinephrine neurons in the locus coeruleus and adjoining A5 and A7 noradrenaline cell groups. In 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine/vehicle-treated animals, almost 40% loss of tyrosine hydroxylase-positive norepinephrine neurons was found in locus coeruleus/A5/A7 noradrenaline cell groups, whereas the extent of neuronal loss was lower than 15% of control values in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine/3-[(2-methyl-1,3-thiazol-4-yl) ethynyl] pyridine-treated monkeys. Our data demonstrate that chronic treatment with the metabotropic glutamate receptor 5 antagonist, 3-[(2-methyl-1,3-thiazol-4-yl) ethynyl] pyridine, significantly reduces 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine toxicity towards dopaminergic and noradrenergic cell groups in non-human primates. This suggests that the use of metabotropic glutamate receptor 5 antagonists may be a useful strategy to reduce degeneration of catecholaminergic neurons in Parkinsons disease.

A novel class of positive allosteric modulators of metabotropic glutamate receptor subtype 1 interact with a site distinct from that of negative allosteric modulators.

We recently reported a novel class of compounds, represented by 3-cyano-N-(1,3-diphenyl-1H-pyrazol-5-yl)benzamide (CD-PPB), that act as positive allosteric modulators (potentiators) of metabotropic glutamate receptor (mGluR) subtype 5. Studies of CDPPB analogs revealed that some compounds in this series serve also as positive allosteric modulators of mGluR1. Although CDPPB is selective for mGluR5 relative to other mGluR subtypes, several CDPPB analogs also showed 2.5-fold potentiation of glutamate-induced calcium transients in cells expressing mGluR1 at 10 μM, with 4-nitro-N-(1,4-diphenyl-1H-pyrazol-5-yl)benzamide (VU-71) being selective for mGluR1. In previous studies, we found that two structural classes of mGluR5-selective allosteric potentiators, including CDPPB, share a common binding site with the allosteric mGluR5 antagonist 2-methyl-6-(phenylethynyl)pyridine. Negative allosteric modulators of mGluR1, regardless of structural class, have been reported to bind to a common allosteric antagonist site on this receptor. However, neither the novel CDPPB analogs nor previously identified allosteric mGluR1 potentiators [e.g., (S)-2-(4-fluorophenyl)-1-(toluene-4-sulfonyl)pyrrolidine (Ro 67-7476), ethyl diphenylacetylcarbamate (Ro 01-6128), and butyl (9H-xanthene-9-carbonyl)carbamate (Ro 67-4853)] displaced the binding of [3H]1-(3,4-dihydro-2H-pyrano[2,3-b]quinolin-7-yl)-2-phenyl-1-ethanone (R214127), a high-affinity radioligand for the allosteric antagonist site on mGluR1 at concentrations several orders of magnitude higher than those required to induce allosteric potentiation of mGluR1 responses. These data suggest that allosteric potentiators of mGluR1 act at a site that is distinct from that of allosteric antagonists of mGluR1. Site-directed mutagenesis revealed that valine at position 757 in transmembrane V of mGluR1a is crucial for the activity of multiple classes of allosteric mGluR1 potentiators.

Local anesthetic effects of cocaethylene and isopropylcocaine on rat peripheral nerves.

Cocaethylene is a naturally occurring cocaine derivative that has been used as a tool in both clinical studies of cocaine reward and as a potential model compound for agonist substitution therapy in cocaine dependence. It is equipotent to cocaine at inhibiting dopamine uptake in-vitro and in-vivo. Because it has been reported that local anesthetic properties may influence the reinforcing effects of dopamine uptake inhibitors, we investigated the local anesthetic properties of cocaethylene as well as isopropylcocaine, another potential pharmacological tool in studies of cocaine reward and agonist substitution therapy. We compared the efficacy of nerve impulse blockade by lidocaine, cocaine, cocaethylene and isopropylcocaine using rat sciatic nerves and dorsal roots (DRs). Nerves were placed in a modified sucrose gap chamber and repetitively stimulated at high frequency. The amplitude of compound action potentials (CAPs) at the beginning and end of each stimulus train was measured before and after exposure to each compound. All compounds produced concentration-dependent and use-dependent decrements in CAP amplitude, but cocaethylene and isopropylcocaine at medium to high concentration (0.375-1.875 mM) showed a more prolonged block after washout relative to cocaine or lidocaine. Patch clamp studies on dorsal root ganglion (DRG) neurons indicated a use-dependent blockade of sodium channels. These studies provide a more complete understanding of the pharmaocology of potential agonist treatment candidates, and suggest a mechanism whereby cocaethylene produces a decreased euphoria in humans compared to cocaine.