Publications by authors named "Ines Ibañez-Tallon"

34 Publications

β4-Nicotinic Receptors Are Critically Involved in Reward-Related Behaviors and Self-Regulation of Nicotine Reinforcement.

J Neurosci 2020 04 17;40(17):3465-3477. Epub 2020 Mar 17.

Université de Bordeaux, Institut de Neurosciences Cognitives et Intégratives d'Aquitaine, 33615 Pessac, France

Nicotine addiction, through smoking, is the principal cause of preventable mortality worldwide. Human genome-wide association studies have linked polymorphisms in the gene cluster, coding for the α, α, and β4 nicotinic acetylcholine receptor (nAChR) subunits, to nicotine addiction. β4*nAChRs have been implicated in nicotine withdrawal, aversion, and reinforcement. Here we show that β4*nAChRs also are involved in non-nicotine-mediated responses that may predispose to addiction-related behaviors. β4 knock-out (KO) male mice show increased novelty-induced locomotor activity, lower baseline anxiety, and motivational deficits in operant conditioning for palatable food rewards and in reward-based Go/No-go tasks. To further explore reward deficits we used intracranial self-administration (ICSA) by directly injecting nicotine into the ventral tegmental area (VTA) in mice. We found that, at low nicotine doses, β4KO self-administer less than wild-type (WT) mice. Conversely, at high nicotine doses, this was reversed and β4KO self-administered more than WT mice, whereas β4-overexpressing mice avoided nicotine injections. Viral expression of β4 subunits in medial habenula (MHb), interpeduncular nucleus (IPN), and VTA of β4KO mice revealed dose- and region-dependent differences: β4*nAChRs in the VTA potentiated nicotine-mediated rewarding effects at all doses, whereas β4*nAChRs in the MHb-IPN pathway, limited VTA-ICSA at high nicotine doses. Together, our findings indicate that the lack of functional β4*nAChRs result in deficits in reward sensitivity including increased ICSA at high doses of nicotine that is restored by re-expression of β4*nAChRs in the MHb-IPN. These data indicate that β4 is a critical modulator of reward-related behaviors. Human genetic studies have provided strong evidence for a relationship between variants in the gene cluster and nicotine addiction. Yet, little is known about the role of β4 nicotinic acetylcholine receptor (nAChR) subunit encoded by this cluster. We investigated the implication of β4*nAChRs in anxiety-, food reward- and nicotine reward-related behaviors. Deletion of the β4 subunit gene resulted in an addiction-related phenotype characterized by low anxiety, high novelty-induced response, lack of sensitivity to palatable food rewards and increased intracranial nicotine self-administration at high doses. Lentiviral vector-induced re-expression of the β4 subunit into either the MHb or IPN restored a "stop" signal on nicotine self-administration. These results suggest that β4*nAChRs provide a promising novel drug target for smoking cessation.
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http://dx.doi.org/10.1523/JNEUROSCI.0356-19.2020DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7178913PMC
April 2020

The habenular G-protein-coupled receptor 151 regulates synaptic plasticity and nicotine intake.

Proc Natl Acad Sci U S A 2020 03 25;117(10):5502-5509. Epub 2020 Feb 25.

Laboratory of Molecular Biology, The Rockefeller University, New York, NY 10065;

The habenula, an ancient small brain area in the epithalamus, densely expresses nicotinic acetylcholine receptors and is critical for nicotine intake and aversion. As such, identification of strategies to manipulate habenular activity may yield approaches to treat nicotine addiction. Here we show that GPR151, an orphan G-protein-coupled receptor (GPCR) highly enriched in the habenula of humans and rodents, is expressed at presynaptic membranes and synaptic vesicles and associates with synaptic components controlling vesicle release and ion transport. Deletion of inhibits evoked neurotransmission but enhances spontaneous miniature synaptic currents and eliminates short-term plasticity induced by nicotine. We find that GPR151 couples to the G-alpha inhibitory protein Gα to reduce cyclic adenosine monophosphate (cAMP) levels in mice and in GPR151-expressing cell lines that are amenable to ligand screens. knockout (KO) mice show diminished behavioral responses to nicotine and self-administer greater quantities of the drug, phenotypes rescued by viral reexpression of in the habenula. These data identify GPR151 as a critical modulator of habenular function that controls nicotine addiction vulnerability.
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http://dx.doi.org/10.1073/pnas.1916132117DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7071848PMC
March 2020

Habenular TCF7L2 links nicotine addiction to diabetes.

Nature 2019 10 16;574(7778):372-377. Epub 2019 Oct 16.

Nash Family Department of Neuroscience, Icahn School of Medicine at Mount Sinai, New York, NY, USA.

Diabetes is far more prevalent in smokers than non-smokers, but the underlying mechanisms of vulnerability are unknown. Here we show that the diabetes-associated gene Tcf7l2 is densely expressed in the medial habenula (mHb) region of the rodent brain, where it regulates the function of nicotinic acetylcholine receptors. Inhibition of TCF7L2 signalling in the mHb increases nicotine intake in mice and rats. Nicotine increases levels of blood glucose by TCF7L2-dependent stimulation of the mHb. Virus-tracing experiments identify a polysynaptic connection from the mHb to the pancreas, and wild-type rats with a history of nicotine consumption show increased circulating levels of glucagon and insulin, and diabetes-like dysregulation of blood glucose homeostasis. By contrast, mutant Tcf7l2 rats are resistant to these actions of nicotine. Our findings suggest that TCF7L2 regulates the stimulatory actions of nicotine on a habenula-pancreas axis that links the addictive properties of nicotine to its diabetes-promoting actions.
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http://dx.doi.org/10.1038/s41586-019-1653-xDOI Listing
October 2019

Cell-Type-Specific Contributions of Medial Prefrontal Neurons to Flexible Behaviors.

J Neurosci 2018 05 12;38(19):4490-4504. Epub 2018 Apr 12.

Laboratory of Molecular Biology, Rockefeller University, New York, New York 10065

Behavioral flexibility and impulse control are necessary for successful execution of adaptive behavior. They are impaired in patients with damage to the prefrontal cortex (PFC) and in some clinically important conditions, such as obsessive-compulsive disorder. Although the medial prefrontal cortex (mPFC) has been investigated as a critical structure for behavioral flexibility and impulse control, the contribution of the underlying pyramidal neuron cell types in the mPFC remained to be understood. Here we show that interneuron-mediated local inactivation of pyramidal neurons in the mPFC of male and female mice induces both premature responses and choice bias, and establish that these impulsive and compulsive responses are modulated independently. Cell-type-specific photoinhibition of pyramidal deep layer corticostriatal or corticothalamic neurons reduces behavioral flexibility without inducing premature responses. Together, our data confirm the role of corticostriatal neurons in behavioral flexibility and demonstrate that flexible behaviors are also modulated by direct projections from deep layer corticothalamic neurons in the mPFC to midline thalamic nuclei. Behavioral flexibility and impulse control are indispensable for animals to adapt to changes in the environment and often affected in patients with PFC damage and obsessive-compulsive disorder. We used a probabilistic reversal task to dissect the underlying neural circuitry in the mPFC. Through characterization of the three major pyramidal cell types in the mPFC with optogenetic silencing, we demonstrated that corticostriatal and corticothalamic but not corticocortical pyramidal neurons are temporally recruited for behavioral flexibility. Together, our findings confirm the role of corticostriatal projections in cognitive flexibility and identify corticothalamic neurons as equally important for behavioral flexibility.
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http://dx.doi.org/10.1523/JNEUROSCI.3537-17.2018DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5943977PMC
May 2018

Retrograde inhibition by a specific subset of interpeduncular α5 nicotinic neurons regulates nicotine preference.

Proc Natl Acad Sci U S A 2017 12 20;114(49):13012-13017. Epub 2017 Nov 20.

Laboratory of Molecular Biology, The Rockefeller University, New York, NY 10065;

Repeated exposure to drugs of abuse can produce adaptive changes that lead to the establishment of dependence. It has been shown that allelic variation in the α5 nicotinic acetylcholine receptor (nAChR) gene is associated with higher risk of tobacco dependence. In the brain, α5-containing nAChRs are expressed at very high levels in the interpeduncular nucleus (IPN). Here we identified two nonoverlapping α5 cell populations (α5- and α5- ) in mouse IPN that respond differentially to nicotine. Chronic nicotine treatment altered the translational profile of more than 1,000 genes in α5- neurons, including neuronal nitric oxide synthase () and somatostatin (). In contrast, expression of few genes was altered in the α5- population. We show that both nitric oxide and SST suppress optically evoked neurotransmitter release from the terminals of habenular (Hb) neurons in IPN. Moreover, in vivo silencing of neurotransmitter release from the α5- but not from the α5- population eliminates nicotine reward, measured using place preference. This loss of nicotine reward was mimicked by shRNA-mediated knockdown of in the IPN. These findings reveal a proaddiction adaptive response to chronic nicotine in which nitric oxide and SST are released by a specific α5 neuronal population to provide retrograde inhibition of the Hb-IPN circuit and thereby enhance the motivational properties of nicotine.
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http://dx.doi.org/10.1073/pnas.1717506114DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5724287PMC
December 2017

A Cortical Circuit for Sexually Dimorphic Oxytocin-Dependent Anxiety Behaviors.

Cell 2016 Sep 15;167(1):60-72.e11. Epub 2016 Sep 15.

Laboratory of Molecular Biology, Howard Hughes Medical Institute, The Rockefeller University, 1230 York Avenue, New York, NY 10065, USA. Electronic address:

The frequency of human social and emotional disorders varies significantly between males and females. We have recently reported that oxytocin receptor interneurons (OxtrINs) modulate female sociosexual behavior. Here, we show that, in male mice, OxtrINs regulate anxiety-related behaviors. We demonstrate that corticotropin-releasing-hormone-binding protein (CRHBP), an antagonist of the stress hormone CRH, is specifically expressed in OxtrINs. Production of CRHBP blocks the CRH-induced potentiation of postsynaptic layer 2/3 pyramidal cell activity of male, but not female, mice, thus producing an anxiolytic effect. Our data identify OxtrINs as critical for modulation of social and emotional behaviors in both females and males and reveal a molecular mechanism that acts on local medial prefrontal cortex (mPFC) circuits to coordinate responses to OXT and CRH. They suggest that additional studies of the impact of the OXT/OXTR and CRHBP/CRH pathways in males and females will be important in development of gender-specific therapies.
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http://dx.doi.org/10.1016/j.cell.2016.08.067DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5220951PMC
September 2016

An essential role of acetylcholine-glutamate synergy at habenular synapses in nicotine dependence.

Elife 2015 Dec 1;4:e11396. Epub 2015 Dec 1.

Molecular Neurobiology Group, Max Delbrück Center for Molecular Medicine, Berlin, Germany.

A great deal of interest has been focused recently on the habenula and its critical role in aversion, negative-reward and drug dependence. Using a conditional mouse model of the ACh-synthesizing enzyme choline acetyltransferase (Chat), we report that local elimination of acetylcholine (ACh) in medial habenula (MHb) neurons alters glutamate corelease and presynaptic facilitation. Electron microscopy and immuno-isolation analyses revealed colocalization of ACh and glutamate vesicular transporters in synaptic vesicles (SVs) in the central IPN. Glutamate reuptake in SVs prepared from the IPN was increased by ACh, indicating vesicular synergy. Mice lacking CHAT in habenular neurons were insensitive to nicotine-conditioned reward and withdrawal. These data demonstrate that ACh controls the quantal size and release frequency of glutamate at habenular synapses, and suggest that the synergistic functions of ACh and glutamate may be generally important for modulation of cholinergic circuit function and behavior.
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http://dx.doi.org/10.7554/eLife.11396DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4718731PMC
December 2015

Excitatory transmission at thalamo-striatal synapses mediates susceptibility to social stress.

Nat Neurosci 2015 Jul 1;18(7):962-4. Epub 2015 Jun 1.

Fishberg Department of Neuroscience, Friedman Brain Institute Graduate School of Biomedical Sciences, Icahn School of Medicine at Mount Sinai, New York, New York, USA.

Postsynaptic remodeling of glutamatergic synapses on ventral striatum (vSTR) medium spiny neurons (MSNs) is critical for shaping stress responses. However, it is unclear which presynaptic inputs are involved. Susceptible mice exhibited increased synaptic strength at intralaminar thalamus (ILT), but not prefrontal cortex (PFC), inputs to vSTR MSNs following chronic social stress. Modulation of ILT-vSTR versus PFC-vSTR neuronal activity differentially regulated dendritic spine plasticity and social avoidance.
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http://dx.doi.org/10.1038/nn.4034DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4482771PMC
July 2015

Suppression of Peripheral Pain by Blockade of Voltage-Gated Calcium 2.2 Channels in Nociceptors Induces RANKL and Impairs Recovery From Inflammatory Arthritis in a Mouse Model.

Arthritis Rheumatol 2015 Jun;67(6):1657-67

The Rockefeller University, New York, New York.

Objective: A hallmark of rheumatoid arthritis (RA) is the chronic pain that accompanies inflammation and joint deformation. Patients with RA rate pain relief as the highest priority; however, few studies have addressed the efficacy and safety of therapies directed specifically toward pain pathways. The ω-conotoxin MVIIA (ziconotide) is used in humans to alleviate persistent pain syndromes, because it specifically blocks the voltage-gated calcium 2.2 (CaV 2.2) channel, which mediates the release of neurotransmitters and proinflammatory mediators from peripheral nociceptor nerve terminals. The aims of this study were to investigate whether blockade of CaV 2.2 can suppress arthritis pain, and to examine the progression of induced arthritis during persistent CaV 2.2 blockade.

Methods: Transgenic mice expressing a membrane-tethered form of MVIIA under the control of a nociceptor-specific gene (MVIIA-transgenic mice) were used in the experiments. The mice were subjected to unilateral induction of joint inflammation using a combination of antigen and collagen.

Results: CaV 2.2 blockade mediated by tethered MVIIA effectively suppressed arthritis-induced pain; however, in contrast to their wild-type littermates, which ultimately regained use of their injured joint as inflammation subsided, MVIIA-transgenic mice showed continued inflammation, with up-regulation of the osteoclast activator RANKL and concomitant joint and bone destruction.

Conclusion: Taken together, our results indicate that alleviation of peripheral pain by blockade of CaV 2.2- mediated calcium influx and signaling in nociceptor sensory neurons impairs recovery from induced arthritis and point to the potentially devastating effects of using CaV 2.2 channel blockers as analgesics during inflammation.
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http://dx.doi.org/10.1002/art.39094DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4446144PMC
June 2015

The habenulo-interpeduncular pathway in nicotine aversion and withdrawal.

Neuropharmacology 2015 Sep 2;96(Pt B):213-22. Epub 2014 Dec 2.

Laboratory of Molecular Biology and Howard Hughes Medical Institute, The Rockefeller University, 1230 York Avenue, NY 10065, New York, USA. Electronic address:

Progress has been made over the last decade in our understanding of the brain areas and circuits involved in nicotine reward and withdrawal, leading to models of addiction that assign different addictive behaviors to distinct, yet overlapping, neural circuits (Koob and Volkow, 2010; Lobo and Nestler, 2011; Tuesta et al., 2011; Volkow et al., 2011). Recently the habenulo-interpeduncular (Hb-IPN) midbrain pathway has re-emerged as a new critical crossroad that influences the brain response to nicotine. This brain area is particularly enriched in nicotinic acetylcholine receptor (nAChR) subunits α5, α3 and β4 encoded by the CHRNA5-A3-B4 gene cluster, which has been associated with vulnerability to tobacco dependence in human genetics studies. This finding, together with studies in mice involving deletion and replacement of nAChR subunits, and investigations of the circuitry, cell types and electrophysiological properties, have begun to identify the molecular mechanisms that take place in the MHb-IPN which underlie critical aspects of nicotine dependence. In the current review we describe the anatomical and functional connections of the MHb-IPN system, as well as the contribution of specific nAChRs subtypes in nicotine-mediated behaviors. Finally, we discuss the specific electrophysiological properties of MHb-IPN neuronal populations and how nicotine exposure alters their cellular physiology, highlighting the unique role of the MHb-IPN in the context of nicotine aversion and withdrawal. This article is part of the Special Issue entitled 'The Nicotinic Acetylcholine Receptor: From Molecular Biology to Cognition'.
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http://dx.doi.org/10.1016/j.neuropharm.2014.11.019DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4452453PMC
September 2015

Conserved expression of the GPR151 receptor in habenular axonal projections of vertebrates.

J Comp Neurol 2015 Feb 8;523(3):359-80. Epub 2014 Sep 8.

Psychiatric Neuromodulation Unit, Clinical Sciences, Lund University, 222 42, Lund, Sweden.

The habenula is a phylogenetically conserved brain structure in the epithalamus. It is a major node in the information flow between fronto-limbic brain regions and monoaminergic brainstem nuclei, and is thus anatomically and functionally ideally positioned to regulate emotional, motivational, and cognitive behaviors. Consequently, the habenula may be critically important in the pathophysiology of psychiatric disorders such as addiction and depression. Here we investigated the expression pattern of GPR151, a G protein-coupled receptor (GPCR), whose mRNA has been identified as highly and specifically enriched in habenular neurons by in situ hybridization and translating ribosome affinity purification (TRAP). In the present immunohistochemical study we demonstrate a pronounced and highly specific expression of the GPR151 protein in the medial and lateral habenula of rodent brain. Specific expression was also seen in efferent habenular fibers projecting to the interpeduncular nucleus, the rostromedial tegmental area, the rhabdoid nucleus, the mesencephalic raphe nuclei, and the dorsal tegmental nucleus. Using confocal microscopy and quantitative colocalization analysis, we found that GPR151-expressing axons and terminals overlap with cholinergic, substance P-ergic, and glutamatergic markers. Virtually identical expression patterns were observed in rat, mouse, and zebrafish brains. Our data demonstrate that GPR151 is highly conserved, specific for a subdivision of the habenular neurocircuitry, and constitutes a promising novel target for psychiatric drug development.
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http://dx.doi.org/10.1002/cne.23664DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4270839PMC
February 2015

Habenular expression of rare missense variants of the β4 nicotinic receptor subunit alters nicotine consumption.

Front Hum Neurosci 2014 27;8:12. Epub 2014 Jan 27.

Molecular Neurobiology Group, Max Delbrück Center for Molecular Medicine Berlin, Germany ; Laboratory of Molecular Biology, The Rockefeller University New York, NY, USA.

The CHRNA5-CHRNA3-CHRNB4 gene cluster, encoding the α5, α3, and β4 nicotinic acetylcholine receptor (nAChR) subunits, has been linked to nicotine dependence. The habenulo-interpeduncular (Hb-IPN) tract is particularly enriched in α3β4 nAChRs. We recently showed that modulation of these receptors in the medial habenula (MHb) in mice altered nicotine consumption. Given that β4 is rate-limiting for receptor activity and that single nucleotide polymorphisms (SNPs) in CHRNB4 have been linked to altered risk of nicotine dependence in humans, we were interested in determining the contribution of allelic variants of β4 to nicotine receptor activity in the MHb. We screened for missense SNPs that had allele frequencies >0.0005 and introduced the corresponding substitutions in Chrnb4. Fourteen variants were analyzed by co-expression with α3. We found that β4A90I and β4T374I variants, previously shown to associate with reduced risk of smoking, and an additional variant β4D447Y, significantly increased nicotine-evoked current amplitudes, while β4R348C, the mutation most frequently encountered in sporadic amyotrophic lateral sclerosis (sALS), showed reduced nicotine currents. We employed lentiviruses to express β4 or β4 variants in the MHb. Immunoprecipitation studies confirmed that β4 lentiviral-mediated expression leads to specific upregulation of α3β4 but not β2 nAChRs in the Mhb. Mice injected with the β4-containing virus showed pronounced aversion to nicotine as previously observed in transgenic Tabac mice overexpressing Chrnb4 at endogenous sites including the MHb. Habenular expression of the β4 gain-of-function allele T374I also resulted in strong aversion, while transduction with the β4 loss-of function allele R348C failed to induce nicotine aversion. Altogether, these data confirm the critical role of habenular β4 in nicotine consumption, and identify specific SNPs in CHRNB4 that modify nicotine-elicited currents and alter nicotine consumption in mice.
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http://dx.doi.org/10.3389/fnhum.2014.00012DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3902282PMC
January 2014

Reexposure to nicotine during withdrawal increases the pacemaking activity of cholinergic habenular neurons.

Proc Natl Acad Sci U S A 2013 Oct 30;110(42):17077-82. Epub 2013 Sep 30.

Laboratory of Molecular Biology, The Rockefeller University, New York, NY 10065.

The discovery of genetic variants in the cholinergic receptor nicotinic CHRNA5-CHRNA3-CHRNB4 gene cluster associated with heavy smoking and higher relapse risk has led to the identification of the midbrain habenula-interpeduncular axis as a critical relay circuit in the control of nicotine dependence. Although clear roles for α3, β4, and α5 receptors in nicotine aversion and withdrawal have been established, the cellular and molecular mechanisms that participate in signaling nicotine use and contribute to relapse have not been identified. Here, using translating ribosome affinity purification (TRAP) profiling, electrophysiology, and behavior, we demonstrate that cholinergic neurons, but not peptidergic neurons, of the medial habenula (MHb) display spontaneous tonic firing of 2-10 Hz generated by hyperpolarization-activated cyclic nucleotide-gated (HCN) pacemaker channels and that infusion of the HCN pacemaker antagonist ZD7288 in the habenula precipitates somatic and affective signs of withdrawal. Further, we show that a strong, α3β4-dependent increase in firing frequency is observed in these pacemaker neurons upon acute exposure to nicotine. No change in the basal or nicotine-induced firing was observed in cholinergic MHb neurons from mice chronically treated with nicotine. We observe, however, that, during withdrawal, reexposure to nicotine doubles the frequency of pacemaking activity in these neurons. These findings demonstrate that the pacemaking mechanism of cholinergic MHb neurons controls withdrawal, suggesting that the heightened nicotine sensitivity of these neurons during withdrawal may contribute to smoking relapse.
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http://dx.doi.org/10.1073/pnas.1313103110DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3800986PMC
October 2013

The biochemical anatomy of cortical inhibitory synapses.

PLoS One 2012 29;7(6):e39572. Epub 2012 Jun 29.

Howard Hughes Medical Institute, Laboratory of Molecular Biology, The Rockefeller University, New York, New York, United States of America.

Classical electron microscopic studies of the mammalian brain revealed two major classes of synapses, distinguished by the presence of a large postsynaptic density (PSD) exclusively at type 1, excitatory synapses. Biochemical studies of the PSD have established the paradigm of the synapse as a complex signal-processing machine that controls synaptic plasticity. We report here the results of a proteomic analysis of type 2, inhibitory synaptic complexes isolated by affinity purification from the cerebral cortex. We show that these synaptic complexes contain a variety of neurotransmitter receptors, neural cell-scaffolding and adhesion molecules, but that they are entirely lacking in cell signaling proteins. This fundamental distinction between the functions of type 1 and type 2 synapses in the nervous system has far reaching implications for models of synaptic plasticity, rapid adaptations in neural circuits, and homeostatic mechanisms controlling the balance of excitation and inhibition in the mature brain.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0039572PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3387162PMC
November 2012

Cholinergic interneurons in the nucleus accumbens regulate depression-like behavior.

Proc Natl Acad Sci U S A 2012 Jul 25;109(28):11360-5. Epub 2012 Jun 25.

Laboratory of Molecular and Cellular Neuroscience, The Rockefeller University, New York, NY 10065, USA.

A large number of studies have demonstrated that the nucleus accumbens (NAC) is a critical site in the neuronal circuits controlling reward responses, motivation, and mood, but the neuronal cell type(s) underlying these processes are not yet known. Identification of the neuronal cell types that regulate depression-like states will guide us in understanding the biological basis of mood and its regulation by diseases like major depressive disorder. Taking advantage of recent findings demonstrating that the serotonin receptor chaperone, p11, is an important molecular regulator of depression-like states, here we identify cholinergic interneurons (CINs) as a primary site of action for p11 in the NAC. Depression-like behavior is observed in mice after decrease of p11 levels in NAC CINs. This phenotype is recapitulated by silencing neuronal transmission in these cells, demonstrating that accumbal cholinergic neuronal activity regulates depression-like behaviors and suggesting that accumbal CIN activity is crucial for the regulation of mood and motivation.
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http://dx.doi.org/10.1073/pnas.1209293109DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3396525PMC
July 2012

Cholinergic receptors in the murine oviduct: inventory and coupling to intracellular calcium concentration.

Life Sci 2012 Nov 28;91(21-22):1003-8. Epub 2012 Mar 28.

Institute of Anatomy and Cell Biology, Justus-Liebig-University, 35385 Giessen, Germany.

Aims: In the oviduct, muscarinic acetylcholine receptors (MR) are linked with motility regulation and nicotinic receptors (nAChR) with ectopic pregnancy. We here aimed to determine the repertoire of cholinergic receptor expression in the murine oviduct and their functional coupling to regulation of intracellular calcium concentration ([Ca(2+)](i)).

Main Methods: Cholinergic receptor transcripts were assessed by RT-PCR in oviductal segments (ampulla, isthmus, uterotubar junction) in all cyclic stages and pregnancy, and in laser-microdissected samples of epithelium and smooth muscle, nAChR subunit α3 distribution in tissue sections using an appropriate genetic reporter mouse strain. [Ca(2+)](i) responses were monitored in ciliated and non-ciliated oviductal cells isolated from wild-type and MR subtypes 1 and 3 gene deficient mice.

Key Findings: Transcripts for all MR subtypes (M1-M5) are constantly expressed whereas there is some variability in nAChR expression from individual to individual. The qualitative expression pattern is independent from the hormonal status of the animal, except for nAChR α7, which is less present during pregnancy. The epithelium expresses M1, M3, nAChR α7 (data from laser-assisted microdissection) and nAChR α3 (ultrastructural investigation of reporter mice). MR dominate over nAChR in increasing [Ca(2+)](i) with being M3 the major, but not sole subtype driving this effect. The general nAChR inhibitor mecamylamine enhances muscarinic and purinergic responses.

Significance: In conclusion, the murine oviduct is endowed with a multiplicity of muscarinic and nicotinic receptors subtypes that, with respect to regulation of [Ca(2+)](i), are inversely linked to each other. The major, but not sole, cholinergic receptor driving increase in [Ca(2+)](i) is M3.
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http://dx.doi.org/10.1016/j.lfs.2012.03.016DOI Listing
November 2012

Tethering toxins and peptide ligands for modulation of neuronal function.

Curr Opin Neurobiol 2012 Feb 24;22(1):72-8. Epub 2011 Nov 24.

Molecular Neurobiology Group, Max-Delbrück-Centrum, Robert-Rössle-Str. 10, 13125 Berlin, Germany.

Tethering genetically encoded peptide toxins or ligands close to their point of activity at the cell plasma membrane provides a new approach to the study of cell networks and neuronal circuits, as it allows selective targeting of specific cell populations, enhances the working concentration of the ligand or blocker peptide, and permits the engineering of a large variety of t-peptides (e.g., including use of fluorescent markers, viral vectors and point mutation variants). This review describes the development of tethered toxins (t-toxins) and peptides derived from the identification of the cell surface nicotinic acetylcholine receptor (nAChR) modulator lynx1, the existence of related endogenous cell surface modulators of nAChR and AMPA receptors, and the application of the t-toxin and t-neuropeptide technology to the dissection of neuronal circuits in metazoans.
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http://dx.doi.org/10.1016/j.conb.2011.11.003DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3294089PMC
February 2012

Cross-reactivity of acid-sensing ion channel and Na⁺-H⁺ exchanger antagonists with nicotinic acetylcholine receptors.

J Physiol 2011 Nov 12;589(Pt 21):5109-23. Epub 2011 Sep 12.

Molecular Neurobiology Group, Department of Neuroscience, Max-Delbrück-Centrum, Berlin, Germany.

Nicotinic acetylcholine receptors (nAChRs) are widely distributed throughout the mammalian central and peripheral nervous systems, where they contribute to neuronal excitability and synaptic communication. It has been reported that nAChRs are modulated by BK channels and that BK channels, in turn, are inhibited by acid-sensing ion channels (ASICs). Here we investigate the possible functional interaction between these channels in medial habenula (MHb) neurones. We report that selective antagonists of large-conductance calcium-activated potassium channels and ASIC1a channels, paxilline and psalmotoxin 1, respectively, did not induce detectable changes in nicotine-evoked currents. In contrast, the non-selective ASIC and Na(+)-H(+) exchanger (NHE1) antagonists, amiloride and its analogues, suppressed nicotine-evoked responses in MHb neurones of wild-type and ASIC2 null mice, excluding a possible involvement of ASIC2 in the nAChR inhibition by amiloride. Zoniporide, a more selective inhibitor of NHE1, reversibly inhibited α3β4-, α7- and α4-containing (*) nAChRs in Xenopus oocytes and in brain slices, as well as in PS120 cells deficient in NHE1 and virally transduced with nAChRs, suggesting a generalized effect of zoniporide in most neuronal nAChR subtypes. Independently from nAChR antagonism, zoniporide profoundly blocked synaptic transmission onto MHb neurones without affecting glutamatergic and GABA receptors. Taken together, these results indicate that amiloride and zoniporide, which are clinically used to treat hypertension and cardiovascular disease, have an inhibitory effect on neuronal nAChRs when used experimentally at high doses. The possible cross-reactivity of these compounds with nAChRs in vivo will require further investigation.
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http://dx.doi.org/10.1113/jphysiol.2011.213272DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3225668PMC
November 2011

Cholinergic chemosensory cells in the trachea regulate breathing.

Proc Natl Acad Sci U S A 2011 Jun 23;108(23):9478-83. Epub 2011 May 23.

Institute of Anatomy and Cell Biology and Institute for Clinical Immunology and Transfusion Medicine, Justus-Liebig-University, Giessen D-35385, Germany.

In the epithelium of the lower airways, a cell type of unknown function has been termed "brush cell" because of a distinctive ultrastructural feature, an apical tuft of microvilli. Morphologically similar cells in the nose have been identified as solitary chemosensory cells responding to taste stimuli and triggering trigeminal reflexes. Here we show that brush cells of the mouse trachea express the receptors (Tas2R105, Tas2R108), the downstream signaling molecules (α-gustducin, phospholipase C(β2)) of bitter taste transduction, the synthesis and packaging machinery for acetylcholine, and are addressed by vagal sensory nerve fibers carrying nicotinic acetylcholine receptors. Tracheal application of an nAChR agonist caused a reduction in breathing frequency. Similarly, cycloheximide, a Tas2R108 agonist, evoked a drop in respiratory rate, being sensitive to nicotinic receptor blockade and epithelium removal. This identifies brush cells as cholinergic sensors of the chemical composition of the lower airway luminal microenvironment that are directly linked to the regulation of respiration.
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http://dx.doi.org/10.1073/pnas.1019418108DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3111311PMC
June 2011

Aversion to nicotine is regulated by the balanced activity of β4 and α5 nicotinic receptor subunits in the medial habenula.

Neuron 2011 May;70(3):522-35

Molecular Neurobiology Group, Max-Delbrück-Centrum, Berlin, Germany.

Nicotine dependence is linked to single nucleotide polymorphisms in the CHRNB4-CHRNA3-CHRNA5 gene cluster encoding the α3β4α5 nicotinic acetylcholine receptor (nAChR). Here we show that the β4 subunit is rate limiting for receptor activity, and that current increase by β4 is maximally competed by one of the most frequent variants associated with tobacco usage (D398N in α5). We identify a β4-specific residue (S435), mapping to the intracellular vestibule of the α3β4α5 receptor in close proximity to α5 D398N, that is essential for its ability to increase currents. Transgenic mice with targeted overexpression of Chrnb4 to endogenous sites display a strong aversion to nicotine that can be reversed by viral-mediated expression of the α5 D398N variant in the medial habenula (MHb). Thus, this study both provides insights into α3β4α5 receptor-mediated mechanisms contributing to nicotine consumption, and identifies the MHb as a critical element in the circuitry controlling nicotine-dependent phenotypes.
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http://dx.doi.org/10.1016/j.neuron.2011.04.013DOI Listing
May 2011

The molecular and cellular identity of peripheral osmoreceptors.

Neuron 2011 Jan;69(2):332-44

Department of Neuroscience, Max-Delbrück Center for Molecular Medicine, Robert-Rössle-Straße 10, D-13092 Berlin-Buch, Germany.

In mammals, the osmolality of the extracellular fluid (ECF) is highly stable despite radical changes in salt/water intake and excretion. Afferent systems are required to detect hypo- or hyperosmotic shifts in the ECF to trigger homeostatic control of osmolality. In humans, a pressor reflex is triggered by simply drinking water which may be mediated by peripheral osmoreceptors. Here, we identified afferent neurons in the thoracic dorsal root ganglia (DRG) of mice that innervate hepatic blood vessels and detect physiological hypo-osmotic shifts in blood osmolality. Hepatic sensory neurons are equipped with an inward current that faithfully transduces graded changes in osmolality within the physiological range (~15 mOsm). In mice lacking the osmotically activated ion channel, TRPV4, hepatic sensory neurons no longer exhibit osmosensitive inward currents and activation of peripheral osmoreceptors in vivo is abolished. We have thus identified a new population of sensory neurons that transduce ongoing changes in hepatic osmolality.
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http://dx.doi.org/10.1016/j.neuron.2010.12.028DOI Listing
January 2011

"The King is dead": Checkmating ion channels with tethered toxins.

Toxicon 2010 Dec 7;56(8):1293-8. Epub 2010 Oct 7.

Molecular Neurobiology Group, Department of Neuroscience, Max Delbrück Center for Molecular Medicine, 13125 Berlin, Germany.

The quickest possible checkmate in the game of chess requires two moves using a pawn and the queen. Metaphorically speaking, the pawn (a membrane tether) and the queen (a toxin) work together to checkmate an ion channel within a neuronal circuit. This strategy termed "tethered toxin" (t-toxin) is based on the use of genetically encoded peptide toxins that are anchored to the cell-membrane via a glycolipid or transmembrane tether. Because of their mode of action at the cell surface, t-toxins act only on ion channels and receptors of the cell that is expressing the t-toxin, and not on identical receptors present in neighboring cells that do not express the t-toxin. In this mini-review we discuss the design of these genetic tools and their application for cell-specific and temporal manipulation of ion channel-mediated activities in vivo.
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http://dx.doi.org/10.1016/j.toxicon.2010.09.016DOI Listing
December 2010

An in vivo tethered toxin approach for the cell-autonomous inactivation of voltage-gated sodium channel currents in nociceptors.

J Physiol 2010 May 22;588(Pt 10):1695-707. Epub 2010 Mar 22.

Molecular Neurobiology group, Department of Neuroscience, Max-Delbrück Center for Molecular Medicine, Robert-Rössle Strasse 10, 13125 Berlin, Germany.

Understanding information flow in sensory pathways requires cell-selective approaches to manipulate the activity of defined neurones. Primary afferent nociceptors, which detect painful stimuli, are enriched in specific voltage-gated sodium channel (VGSC) subtypes. Toxins derived from venomous animals can be used to dissect the contributions of particular ion currents to cell physiology. Here we have used a transgenic approach to target a membrane-tethered isoform of the conotoxin MrVIa (t-MrVIa) only to nociceptive neurones in mice. T-MrVIa transgenic mice show a 44 +/- 7% reduction of tetrodotoxin-resistant (TTX-R) VGSC current densities. This inhibition is permanent, reversible and does not result in functional upregulation of TTX-sensitive (TTX-S) VGSCs, voltage-gated calcium channels (VGCCs) or transient receptor potential (TRP) channels present in nociceptive neurones. As a consequence of the reduction of TTX-R VGSC currents, t-MrVIa transgenic mice display decreased inflammatory mechanical hypersensitivity, cold pain insensitivity and reduced firing of cutaneous C-fibres sensitive to noxious cold temperatures. These data validate the use of genetically encoded t-toxins as a powerful tool to manipulate VGSCs in specific cell types within the mammalian nervous system. This novel genetic methodology can be used for circuit mapping and has the key advantage that it enables the dissection of the contribution of specific ionic currents to neuronal function and to behaviour.
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http://dx.doi.org/10.1113/jphysiol.2010.187112DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2887988PMC
May 2010

Silencing neurotransmission with membrane-tethered toxins.

Nat Methods 2010 Mar 7;7(3):229-36. Epub 2010 Feb 7.

Molecular Neurobiology group, Max Delbrück Center for Molecular Medicine, Berlin, Germany.

At synaptic terminals, high voltage-activated Ca(v)2.1 and Ca(v)2.2 calcium channels have an essential and joint role in coupling the presynaptic action potential to neurotransmitter release. Here we show that membrane-tethered toxins allowed cell-autonomous blockade of each channel individually or simultaneously in mouse neurons in vivo. We report optimized constitutive, inducible and Cre recombinase-dependent lentiviral vectors encoding fluorescent recombinant toxins, and we also validated the toxin-based strategy in a transgenic mouse model. Toxins delivered by lentiviral vectors selectively inhibited the dopaminergic nigrostriatal pathway, and transgenic mice with targeted expression in nociceptive peripheral neurons displayed long-lasting suppression of chronic pain. Optimized tethered toxins are tools for cell-specific and temporal manipulation of ion channel-mediated activities in vivo, including blockade of neurotransmitter release.
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http://dx.doi.org/10.1038/nmeth.1425DOI Listing
March 2010

Prostate stem cell antigen is an endogenous lynx1-like prototoxin that antagonizes alpha7-containing nicotinic receptors and prevents programmed cell death of parasympathetic neurons.

J Neurosci 2009 Nov;29(47):14847-54

Department of Anatomy, University of Vermont College of Medicine, Burlington, Vermont 05405, USA.

Vertebrate alpha-bungarotoxin-like molecules of the Ly-6 superfamily have been implicated as balancers of activity and survival in the adult nervous system. To determine whether a member of this family could be involved in the development of the avian ciliary ganglion, we identified 6 Gallus genes by their homology in structure to mouse lynx1 and lynx2. One of these genes, an ortholog of prostate stem cell antigen (psca), is barely detectable at embryonic day (E) 8, before neuronal cell loss in the ciliary ganglion, but increases >100-fold as the number of neurons begins to decline between E9 and E14. PSCA is highly expressed in chicken and mouse telencephalon and peripheral ganglia and correlates with expression of alpha7-containing nicotinic acetylcholine receptors (alpha7-nAChRs). Misexpressing PSCA before cell death in the ciliary ganglion blocks alpha7-nAChR activation by nicotine and rescues the choroid subpopulation from dying. Thus, PSCA, a molecule previously identified as a marker of prostate cancer, is a member of the Ly-6 neurotoxin-like family in the nervous system, and is likely to play a role as a modulator of alpha7 signaling-induced cell death during development.
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http://dx.doi.org/10.1523/JNEUROSCI.2271-09.2009DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2848080PMC
November 2009

Manipulating neuronal circuits with endogenous and recombinant cell-surface tethered modulators.

Front Mol Neurosci 2009 30;2:21. Epub 2009 Oct 30.

York College and The Graduate Center, The American Museum of Natural History, The City University of New York New York, NY, USA.

Neuronal circuits depend on the precise regulation of cell-surface receptors and ion channels. An ongoing challenge in neuroscience research is deciphering the functional contribution of specific receptors and ion channels using engineered modulators. A novel strategy, termed "tethered toxins", was recently developed to characterize neuronal circuits using the evolutionary derived selectivity of venom peptide toxins and endogenous peptide ligands, such as lynx1 prototoxins. Herein, the discovery and engineering of cell-surface tethered peptides is reviewed, with particular attention given to their cell-autonomy, modular composition, and genetic targeting in different model organisms. The relative ease with which tethered peptides can be engineered, coupled with the increasing number of neuroactive venom toxins and ligand peptides being discovered, imply a multitude of potentially innovative applications for manipulating neuronal circuits and tissue-specific cell networks, including treatment of disorders caused by malfunction of receptors and ion channels.
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http://dx.doi.org/10.3389/neuro.02.021.2009DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2776481PMC
July 2011

A role for LYNX2 in anxiety-related behavior.

Proc Natl Acad Sci U S A 2009 Mar 25;106(11):4477-82. Epub 2009 Feb 25.

Laboratory of Molecular Biology, Howard Hughes Medical Institute, The Rockefeller University, New York, NY 10065, USA.

Anxiety disorders are the most prevalent mental disorders in developed societies. Although roles for the prefrontal cortex, amygdala, hippocampus and mediodorsal thalamus in anxiety disorders are well documented, molecular mechanisms contributing to the functions of these structures are poorly understood. Here we report that deletion of Lynx2, a mammalian prototoxin gene that is expressed at high levels in anxiety associated brain areas, results in elevated anxiety-like behaviors. We show that LYNX2 can bind to and modulate neuronal nicotinic receptors, and that loss of Lynx2 alters the actions of nicotine on glutamatergic signaling in the prefrontal cortex. Our data identify Lynx2 as an important component of the molecular mechanisms that control anxiety, and suggest that altered glutamatergic signaling in the prefrontal cortex of Lynx2 mutant mice contributes to increased anxiety-related behaviors.
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http://dx.doi.org/10.1073/pnas.0813109106DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2657456PMC
March 2009

The prototoxin lynx1 acts on nicotinic acetylcholine receptors to balance neuronal activity and survival in vivo.

Neuron 2006 Sep;51(5):587-600

The Laboratory of Molecular Biology, Howard Hughes Medical Institute, The Rockefeller University, New York, NY 10021, USA.

Nicotinic acetylcholine receptors (nAChRs) affect a wide array of biological processes, including learning and memory, attention, and addiction. lynx1, the founding member of a family of mammalian prototoxins, modulates nAChR function in vitro by altering agonist sensitivity and desensitization kinetics. Here we demonstrate, through the generation of lynx1 null mutant mice, that lynx1 modulates nAChR signaling in vivo. Its loss decreases the EC(50) for nicotine by approximately 10-fold, decreases receptor desensitization, elevates intracellular calcium levels in response to nicotine, and enhances synaptic efficacy. lynx1 null mutant mice exhibit enhanced performance in specific tests of learning and memory. Consistent with reports that mutations resulting in hyperactivation of nAChRs can lead to neurodegeneration, aging lynx1 null mutant mice exhibit a vacuolating degeneration that is exacerbated by nicotine and ameliorated by null mutations in nAChRs. We conclude that lynx1 functions as an allosteric modulator of nAChR function in vivo, balancing neuronal activity and survival in the CNS.
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http://dx.doi.org/10.1016/j.neuron.2006.07.025DOI Listing
September 2006

Tethering naturally occurring peptide toxins for cell-autonomous modulation of ion channels and receptors in vivo.

Neuron 2004 Aug;43(3):305-11

Howard Hughes Medical Institute, Laboratory of Molecular Biology, The Rockefeller University, New York, NY 10021, USA.

The physiologies of cells depend on electrochemical signals carried by ion channels and receptors. Venomous animals produce an enormous variety of peptide toxins with high affinity for specific ion channels and receptors. The mammalian prototoxin lynx1 shares with alpha-bungarotoxin the ability to bind and modulate nicotinic receptors (nAChRs); however, lynx1 is tethered to the membrane via a GPI anchor. We show here that several classes of neurotoxins, including bungarotoxins and cobratoxins, retain their selective antagonistic properties when tethered to the membrane. Targeted elimination of nAChR function in zebrafish can be achieved with tethered alpha-bungarotoxin, silencing synaptic transmission without perturbing synapse formation. These studies harness the pharmacological properties of peptide toxins for use in genetic experiments. When combined with specific methods of cell and temporal expression, the extension of this approach to hundreds of naturally occurring peptide toxins opens a new landscape for cell-autonomous regulation of cellular physiology in vivo.
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http://dx.doi.org/10.1016/j.neuron.2004.07.015DOI Listing
August 2004

Dysfunction of axonemal dynein heavy chain Mdnah5 inhibits ependymal flow and reveals a novel mechanism for hydrocephalus formation.

Hum Mol Genet 2004 Sep 21;13(18):2133-41. Epub 2004 Jul 21.

Laboratory of Molecular Biology, Howard Hughes Medical Institute, Rockefeller University, New York, NY 10021, USA.

Motility of unicellular organisms occurred early in evolution with the emergence of cilia and flagella. In vertebrates, motile cilia are required for numerous functions such as clearance of the airways and determination of left-right body asymmetry. Ependymal cells lining the brain ventricles also carry motile cilia, but their biological function has remained obscure. Here, we show that ependymal cilia generate a laminar flow of cerebrospinal fluid through the cerebral aqueduct, which we term as 'ependymal flow'. The axonemal dynein heavy chain gene Mdnah5 is specifically expressed in ependymal cells, and is essential for ultrastructural and functional integrity of ependymal cilia. In Mdnah5-mutant mice, lack of ependymal flow causes closure of the aqueduct and subsequent formation of triventricular hydrocephalus during early postnatal brain development. The higher incidence of aqueduct stenosis and hydrocephalus formation in patients with ciliary defects proves the relevance of this novel mechanism in humans.
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http://dx.doi.org/10.1093/hmg/ddh219DOI Listing
September 2004