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The mammalian brain is a very complex organ containing an estimated 200 billion cells in humans. Therefore, studying human brain development has become very challenging given all the data that are available from different approaches, notably genetic studies. Recent pluripotent stem cell methods have given rise to the possibility of modeling neurodevelopmental diseases associated with genetic defects. Fibroblasts from patients have been reprogrammed into pluripotent stem cells to derive appropriate neuronal lineages. They specifically include different subtypes of cortical neurons that are at the core of human-specific cognitive abilities. The use of neurons derived from induced pluripotent stem cells (iPSC) has led to deciphering convergent and pleiotropic neuronal synaptic phenotypes found in neurodevelopmental disorders such as autism spectrum disorders (ASD) and their associated syndromes. In addition to these initial studies, remarkable progress has been made in the field of stem cells, with the major objective of reproducing the in vivo maturation steps of human neurons. Recently, several studies have demonstrated the ability of human progenitors to respond to guidance cues and signals in vivo that can direct neurons to their appropriate sites of differentiation where they become fully mature neurons. We provide a brief overview on research using human iPSC in ASD and associated syndromes and on the current understanding of new theories using the re-implantation of neural precursors in mouse brain.

Using human induced pluripotent stem cells (iPSC), recent studies have shown that the events underlying autism spectrum disorders (ASD) can occur during neonatal development. We previously analyzed the iPSC-derived pyramidal cortical neurons of a subset of patients with ASD carrying de novo heterozygous mutations in postsynaptic SHANK3 protein, in culture. We reported altered spinogenesis of those neurons. The transplantation of human iPSC-derived neuronal precursors into mouse brain represents a novel option for in vivo analysis of mutations affecting the human brain. In this study, we transplanted the neuronal precursor cells (NPC) into the cortex of newborn mice to analyze their integration and maturation at early stages of development and studied axonal projections of transplanted human neurons into adult mouse brain. We then co-transplanted NPC from a control individual and from a patient carrying a de novo heterozygous SHANK3 mutation. We observed a reduction in cell soma size of selective neuronal categories and in axonal projections at 30 days post-transplantation. In contrast to previous in vitro studies, we did not observe any alteration in spinogenesis at this early age. The humanized chimeric mouse models offer the means to analyze ASD-associated mutations further and provide the opportunity to visualize phenotypes in vivo.

The transplantation of neural progenitors into a host brain represents a useful tool to evaluate the involvement of cell-autonomous processes and host local cues in the regulation of neuronal differentiation during the development of the mammalian brain. Human brain development starts at the embryonic stages, in utero, with unique properties at its neotenic stages. We analyzed the engraftment and differentiation of human neuronal progenitor cells (hNPCs) transplanted in utero into the mouse brain. The influence of the environment was studied by transplanting human NPCs within the lateral ventricles (LV), compared with the prefrontal cortex (PFC) of immunocompetent mice. We developed a semi-automated method to accurately quantify the number of cell bodies and the distribution of neuronal projections among the different mouse brain structures, at 1 and 3 months post-transplantation (MPT). Our data show that human NPCs can differentiate between immature “juvenile” neurons and more mature pyramidal cells in a reproducible manner. Depending on the injection site, LV vs. PFC, specific fetal local environments could modify the synaptogenesis processes while maintaining human neoteny. The use of immunocompetent mice as host species allows us to investigate further neuropathological conditions making use of all of the engineered mouse models already available.

Loss or dysfunction of the cystic fibrosis (CF) transmembrane conductance regulator (CFTR) leads to impairment of airway mucus transport and to chronic lung diseases resulting in progressive respiratory failure. Nicotinic acetylcholine receptors (nAChRs) bind nicotine and nicotine-derived nitrosamines and thus mediate many of the tobacco-related deleterious effects in the lung. Here we identify α7 nAChR as a key regulator of CFTR in the airways. The airway epithelium in α7 knockout mice is characterized by a higher transepithelial potential difference, an increase of amiloride-sensitive apical Na ⁺ absorption, a defective cAMP-dependent Cl ⁻ conductance, higher concentrations of Na ⁺, Cl ⁻, K ⁺, and Ca ²⁺ in secretions, and a decreased mucus transport, all relevant to a deficient CFTR activity. Moreover, prolonged nicotine exposure mimics the absence of α7 nAChR in mice or its inactivation in vitro in human airway epithelial cell cultures. The functional coupling of α7 nAChR to CFTR occurs through Ca ²⁺ entry and activation of adenylyl cyclases, protein kinase A, and PKC. α7 nAChR, CFTR, and adenylyl cyclase-1 are physically and functionally associated in a macromolecular complex within lipid rafts at the apical membrane of surface and glandular airway epithelium. This study establishes the potential role of α7 nAChR in the regulation of CFTR function and in the pathogenesis of smoking-related chronic lung diseases.

Chronic nicotine exposure results in long-term homeostatic regulation of nicotinic acetylcholine receptors (nAChRs) that play a key role in the adaptative cellular processes leading to addiction. However, the relative contribution of the different nAChR subunits in this process is unclear. Using genetically modified mice and pharmacological manipulations, we provide behavioral, electrophysiological, and pharmacological evidence for a long-term mechanism by which chronic nicotine triggers opposing processes differentially mediated by β2*- vs. α7*nAChRs. These data offer previously undescribed insights into the understanding of nicotine addiction and the treatment of several human pathologies by nicotine-like agents chronically acting on β2*- or α7*nAChRs.

The two sides of nicotine The bad news, nicotine addiction has caused the deaths of millions from smoking-related diseases. The good news, nicotine can enhance cognitive performance. In an attempt to disentangle the brain networks that mediate nicotine reward and relevant cognitive functions, Maskos et al . have developed an injectable lentiviral vector that delivers functional nicotinic acetylcholine receptors to defined regions of the mouse brain. The technique was used to generate mice that express nicotinic receptors exclusively in the midbrain ventral tegmental area, the VTA, which contains dopamine reward neurons and is associated with the response to drugs of abuse. The experiment showed that these receptors in the VTA are sufficient for all behavioural and physiological phenomena associated with nicotine dependence. And they are also involved in the higher brain or ‘cognitive’ functions in the mouse. Nicotine's ‘good’ and ‘bad’ sides are, it seems, intimately entangled and originate from a phylogenetically ancient part of the brain. Worldwide, 100 million people are expected to die this century from the consequences of nicotine addiction 1 , but nicotine is also known to enhance cognitive performance 2 . Identifying the molecular mechanisms involved in nicotine reinforcement and cognition is a priority and requires the development of new in vivo experimental paradigms. The ventral tegmental area (VTA) of the midbrain is thought to mediate the reinforcement properties of many drugs of abuse. Here we specifically re-expressed the β2-subunit of the nicotinic acetylcholine receptor (nAChR) by stereotaxically injecting a lentiviral vector into the VTA of mice carrying β2-subunit deletions 3 , 4 . We demonstrate the efficient re-expression of electrophysiologically responsive, ligand-binding nicotinic acetylcholine receptors in dopamine-containing neurons of the VTA, together with the recovery of nicotine-elicited dopamine release and nicotine self-administration. We also quantified exploratory behaviours of the mice, and showed that β2-subunit re-expression restored slow exploratory behaviour (a measure of cognitive function) to wild-type levels, but did not affect fast navigation behaviour. We thus demonstrate the sufficient role of the VTA in both nicotine reinforcement and endogenous cholinergic regulation of cognitive functions.

5-HT-moduline is an endogenous tetrapeptide [Leu-Ser-Ala-Leu (LSAL)] that was first isolated from bovine brain tissue. To understand the physiological role of this tetrapeptide, we studied the localization of 5-HT-moduline binding sites in rat and mouse brains. Quantitative data obtained with a gaseous detector of β-particles (β-imager) indicated that [ 3 H]-5-HT-moduline bound specifically to rat brain sections with high affinity ( K d = 0.77 nM and B max = 0.26 dpm/mm 2 ). Using film autoradiography in parallel, we found that 5-HT-moduline binding sites were expressed in a variety of rat and mouse brain structures. In 5-HT 1B receptor knock-out mice, the specific binding of [ 3 H]-5-HT-moduline was not different from background labeling, indicating that 5-HT-moduline targets are exclusively located on the 5-HT 1B receptors. Although the distribution of 5-HT-moduline binding sites was similar to that of 5-HT 1B receptors, they did not overlap totally. Differences in distribution patterns were found in regions containing either high levels of 5-HT 1B receptors such as globus pallidus and subiculum that were poorly labeled or in other regions such as dentate gyrus of hippocampus and cortex where the relative density of 5-HT-moduline binding sites was higher than that of 5-HT 1B receptors. In conclusion, our data, based on autoradiographic localization, indicate that 5-HT-moduline targets are located on 5-HT 1B receptors present both on 5-HT afferents and postsynaptic neurons. By interacting specifically with 5-HT 1B receptors, this tetrapeptide may play a pivotal role in pathological states such as stress that involves the dysfunction of 5-HT neurotransmission. 5-HT-moduline β-imager serotonergic system 5-HT1B receptor 5-HT1B receptor knock-out mice

Rationale Nicotine is the main addictive component of tobacco and modifies brain function via its action on neuronal acetylcholine nicotinic receptors (nAChRs). The mesolimbic dopamine (DA) system, where neurons of the ventral tegmental area (VTA) project to the nucleus accumbens (ACb), is considered a core site for the processing of nicotine’s reinforcing properties. However, the precise subtypes of nAChRs that mediate the rewarding properties of nicotine and that contribute to the development of addiction remain to be identified. Objectives We investigated the role of the nAChRs containing the α7 nicotinic subunit (α7*nAChRs) in the reinforcing properties of nicotine within the VTA and in the nicotine-induced changes in ACb DA outflow in vivo. Methods We performed intra-VTA self-administration and microdialysis experiments in genetically modified mice lacking the α7 nicotinic subunit or after pharmacological blockade of α7*nAChRs in wild-type mice. Results We show that the reinforcing properties of nicotine within the VTA are lower in the absence or after pharmacological blockade of α7*nAChRs. We also report that nicotine-induced increases in ACb DA extracellular levels last longer in the absence of these receptors, suggesting that α7*nAChRs regulate the action of nicotine on DA levels over time. Conclusions The present results reveal new insights for the role of α7*nAChRs in modulating the action of nicotine within the mesolimbic circuit. These receptors appear to potentiate the reinforcing action of nicotine administered into the VTA while regulating its action over time on DA outflow in the ACb.

Loss or dysfunction of the cystic fibrosis (CF) transmembrane conductance regulator (CFTR) leads to impairment of airway mucus transport and to chronic lung diseases resulting in progressive respiratory failure. Nicotinic acetylcholine receptors (nAChRs) bind nicotine and nicotine-derived nitrosamines and thus mediate many of the tobacco-related deleterious effects in the lung. Here we identify α7 nAChR as a key regulator of CFTR in the airways. The airway epithelium in α7 knockout mice is characterized by a higher transepithelial potential difference, an increase of amiloride-sensitive apical Na+ absorption, a defective cAMP-dependent Cl- conductance, higher concentrations of Na+, Cl-, K+, and ... in secretions, and a decreased mucus transport, all relevant to a deficient CFTR activity. Moreover, prolonged nicotine exposure mimics the absence of α7 nAChR in mice or its inactivation in vitro in human airway epithelial cell cultures. The functional coupling of α7 nAChR to CFTR occurs through ... entry and activation of adenylyl cyclases, protein kinase A, and PKC. α7 nAChR, CFTR, and adenylyl cyclase-1 are physically and functionally associated in a macromolecular complex within lipid rafts at the apical membrane of surface and glandular airway epithelium. This study establishes the potential role of α7 nAChR in the regulation of CFTR function and in the pathogenesis of smoking-related chronic lung diseases. (ProQuest: ... denotes formulae/symbols omitted.)

Nicotinic acetylcholine receptors (nAChRs) in the brain exhibit diverse functional properties and ubiquitous distribution. Yet, except for providing a receptor for the exogenously applied nicotine of tobacco products, their role in the normal functioning of the brain has remained elusive. We have used a lentiviral expression vector to re-express the beta2 subunit specifically in the ventral tegmental area (VTA) of beta2-/- mice. The viral vector efficiently expresses beta2- subunit protein leading to new nAChR-binding sites. VTA neurons transduced by the lentiviral vector are responsive to intravenous nicotine when analyzed using in vivo electrophysiology. Nicotine-induced dopamine release from the nucleus accumbens (NuAcc) was also restored in re-expressing beta2-/- mice. Intra-VTA injection of nicotine was found to be reinforcing in both wild-type and beta2-subunit re-expressing beta2-/- mice, but not in beta2-/- mice. Furthermore, in the absence of applied nicotine, the spontaneous slow exploratory behavior of the mice was restored, whereas fast navigation did not change. This latter behavioral analysis suggests a role for beta2* nAChR, specifically expressed in the VTA, in mammalian cognitive function.