Zoltán Molnár

Genetic ablation of the t-SNARE SNAP-25 distinguishes mechanisms of neuroexocytosis.

Axon outgrowth during development and neurotransmitter release depends on exocytotic mechanisms, although what protein machinery is common to or differentiates these processes remains unclear. Here we show that the neural t-SNARE (target-membrane-associated–soluble N-ethylmaleimide fusion protein attachment protein (SNAP) receptor) SNAP-25 is not required for nerve growth or stimulus-independent neurotransmitter release, but is essential for evoked synaptic transmission at neuromuscular junctions and central synapses. These results demonstrate that the development of neurotransmission requires the recruitment of a specialized SNARE core complex to meet the demands of regulated exocytosis.

Satb2 Is a Postmitotic Determinant for Upper-Layer Neuron Specification in the Neocortex

Pyramidal neurons of the neocortex can be subdivided into two major groups: deep- (DL) and upper-layer (UL) neurons. Here we report that the expression of the AT-rich DNA-binding protein Satb2 defines two subclasses of UL neurons: UL1 (Satb2 positive) and UL2 (Satb2 negative). In the absence of Satb2, UL1 neurons lose their identity and activate DL- and UL2-specific genetic programs. UL1 neurons in Satb2 mutants fail to migrate to superficial layers and do not contribute to the corpus callosum but to the corticospinal tract, which is normally populated by DL axons. Ctip2, a gene required for the formation of the corticospinal tract, is ectopically expressed in all UL1 neurons in the absence of Satb2. Satb2 protein interacts with the Ctip2 genomic region and controls chromatin remodeling at this locus. Satb2 therefore is required for the initiation of the UL1-specific genetic program and for the inactivation of DL- and UL2-specific genes.

Thalamocortical development: how are we going to get there?

The arealization of the mammalian cortex is believed to be controlled by a combination of intrinsic factors that are expressed in the cortex, and external signals, some of which are mediated through thalamic input. Recent studies on transgenic mice have identified families of molecules that are involved in thalamic axon growth, pathfinding and cortical target selection, and we are beginning to understand how thalamic projections impose cytoarchitectonic differentiation on the developing cortex. By unravelling these mechanisms further, we should be able to increase our understanding of the principles of cortical organization.

Engaging neuroscience to advance translational research in brain barrier biology

The delivery of many potentially therapeutic and diagnostic compounds to specific areas of the brain is restricted by brain barriers, of which the most well known are the blood–brain barrier (BBB) and the blood–cerebrospinal fluid (CSF) barrier. Recent studies have shown numerous additional roles of these barriers, including an involvement in neurodevelopment, in the control of cerebral blood flow, and — when barrier integrity is impaired — in the pathology of many common CNS disorders such as Alzheimers disease, Parkinsons disease and stroke.

Mutations in α-Tubulin Cause Abnormal Neuronal Migration in Mice and Lissencephaly in Humans

Summary The development of the mammalian brain is dependent on extensive neuronal migration. Mutations in mice and humans that affect neuronal migration result in abnormal lamination of brain structures with associated behavioral deficits. Here, we report the identification of a hyperactive N-ethyl-N-nitrosourea (ENU)-induced mouse mutant with abnormalities in the laminar architecture of the hippocampus and cortex, accompanied by impaired neuronal migration. We show that the causative mutation lies in the guanosine triphosphate (GTP) binding pocket of α-1 tubulin (Tuba1) and affects tubulin heterodimer formation. Phenotypic similarity with existing mouse models of lissencephaly led us to screen a cohort of patients with developmental brain anomalies. We identified two patients with de novo mutations in TUBA3, the human homolog of Tuba1. This study demonstrates the utility of ENU mutagenesis in the mouse as a means to discover the basis of human neurodevelopmental disorders.

Towards the classification of subpopulations of layer V pyramidal projection neurons

The nature of cerebral cortical circuitry has been increasingly clarified by markers for the identification of precise cell types with specific morphology, connectivity and distinct physiological properties. Molecular markers are not only helpful in dissecting cortical circuitry, but also give insight into the mechanisms of cortical neuronal specification and differentiation. The two principal neuronal types of the cerebral cortex are the pyramidal and GABAergic cells. Pyramidal cells are excitatory and project to distant targets, while GABAergic neurons are mostly inhibitory non-pyramidal interneurons. Reliable markers for specific subtypes of interneurons are available and have been employed in the classification and functional analysis of cortical circuitry. Until recently, cortical pyramidal neurons have been considered a homogeneous class of cells. This concept is now changing as the powerful tools of molecular biology and genetics identify molecular tags for subtypes of pyramidal cells such as: Otx-1 [Frantz, G.D., Bohner, A.P., Akers, R.M., McConnell, S.K., 1994. Regulation of the POU domain gene SCIP during cerebral cortical development. J. Neurosci. 14, 472-485; Weimann, J.M., Zhang, Y.A., Levin, M.E., Devine, W.P., Brulet, P., McConnell, S.K., 1999. Cortical neurons require Otx1 for the refinement of exuberant axonal projections to subcortical targets. Neuron 24, 819-831]; SMI-32, N200 and FNP-7 [Voelker, C.C., Garin, N., Taylor, J.S., Gahwiler, B.H., Hornung, J.P., Molnár, Z., 2004. Selective neurofilament (SMI-32, FNP-7 and N200) expression in subpopulations of layer V pyramidal neurons in vivo and in vitro. Cereb. Cortex 14, 1276-1286]; ER81 [Hevner, R.F., Daza, R.A., Rubenstein, J.L., Stunnenberg, H., Olavarria, J.F., Englund, C., 2003. Beyond laminar fate: toward a molecular classification of cortical projection/pyramidal neurons. Dev. Neurosci. 25 (2-4), 139-151; Yoneshima, H., Yamasaki, S., Voelker, C., Molnár, Z., Christophe, E., Audinat, E., Takemoto, M., Tsuji, S., Fujita, I., Yamamoto, N., 2006. ER81 is expressed in a subpopulation of layer 5 projection neurons in rodent cerebral cortices. Neuroscience, 137, 401-412]; Lmo4 [Bulchand, S., Subramanian, L., Tole, S., 2003. Dynamic spatiotemporal expression of LIM genes and cofactors in the embryonic and postnatal cerebral cortex. Dev. Dyn. 226, 460-469; Arlotta, P., Molyneaux, B.J., Chen, J., Inoue, J., Kominami, R., Macklis, J.D., 2005. Neuronal subtype-specific genes that control corticospinal motor neuron development in vivo. Neuron 45 (2), 207-221]; CTIP2 [Arlotta, P., Molyneaux, B.J., Chen, J., Inoue, J., Kominami, R., Macklis, J.D., 2005. Neuronal subtype-specific genes that control corticospinal motor neuron development in vivo. Neuron 45 (2), 207-221]; Fez1 [Molyneaux, B.J., Arlotta, P., Hirata, T., Hibi, M., Macklis, J.D., 2005. Fez1 is required for the birth and specification of corticospinal motor neurons. Neuron 47 (6), 817-831; Chen, B., Schaevitz, L.R., McConnell, S.K., 2005. Fez1 regulates the differentiation and axon targeting of layer 5 subcortical projection neurons in cerebral cortex. Proc. Natl. Acad. Sci. U.S.A. 102 (47), 17184-17189]. These genes outline the numerous subtypes of pyramidal cells and are increasingly refining our previous classifications. They also indicate specific developmental programs operate in cell fate decisions. This review will describe the progress made on the correlation of these markers to each other within a specific subtype of layer V neurons with identified, stereotypic projections. Further work is needed to link these data with observations on somatodendritic morphology and physiological properties. The integrated molecular, anatomical and physiological characterisation of pyramidal neurons will lead to a much better appreciation of functional cortical circuits.

The T-box transcription factor Eomes/Tbr2 regulates neurogenesis in the cortical subventricular zone

The embryonic subventricular zone (SVZ) is a critical site for generating cortical projection neurons; however, molecular mechanisms regulating neurogenesis specifically in the SVZ are largely unknown. The transcription factor Eomes/Tbr2 is transiently expressed in cortical SVZ progenitor cells. Here we demonstrate that conditional inactivation of Tbr2 during early brain development causes microcephaly and severe behavioral deficits. In Tbr2 mutants the number of SVZ progenitor cells is reduced and the differentiation of upper cortical layer neurons is disturbed. Neurogenesis in the adult dentate gyrus but not the subependymal zone is abolished. These studies establish Tbr2 as a key regulator of neurogenesis in the SVZ.

Comparative aspects of cerebral cortical development.

This review aims to provide examples of how both comparative and genetic analyses contribute to our understanding of the rules for cortical development and evolution. Genetic studies have helped us to realize the evolutionary rules of telencephalic organization in vertebrates. The control of the establishment of conserved telencephalic subdivisions and the formation of boundaries between these subdivisions has been examined and the very specific alterations at the striatocortical junction have been revealed. Comparative studies and genetic analyses both demonstrate the differential origin and migratory pattern of the two basic neuron types of the cerebral cortex. GABAergic interneurons are mostly generated in the subpallium and a common mechanism governs their migration to the dorsal cortex in both mammals and sauropsids. The pyramidal neurons are generated within the cortical germinal zone and migrate radially, the earliest generated cell layers comprising preplate cells. Reelin‐positive Cajal–Retzius cells are a general feature of all vertebrates studied so far; however, there is a considerable amplification of the Reelin signalling with cortical complexity, which might have contributed to the establishment of the basic mammalian pattern of cortical development. Based on numerous recent observations we shall present the argument that specialization of the mitotic compartments may constitute a major drive behind the evolution of the mammalian cortex. Comparative developmental studies have revealed distinct features in the early compartments of the developing macaque brain, drawing our attention to the limitations of some of the current model systems for understanding human developmental abnormalities of the cortex. Comparative and genetic aspects of cortical development both reveal the workings of evolution.

A Transcriptomic Atlas of Mouse Neocortical Layers

Summary In the mammalian cortex, neurons and glia form a patterned structure across six layers whose complex cytoarchitectonic arrangement is likely to contribute to cognition. We sequenced transcriptomes from layers 1-6b of different areas (primary and secondary) of the adult (postnatal day 56) mouse somatosensory cortex to understand the transcriptional levels and functional repertoires of coding and noncoding loci for cells constituting these layers. A total of 5,835 protein-coding genes and 66 noncoding RNA loci are differentially expressed (“patterned”) across the layers, on the basis of a machine-learning model (naive Bayes) approach. Layers 2-6b are each associated with specific functional and disease annotations that provide insights into their biological roles. This new resource (http://genserv.anat.ox.ac.uk/layers) greatly extends currently available resources, such as the Allen Mouse Brain Atlas and microarray data sets, by providing quantitative expression levels, by being genome-wide, by including novel loci, and by identifying candidate alternatively spliced transcripts that are differentially expressed across layers.

Long noncoding RNA genes: conservation of sequence and brain expression among diverse amniotes

BackgroundLong considered to be the building block of life, it is now apparent that protein is only one of many functional products generated by the eukaryotic genome. Indeed, more of the human genome is transcribed into noncoding sequence than into protein-coding sequence. Nevertheless, whilst we have developed a deep understanding of the relationships between evolutionary constraint and function for protein-coding sequence, little is known about these relationships for non-coding transcribed sequence. This dearth of information is partially attributable to a lack of established non-protein-coding RNA (ncRNA) orthologs among birds and mammals within sequence and expression databases.ResultsHere, we performed a multi-disciplinary study of four highly conserved and brain-expressed transcripts selected from a list of mouse long intergenic noncoding RNA (lncRNA) loci that generally show pronounced evolutionary constraint within their putative promoter regions and across exon-intron boundaries. We identify some of the first lncRNA orthologs present in birds (chicken), marsupial (opossum), and eutherian mammals (mouse), and investigate whether they exhibit conservation of brain expression. In contrast to conventional protein-coding genes, the sequences, transcriptional start sites, exon structures, and lengths for these non-coding genes are all highly variable.ConclusionsThe biological relevance of lncRNAs would be highly questionable if they were limited to closely related phyla. Instead, their preservation across diverse amniotes, their apparent conservation in exon structure, and similarities in their pattern of brain expression during embryonic and early postnatal stages together indicate that these are functional RNA molecules, of which some have roles in vertebrate brain development.