Toshio Terashima

Ptf1a, a bHLH transcriptional gene, defines GABAergic neuronal fates in cerebellum

The molecular machinery governing glutamatergic-GABAergic neuronal subtype specification is unclear. Here we describe a cerebellar mutant, cerebelless, which lacks the entire cerebellar cortex in adults. The primary defect of the mutant brains was a specific inhibition of GABAergic neuron production from the cerebellar ventricular zone (VZ), resulting in secondary and complete loss of external germinal layer, pontine, and olivary nuclei during development. We identified the responsible gene, Ptf1a, whose expression was lost in the cerebellar VZ but was maintained in the pancreas in cerebelless. Lineage tracing revealed that two types of neural precursors exist in the cerebellar VZ: Ptf1a-expressing and -nonexpressing precursors, which generate GABAergic and glutamatergic neurons, respectively. Introduction of Ptf1a into glutamatergic neuron precursors in the dorsal telencephalon generated GABAergic neurons with representative morphological and migratory features. Our results suggest that Ptf1a is involved in driving neural precursors to differentiate into GABAergic neurons in the cerebellum.

Visualization, direct isolation, and transplantation of midbrain dopaminergic neurons

To visualize and isolate live dopamine (DA)-producing neurons in the embryonic ventral mesencephalon, we generated transgenic mice expressing green fluorescent protein (GFP) under the control of the rat tyrosine hydroxylase gene promoter. In the transgenic mice, GFP expression was observed in the developing DA neurons containing tyrosine hydroxylase. The outgrowth and cue-dependent guidance of GFP-labeled axons was monitored in vitro with brain culture systems. To isolate DA neurons expressing GFP from brain tissue, cells with GFP fluorescence were sorted by fluorescence-activated cell sorting. More than 60% of the sorted GFP+ cells were positive for tyrosine hydroxylase, confirming that the population had been successfully enriched with DA neurons. The sorted GFP+ cells were transplanted into a rat model of Parkinsons disease. Some of these cells survived and innervated the host striatum, resulting in a recovery from Parkinsonian behavioral defects. This strategy for isolating an enriched population of DA neurons should be useful for cellular and molecular studies of these neurons and for clinical applications in the treatment of Parkinsons disease.

Mouse Ror2 receptor tyrosine kinase is required for the heart development and limb formation

A mouse receptor tyrosine kinase (RTK), mRor2, which belongs to the Ror‐family of RTKs consisting of at least two structurally related members, is primarily expressed in the heart and nervous system during mouse development. To elucidate the function of mRor2, we generated mice with a mutated mRor2 locus.

Expression of reelin, the gene responsible for the reeler mutation, in embryonic development and adulthood in the mouse

reelin has recently been isolated as a candidate gene, the mutation of which gives rise to the reeler phenotype in mice. In this study, we analyzed the expression of reelin during embryonic development in the mouse and in adult mouse tissues, by in situ hybridization. reelin transcripts were present on embryonic day (E) 8.5 in the somite, foregut, yolk sac, and unclosed neural plate. reelin was expressed in the brain, spinal cord, liver, and kidney throughout embryonic development, and transiently in many developing organs such as the optic cup, blood vessels, precartilage, stomach, pituitary, vibrissae, tooth germ, and in cells along growing nerve fibers. These observations indicate a role for reelin in development of organs in addition to that in neuronal migration. Furthermore, we demonstrated the existence of reelin mRNA and its cellular distribution in the adult brain, spinal cord, liver, kidney, testis, and ovary, suggesting additional roles for reelin in stabilizing the cytoarchitecture and in remolding in adult organs. However, we detected no obvious phenotype of the reelin‐expressing organs except for the brain in the reeler mouse, indicating the functional redundancy of this gene during the development of these organs. Dev. Dyn. 1997;210:157–172.

Target Selection by Cortical Axons: Alternative Mechanisms to Establish Axonal Connections in the Developing Brain

We have described our studies of the development of projections from layer 5 of the rat neocortex to subcortical targets in the midbrain and hindbrain. The major points are briefly summarized here. 1. Layer-5 neurons extend a primary axon out of cortex and along a spinally directed trajectory, bypassing all of their targets in the midbrain and hindbrain. These targets are later contacted exclusively by collaterals formed by a delayed interstitial branching of the primary axon, not by growth cone bifurcation. 2. Collateral branches only form at stereotypic positions, not randomly along the length of the axon. Thus, specific cues identify branch points, and the length of the primary axon well behind its growth cone responds to these cues. 3. Layer-5 neurons in diverse areas of cortex initially develop the same basic set of collateral branches, although they will permanently retain different subsets of the initial common set. Therefore, branch cues are recognized by layer-5 neurons independent of whether the collateral projection formed is functionally appropriate for the cortical region in which the neuron resides. 4. In vitro and in vivo evidence indicates that one of the major branches, which forms the corticopontine projection, is induced and directed into its target, the basilar pons, by a diffusible, target-derived, tropic signal. Thus, a chemotropic cue promotes recognition of the basilar pontine target by the primary layer-5 axons. 5. In this system, then, target selection is not the responsibility of the growth cone of the primary axon.(ABSTRACT TRUNCATED AT 250 WORDS)

Expression of the receptor tyrosine kinase genes, Ror1 and Ror2, during mouse development.

In mammals, the Ror-family receptor tyrosine kinases consist of two structurally related proteins, Ror1 and Ror2, characterized by the extracellular Frizzled-like cysteine-rich domain and membrane proximal kringle domains. As an attempt to gain insights into their roles in mouse development, expression patterns of Ror1 and Ror2 during early embryogenesis were examined and compared. Interestingly, at early stages, Ror1 and Ror2 exhibit similar expression patterns in the developing face, including the frontonasal process and pharyngeal arches, which are derived from cephalic neural crest cells. On the other hand, they exhibit different expression patterns in the developing limbs and brain, where the expression of Ror2 was detected broadly compared with that of Ror1. At a later stage, both genes are expressed in a similar fashion in the developing heart and lung, yet in a distinct manner in the brain and eye.

Draxin, a repulsive guidance protein for spinal cord and forebrain commissures.

Axon guidance proteins are critical for the correct wiring of the nervous system during development. Several axon guidance cues and their family members have been well characterized. More unidentified axon guidance cues are assumed to participate in the formation of the extremely complex nervous system. We identified a secreted protein, draxin, that shares no homology with known guidance cues. Draxin inhibited or repelled neurite outgrowth from dorsal spinal cord and cortical explants in vitro. Ectopically expressed draxin inhibited growth or caused misrouting of chick spinal cord commissural axons in vivo. draxin knockout mice showed defasciculation of spinal cord commissural axons and absence of all forebrain commissures. Thus, draxin is a previously unknown chemorepulsive axon guidance molecule required for the development of spinal cord and forebrain commissures.

Apolipoprotein E and Reelin ligands modulate tau phosphorylation through an Apolipoprotein E receptor/disabled-1/glycogen synthase kinase-3β cascade

Neurofibrillary tangles comprised of highly phosphorylated tau proteins are a key component of Alzheimers disease pathology. Mice lacking Reelin (Reln), double‐knockouts lacking the VLDL receptor (VLDLR) and ApoE receptor2 (ApoER2), and mice lacking disabled‐1 (Dab1) display increased levels of phosphorylated tau. Because Reln binds to recombinant ApoE receptors, assembly of a Reln/ApoE‐receptor/Dab1 (RAD) complex may initiate a signal transduction cascade that controls tau phosphorylation. Conversely, disruption of this RAD complex may increase tau phosphorylation and lead to neurodegeneration. To substantiate this concept, we mated Reln‐deficient mice to ApoE‐deficient mice and found that in the absence of Reln, tau phosphorylation increased as the amount of ApoE decreased. Paralleling the change in tau phosphorylation levels, we found that GSK‐3β activity increased in Reln‐deficient mice and further increased in mice lacking both Reln and ApoE. CDK‐5 activity was similar in mice lacking Reln, ApoE, or both. GSK‐3β and CDK‐5 activity increased in Dab1‐deficient mice, independent of ApoE levels. Further supporting the idea that increased tau phosphorylation results primarily from increased kinase activity, the activity of two phosphatases was similar in all conditions tested. These data support a novel, ligand‐mediated signal transduction cascade— initiated by the assembly of a RAD complex that suppresses kinase activity and controls tau phosphorylation.

Anatomy, development and lesion-induced plasticity of rodent corticospinal tract

In this review the current knowledge of the anatomy, development and plasticity of the rodent corticospinal tract is summarised. Recent technical advancements, especially in neuronal tracing methods, have provided much new data concerning the anatomy of the corticospinal tract. The rodent corticospinal axons project to the subcortical nuclei via collateral branches. These collateral branches of corticospinal axons are formed by delayed interstitial budding during early postnatal periods. Corticospinal neurons are generated in the ventricular zone during a short time lag, migrate into the cortical plate, and settle in layer V of the cerebral cortex. The migration of corticospinal neurons is experimentally deranged by prenatal exposure to alcohol or genetically affected by the reeler genetic locus (rl), resulting in generation of ectopic corticospinal neurons. Such experimentally or genetically induced ectopic corticospinal neurons are a good model for examining whether target recognition and path finding are affected by the intracortical position of corticospinal neurons. Some chemical molecules (e.g. L1 and B-50/GAP43) are transiently expressed in the corticospinal tract during the perinatal period, while others (e.g. protein kinase C gamma subspecies and alpha CaM kinase II) are permanently expressed in the adult corticospinal tract. The only chemical marker specific for layer V corticofugal neurons is an antibody to a soluble protein, protein 35. Since the corticospinal tract in the rodent is an easily identified group of fibers situated in the most ventral portion of the dorsal funiculus of the spinal cord and exhibits considerable postnatal development, it has often been utilized in the neurological studies on plasticity and regenerative capacity of the lesioned central nervous system. Recently, it has been clarified that growing corticospinal fibers have the ability to penetrate and traverse across the lesion sites under certain special conditions.

Origin of Climbing Fiber Neurons and Their Developmental Dependence on Ptf1a

Climbing fiber (CF) neurons in the inferior olivary nucleus (ION) extend their axons to Purkinje cells, playing a crucial role in regulating cerebellar function. However, little is known about their precise place of birth and developmental molecular machinery. Here, we describe the origin of the CF neuron lineage and the involvement of Ptf1a (pancreatic transcription factor 1a) in CF neuron development. Ptf1a protein was found to be expressed in a discrete dorsolateral region of the embryonic caudal hindbrain neuroepithelium. Because expression of Ptf1a is not overlapping other transcription factors such as Math1 (mouse atonal homolog 1) and Neurogenin1, which are suggested to define domains within caudal hindbrain neuroepithelium (Landsberg et al., 2005), we named the neuroepithelial region the Ptf1a domain. Analysis of mice that express β-galactosidase from the Ptf1a locus revealed that CF neurons are derived from the Ptf1a domain. In contrast, retrograde labeling of precerebellar neurons indicated that mossy fiber neurons are not derived from Ptf1a-expressing progenitors. We could observe a detailed migratory path of CF neurons from the Ptf1a domain to the ION during embryogenesis. In Ptf1a null mutants, putative immature CF neurons produced from this domain were unable to migrate or differentiate appropriately, resulting in a failure of ION formation. Apoptotic cells were observed in the mutant hindbrain. Furthermore, the fate of some cells in the Ptf1a lineage were changed to mossy fiber neurons in Ptf1a null mutants. These findings clarify the precise origin of CF neurons and suggest that Ptf1a controls their fate, survival, differentiation, and migration during development.