Regulatory cocktail for dopaminergic neurons in a protovertebrate identified by whole-embryo single-cell transcriptomics.

The CNS of the protovertebrate Ciona intestinalis contains a single cluster of dopaminergic (DA) neurons, the coronet cells, which have been likened to the hypothalamus of vertebrates. Whole-embryo single-cell RNA sequencing (RNA-seq) assays identified Ptf1a as the most strongly expressed cell-specific transcription factor (TF) in DA/coronet cells. Knockdown of Ptf1a activity results in their loss, while misexpression results in the appearance of supernumerary DA/coronet cells. Photoreceptor cells and ependymal cells are the most susceptible to transformation, and both cell types express high levels of Meis Coexpression of both Ptf1a and Meis caused the wholesale transformation of the entire CNS into DA/coronet cells. We therefore suggest that the reiterative use of functional manipulations and single-cell RNA-seq assays is an effective means for the identification of regulatory cocktails underlying the specification of specific cell identities.

Neuronal identities derived by misexpression of the POU IV sensory determinant in a protovertebrate.

The protovertebrate Ciona intestinalis type A (sometimes called Ciona robusta) contains a series of sensory cell types distributed across the head-tail axis of swimming tadpoles. They arise from lateral regions of the neural plate that exhibit properties of vertebrate placodes and neural crest. The sensory determinant POU IV/Brn3 is known to work in concert with regional determinants, such as Foxg and Neurogenin, to produce palp sensory cells (PSCs) and bipolar tail neurons (BTNs), in head and tail regions, respectively. A combination of single-cell RNA-sequencing (scRNA-seq) assays, computational analysis, and experimental manipulations suggests that misexpression of POU IV results in variable transformations of epidermal cells into hybrid sensory cell types, including those exhibiting properties of both PSCs and BTNs. Hybrid properties are due to coexpression of Foxg and Neurogenin that is triggered by an unexpected POU IV feedback loop. Hybrid cells were also found to express a synthetic gene battery that is not coexpressed in any known cell type. We discuss these results with respect to the opportunities and challenges of reprogramming cell types through the targeted misexpression of cellular determinants.

Shared evolutionary origin of vertebrate neural crest and cranial placodes.

Placodes and neural crests represent defining features of vertebrates, yet their relationship remains unclear despite extensive investigation1-3. Here we use a combination of lineage tracing, gene disruption and single-cell RNA-sequencing assays to explore the properties of the lateral plate ectoderm of the proto-vertebrate, Ciona intestinalis. There are notable parallels between the patterning of the lateral plate in Ciona and the compartmentalization of the neural plate ectoderm in vertebrates4. Both systems exhibit sequential patterns of Six1/2, Pax3/7 and Msxb expression that depend on a network of interlocking regulatory interactions4. In Ciona, this compartmentalization network produces distinct but related types of sensory cells that share similarities with derivatives of both cranial placodes and the neural crest in vertebrates. Simple genetic disruptions result in the conversion of one sensory cell type into another. We focused on bipolar tail neurons, because they arise from the tail regions of the lateral plate and possess properties of the dorsal root ganglia, a derivative of the neural crest in vertebrates5. Notably, bipolar tail neurons were readily transformed into palp sensory cells, a proto-placodal sensory cell type that arises from the anterior-most regions of the lateral plate in the Ciona tadpole6. Proof of transformation was confirmed by whole-embryo single-cell RNA-sequencing assays. These findings suggest that compartmentalization of the lateral plate ectoderm preceded the advent of vertebrates, and served as a common source for the evolution of both cranial placodes and neural crest3,4.

Home High-Flow Nasal Cannula Oxygen Therapy for Stable Hypercapnic COPD: A Randomized Clinical Trial.

Rationale: The long-term effects of using a high-flow nasal cannula for chronic hypercapnic respiratory failure caused by chronic obstructive pulmonary disease remain unclear. Objectives: To assess whether long-term high-flow nasal cannula use reduces the number of exacerbations and improves other physiological parameters in patients with chronic hypercapnic respiratory failure caused by chronic obstructive pulmonary disease. Methods: We enrolled 104 participants (aged ⩾40 yr) with daytime hypercapnia (Global Initiative for Chronic Obstructive Lung Disease stages 2-4) receiving long-term oxygen therapy (⩾16 h/d for ⩾1 mo) and randomly assigned them to high-flow nasal cannula/long-term oxygen therapy and long-term oxygen therapy groups. The primary endpoint was the moderate or severe exacerbation rate. We compared changes from baseline in arterial blood gas values, peripheral oxygen saturation, pulmonary function, health-related quality-of-life scores, and the 6-minute-walk test. Measurements and Main Results: High-flow nasal cannula use significantly reduced the rate of moderate/severe exacerbations (unadjusted mean count 1.0 vs. 2.5, a ratio of the adjusted mean count between groups [95% confidence interval] of 2.85 [1.48-5.47]) and prolonged the duration without moderate or severe exacerbations. The median time to first moderate or severe exacerbation in the long-term oxygen therapy group was 25 (14.1-47.4) weeks; this was not reached in the high-flow nasal cannula/long-term oxygen therapy group. High-flow nasal cannula use significantly improved health-related quality of life scores, peripheral oxygen saturation, and specific pulmonary function parameters. No safety concerns were identified. Conclusions: A high-flow nasal cannula is a reasonable therapeutic option for patients with stable hypercapnic chronic obstructive pulmonary disease and a history of exacerbations. Clinical trial registered with www.umin/ac.jp (UMIN000028581) and www.clinicaltrials.gov (NCT03282019).

The TRP channel PKD2 is involved in sensing the mechanical stimulus of adhesion for initiating metamorphosis in the chordate Ciona.

Metamorphosis is the dramatic and irreversible reconstruction of animal bodies transitioning from the larval stage. Because of the significant impact of metamorphosis on animal life, its timing is strictly regulated. Invertebrate chordate ascidians are the closest living relatives of vertebrates. Ascidians exhibit metamorphosis that converts their swimming larvae into sessile adults. Ascidian metamorphosis is triggered by a mechanical stimulus generated when adhesive papillae adhere to a substrate. However, it is not well understood how the mechanical stimulus is generated and how ascidian larvae sense the stimulus. In this study, we addressed these issues by a combination of embryological, molecular, and genetic experiments in the model ascidian Ciona intestinalis Type A, also called Ciona robusta. We here showed that the epidermal neuronal network starting from the sensory neurons at the adhesive papillae is responsible for the sensing of adhesion. We also found that the transient receptor potential (TRP) channel PKD2 is involved in sensing the stimulus of adhesion. Our results provide a better understanding of the mechanisms underlying the regulation of the timing of ascidian metamorphosis.

Comparison of differentiation gene batteries for migratory mechanosensory neurons across bilaterians.

In embryos of distantly related bilaterian phyla, their lateral neural borders give rise to the peripheral nervous system elements, including various mechanosensory cells derived from migratory precursors, such as hair cells and dorsal root ganglion (DRG) neurons in vertebrates, bipolar tail neuron (BTN) in Ciona, chordotonal organ in Drosophila, and AVM/PVM in Caenorhabditis elegans. Developmental genetics studies had revealed a couple of transcription factors (TFs) regulating differentiation of mechanosensory cells shared by vertebrates and arthropods. However, unbiased systematic profiling of regulators is needed to demonstrate conservation of differentiation gene batteries for mechanosensory cells across bilaterians. At first, we observed that in both C. elegans Q neuroblasts and Drosophila lateral neuroectoderm, conserved NPB specifier Msx/vab-15 regulates Atoh1/lin-32, supporting the homology of mechanosensory neuron development in lateral neural border lineage of Ecdysozia. So we used C. elegans as a protostomia model. Single-cell resolution expression profiling of TFs and genetic analysis revealed a differentiation gene battery (Atonh1/lin-32, Drg11/alr-1, Gfi1/pag-3, Lhx5/mec-3, and Pou4/unc-86) for AVM/PVM mechanosensory neurons. The worm-gene battery significantly overlaps with both that of placode-derived Atonh1/lin-32-dependent hair cells and that of NPB-derived Neurogenin-dependent DRG neurons in vertebrates, supporting the homology of molecular mechanisms underlying the differentiation of neural border-derived mechanosensory cells between protostome and deuterostome. At last, Ciona BTN, the homolog of vertebrate DRG, also expresses Atonh1/lin-32, further supporting the homology notion and indicating a common origin of hair cells and DRG in vertebrate lineage.

Conserved gene regulatory module specifies lateral neural borders across bilaterians.

The lateral neural plate border (NPB), the neural part of the vertebrate neural border, is composed of central nervous system (CNS) progenitors and peripheral nervous system (PNS) progenitors. In invertebrates, PNS progenitors are also juxtaposed to the lateral boundary of the CNS. Whether there are conserved molecular mechanisms determining vertebrate and invertebrate lateral neural borders remains unclear. Using single-cell-resolution gene-expression profiling and genetic analysis, we present evidence that orthologs of the NPB specification module specify the invertebrate lateral neural border, which is composed of CNS and PNS progenitors. First, like in vertebrates, the conserved neuroectoderm lateral border specifier Msx/vab-15 specifies lateral neuroblasts in Caenorhabditis elegans Second, orthologs of the vertebrate NPB specification module (Msx/vab-15, Pax3/7/pax-3, and Zic/ref-2) are significantly enriched in worm lateral neuroblasts. In addition, like in other bilaterians, the expression domain of Msx/vab-15 is more lateral than those of Pax3/7/pax-3 and Zic/ref-2 in C. elegans Third, we show that Msx/vab-15 regulates the development of mechanosensory neurons derived from lateral neural progenitors in multiple invertebrate species, including C. elegans, Drosophila melanogaster, and Ciona intestinalis We also identify a novel lateral neural border specifier, ZNF703/tlp-1, which functions synergistically with Msx/vab-15 in both C. elegans and Xenopus laevis These data suggest a common origin of the molecular mechanism specifying lateral neural borders across bilaterians.

Possible Serological Markers to Predict Mortality in Acute Exacerbation of Idiopathic Pulmonary Fibrosis.

Background and objectives: Idiopathic pulmonary fibrosis (IPF) has a particularly poor prognosis, and most IPF-related deaths are due to acute exacerbation (AE) of this condition. Few reports about biomarkers to predict prognosis of AE-IPF have been published since the release of the new AE-IPF criteria in 2016. The present study investigated relationships between serological markers and in-hospital mortality after the onset of AE-IPF. Methods: Demographic, serological, and imaging data from patients hospitalized at the Maebashi Red Cross Hospital (Gunma, Japan) between 1 January 2013, and 31 December 2017, were retrospectively reviewed. Subjects fulfilling the diagnostic criteria for AE-IPF were divided into those who survived or died; statistical analysis of risk factors was performed using data from these two groups. Results: Diagnostic criteria for AE-IPF were fulfilled by 84 patients (59 males (70.2%)), with a median age of 78 years (range, 56-95 years). IPF was diagnosed before hospitalization in 50 (59.5%) patients and 38 (45.2%) died in hospital. Among the serological markers at hospitalization in the deceased group, C-reactive protein (CRP) was significantly higher than in the survivor group (p = 0.002), while total serum protein (p = 0.031), albumin (p = 0.047) and total cholesterol (p = 0.039) were significantly lower. Cox hazard analysis of factors predicting mortality, corrected for age, sex and BMI, revealed the following: CRP (hazard ratio (HR) 1.080 (95% confidence interval (CI) 1.022-1.141); p = 0.006), LDH (HR 1.003 (95% CI 1.000-1.006); p = 0.037), and total cholesterol (HR 0.985 (95% CI 0.972-0.997); p = 0.018). Conclusions: Our data suggest that CRP, LDH, and total cholesterol may be biomarkers predicting mortality in patients with AE-IPF. However, only prospective controlled studies can confirm or not our observation as a generalizable one.

Ependymal cells of chordate larvae are stem-like cells that form the adult nervous system.

In ascidian tunicates, the metamorphic transition from larva to adult is accompanied by dynamic changes in the body plan. For instance, the central nervous system (CNS) is subjected to extensive rearrangement because its regulating larval organs are lost and new adult organs are created. To understand how the adult CNS is reconstructed, we traced the fate of larval CNS cells during ascidian metamorphosis by using transgenic animals and imaging technologies with photoconvertible fluorescent proteins. Here we show that most parts of the ascidian larval CNS, except for the tail nerve cord, are maintained during metamorphosis and recruited to form the adult CNS. We also show that most of the larval neurons disappear and only a subset of cholinergic motor neurons and glutamatergic neurons are retained. Finally, we demonstrate that ependymal cells of the larval CNS contribute to the construction of the adult CNS and that some differentiate into neurons in the adult CNS. An unexpected role of ependymal cells highlighted by this study is that they serve as neural stem-like cells to reconstruct the adult nervous network during chordate metamorphosis. Consequently, the plasticity of non-neuronal ependymal cells and neuronal cells in chordates should be re-examined by future studies.

Neuronal map reveals the highly regionalized pattern of the juvenile central nervous system of the ascidian Ciona intestinalis.

BACKGROUND: The dorsally located central nervous system (CNS) is an important hallmark of chordates. Among chordates, tunicate ascidians change their CNS remarkably by means of a metamorphosis from a highly regionalized larval CNS to an oval-shaped juvenile CNS without prominent morphological features. The neuronal organization of the CNS of ascidian tadpole larvae has been well described, but that in the CNS of postmetamorphosis juveniles has not been characterized well. RESULTS: We investigated the number of neural cells, the number and position of differentiated neurons, and their axonal trajectories in the juvenile CNS of the ascidian Ciona intestinalis. The cell bodies of cholinergic, glutamatergic, and GABAergic/glycinergic neurons exhibited different localization patterns along the anterior-posterior axis in the juvenile CNS. Cholinergic neurons extended their axons toward the oral, atrial and body wall muscles and pharyngeal gill to regulate muscle contraction and ciliary movement. CONCLUSIONS: Unlike its featureless shape, the juvenile CNS is highly patterned along the anterior-posterior axis. This patterning may be necessary for exerting multiple roles in the regulation of adult tissues distributed throughout the body. This basic information of the juvenile CNS of Ciona will allow in-depth studies of molecular mechanisms underlying the reconstruction of the CNS during ascidian metamorphosis.

Outline of pharmacotherapy of adult bronchial asthma based on Japanese asthma guideline 2015.

Recent Japanese asthma guideline was published in 2015 (JGL2015). Variability of asthma symptom and airflow limitation was added to its definition. Updated information of pharma- cotherapy in adult asthma was documented. Long-acting anticholinergics as add-on therapy to inhaled corticosteroids combined with a LABA were incorporated into the step 3 and 4 in adult patients. Clinician should confirm treatment adherence and correct inhaler technique, and take enough time to discuss about treatment at every visit.

Retinoic acid-driven Hox1 is required in the epidermis for forming the otic/atrial placodes during ascidian metamorphosis.

Retinoic acid (RA)-mediated expression of the homeobox gene Hox1 is a hallmark of the chordate central nervous system (CNS). It has been suggested that the RA-Hox1 network also functions in the epidermal ectoderm of chordates. Here, we show that in the urochordate ascidian Ciona intestinalis, RA-Hox1 in the epidermal ectoderm is necessary for formation of the atrial siphon placode (ASP), a structure homologous to the vertebrate otic placode. Loss of Hox1 function resulted in loss of the ASP, which could be rescued by expressing Hox1 in the epidermis. As previous studies showed that RA directly upregulates Hox1 in the epidermis of Ciona larvae, we also examined the role of RA in ASP formation. We showed that abolishment of RA resulted in loss of the ASP, which could be rescued by forced expression of Hox1 in the epidermis. Our results suggest that RA-Hox1 in the epidermal ectoderm played a key role in the acquisition of the otic placode during chordate evolution.

Nonreproductive role of gonadotropin-releasing hormone in the control of ascidian metamorphosis.

BACKGROUND: Gonadotropin-releasing hormones (GnRHs) are neuropeptides that play central roles in the reproduction of vertebrates. In the ascidian Ciona intestinalis, GnRHs and their receptors are expressed in the nervous systems at the larval stage, when animals are not yet capable of reproduction, suggesting that the hormones have non-reproductive roles. RESULTS: We showed that GnRHs in Ciona are involved in the animal's metamorphosis by regulating tail absorption and adult organ growth. Absorption of the larval tail and growth of the adult organs are two major events in the metamorphosis of ascidians. When larvae were treated with GnRHs, they completed tail absorption more frequently than control larvae. cAMP was suggested to be a second messenger for the induction of tail absorption by GnRHs. tGnRH-3 and tGnRH-5 (the "t" indicates "tunicate") inhibited the growth of adult organs by arresting cell cycle progression in parallel with the promotion of tail absorption. CONCLUSIONS: This study provides new insights into the molecular mechanisms of ascidian metamorphosis conducted by non-reproductive GnRHs.

Enhancer activity sensitive to the orientation of the gene it regulates in the chordate genome.

Enhancers are flexible in terms of their location and orientation relative to the genes they regulate. However, little is known about whether the flexibility can be applied in every combination of enhancers and genes. Enhancer detection with transposable elements is a powerful method to identify enhancers in the genome and to create marker lines expressing fluorescent proteins in a tissue-specific manner. In the chordate Ciona intestinalis, this method has been established with a Tc1/mariner superfamily transposon Minos. Previously, we created the enhancer detection line E[MiTSAdTPOG]15 (E15) that specifically expresses green fluorescent protein (GFP) in the central nervous system (CNS) after metamorphosis. In this study, we identified the causal insertion site of the transgenic line. There are two genes flanking the causal insertion of the E15 line, and the genomic region around the insertion site contains the enhancers responsible for the expression in the endostyle and gut in addition to the CNS. We found that the endostyle and gut enhancers show sensitivity to the orientation of the GFP gene for their enhancer activity. Namely, the enhancers cannot enhance the expression of GFP which is inserted at the same orientation as the E15 line, while the enhancers can enhance GFP expression inserted at the opposite orientation. The CNS enhancer can enhance GFP expression in both orientations. The DNA element adjacent to the endostyle enhancer is responsible for the orientation sensitivity of the enhancer. The different sensitivity of the enhancers to the orientation of the transgene is a cause of CNS-specific GFP expression in the E15 line.

[Current status of "hospital-clinic" and "hospital-pharmacy" cooperation for inhalation therapy -based on hospital surveys throughout Japan].

BACKGROUND: The "zero death from asthma strategy" in the medical treatment for bronchial asthma has been promoted by the Ministry of Health, Labour, and Welfare from 2006, and it indicates that medical and non-medical specialists, as well as pharmacists, should cooperate, and strives to build cooperation which is suited the actual conditions of an area. It is also important for COPD. Although hospitals in some areas cooperate with clinics and pharmacies, the overall concept of cooperation appears to be absent in most Japanese hospitals. METHOD: A questionnaire was administered in early March, 2012 to 477 allergology institutions, and was authorized by an educational establishment. RESULT: Among 246 replies from the institutions, cooperation between hospitals and clinics was carried out by 98 institutions (39.8%) specializing in bronchial asthma, and in 64 institutions (37.2%) specializing in COPD. However, cooperation tools were used in only 37 of these institutions (15.0%). The ability to fill prescriptions outside the hospital was available in 209 institutions (85.0%). One-hundred and seventeen institutions (47.6%) replied that they have no tools for hospital-pharmacy cooperation. Direct indications were written in prescriptions by 82 institutions (33.3). CONCLUSION: In order to build inter-regional association and to equalize medical treatment, we suggest that developing tools and organization for cooperation between health professionals who treat patients with bronchial asthma and COPD is necessary.

doublesex/mab3 related-1 (dmrt1) is essential for development of anterior neural plate derivatives in Ciona.

Ascidian larvae have a hollow, dorsal central nervous system that shares many morphological features with vertebrate nervous systems yet is composed of very few cells. We show here that a null mutation in the gene dmrt1 in the ascidian Ciona savignyi results in profound abnormalities in the development of the sensory vesicle (brain), as well as other anterior ectodermal derivatives, including the palps and oral siphon primordium (OSP). Although the phenotype of the mutant embryos is variable, the majority have a complete loss of the most anterior structures (palps and OSP) and extensive disruption of sensory structures, such as the light-sensitive ocellus, in the sensory vesicle. dmrt1 is expressed early in the blastula embryo in a small group of presumptive ectodermal cells as they become restricted to anterior neural, OSP and palp fates. Despite the early and restricted expression of dmrt1, we were unable, using several independent criteria, to observe a defect in the mutant embryos until the early tailbud stage. We speculate that the variability and late onset in the phenotype may be due to partially overlapping activities of other gene products.

Monoaminergic modulation of photoreception in ascidian: evidence for a proto-hypothalamo-retinal territory.

BACKGROUND: The retina of craniates/vertebrates has been proposed to derive from a photoreceptor prosencephalic territory in ancestral chordates, but the evolutionary origin of the different cell types making the retina is disputed. Except for photoreceptors, the existence of homologs of retinal cells remains uncertain outside vertebrates. METHODS: The expression of genes expressed in the sensory vesicle of the ascidian Ciona intestinalis including those encoding components of the monoaminergic neurotransmission systems, was analyzed by in situ hybridization or in vivo transfection of the corresponding regulatory elements driving fluorescent reporters. Modulation of photic responses by monoamines was studied by electrophysiology combined with pharmacological treatments. RESULTS: We show that many molecular characteristics of dopamine-synthesizing cells located in the vicinity of photoreceptors in the sensory vesicle of the ascidian Ciona intestinalis are similar to those of amacrine dopamine cells of the vertebrate retina. The ascidian dopamine cells share with vertebrate amacrine cells the expression of the key-transcription factor Ptf1a, as well as that of dopamine-synthesizing enzymes. Surprisingly, the ascidian dopamine cells accumulate serotonin via a functional serotonin transporter, as some amacrine cells also do. Moreover, dopamine cells located in the vicinity of the photoreceptors modulate the light-off induced swimming behavior of ascidian larvae by acting on alpha2-like receptors, instead of dopamine receptors, supporting a role in the modulation of the photic response. These cells are located in a territory of the ascidian sensory vesicle expressing genes found both in the retina and the hypothalamus of vertebrates (six3/6, Rx, meis, pax6, visual cycle proteins). CONCLUSION: We propose that the dopamine cells of the ascidian larva derive from an ancestral multifunctional cell population located in the periventricular, photoreceptive field of the anterior neural tube of chordates, which also gives rise to both anterior hypothalamus and the retina in craniates/vertebrates. It also shows that the existence of multiple cell types associated with photic responses predates the formation of the vertebrate retina.

Improved Genome Editing in the Ascidian Ciona with CRISPR/Cas9 and TALEN.

The ascidian Ciona intestinalis type A (or Ciona robusta) is an important organism for elucidating the mechanisms that make the chordate body plan. CRISPR/Cas9 and TAL effector nuclease (TALEN) are widely used to quickly address genetic functions in Ciona. Our previously reported method of CRISPR/Cas9-mediated mutagenesis in this animal has inferior mutation rates compared to those of TALENs. We here describe an updated way to effectively mutate genes with CRISPR/Cas9 in Ciona. Although the construction of TALENs is much more laborious than that of CRISPR/Cas9, this technique is useful for tissue-specific knockouts that are not easy even by the optimized CRISPR/Cas9 method.

Expression of neuropeptide- and hormone-encoding genes in the Ciona intestinalis larval brain.

Despite containing only approximately 330 cells, the central nervous system (CNS) of Ciona intestinalis larvae has an architecture that is similar to the vertebrate CNS. Although only vertebrates have a distinct hypothalamus-the source of numerous neurohormone peptides that play pivotal roles in the development, function, and maintenance of various neuronal and endocrine systems, it is suggested that the Ciona brain contains a region that corresponds to the vertebrate hypothalamus. To identify genes expressed in the brain, we isolated brain vesicles using transgenic embryos carrying Ci-ß-tubulin(promoter)::Kaede, which resulted in robust Kaede expression in the larval CNS. The associated transcriptome was investigated using microarray analysis. We identified 565 genes that were preferentially expressed in the larval brain. Among these genes, 11 encoded neurohormone peptides including such hypothalamic peptides as gonadotropin-releasing hormone and oxytocin/vasopressin. Six of the identified peptide genes had not been previously described. We also found that genes encoding receptors for some of the peptides were expressed in the brain. Interestingly, whole-mount in situ hybridization showed that most of the peptide genes were expressed in the ventral brain. This catalog of the genes expressed in the larval brain should help elucidate the evolution, development, and functioning of the chordate brain.

Ascidians as excellent chordate models for studying the development of the nervous system during embryogenesis and metamorphosis.

The swimming larvae of the chordate ascidians possess a dorsal hollowed central nervous system (CNS), which is homologous to that of vertebrates. Despite the homology, the ascidian CNS consists of a countable number of cells. The simple nervous system of ascidians provides an excellent experimental system to study the developmental mechanisms of the chordate nervous system. The neural fate of the cells consisting of the ascidian CNS is determined in both autonomous and non-autonomous fashion during the cleavage stage. The ascidian neural plate performs the morphogenetic movement of neural tube closure that resembles that in vertebrate neural tube formation. Following neurulation, the CNS is separated into five distinct regions, whose homology with the regions of vertebrate CNS has been discussed. Following their larval stage, ascidians undergo a metamorphosis and become sessile adults. The metamorphosis is completed quickly, and therefore the metamorphosis of ascidians is a good experimental system to observe the reorganization of the CNS during metamorphosis. A recent study has shown that the major parts of the larval CNS remain after the metamorphosis to form the adult CNS. In contrast to such a conserved manner of CNS reorganization, most larval neurons disappear during metamorphosis. The larval glial cells in the CNS are the major source for the formation of the adult CNS, and some of the glial cells produce adult neurons.