Dorsal Forebrain-Specific Deficiency of Reelin-Dab1 Signal Causes Behavioral Abnormalities Related to Psychiatric Disorders.

Reelin-Dab1 signaling is involved in brain development and neuronal functions. The abnormalities in the signaling through either reduction of Reelin and Dab1 gene expressions or the genomic mutations in the brain have been reported to be associated with psychiatric disorders. However, it has not been clear if the deficiency in Reelin-Dab1 signaling is responsible for symptoms of the disorders. Here, to examine the function of Reelin-Dab1 signaling in the forebrain, we generated dorsal forebrain-specific Dab1 conditional knockout mouse (Dab1 cKO) and performed a behavioral test battery on the Dab1 cKO mice. Although conventional Dab1 null mutant mice exhibit cerebellar atrophy and cerebellar ataxia, the Dab1 cKO mice had normal cerebellum and showed no motor dysfunction. Dab1 cKO mice exhibited behavioral abnormalities, including hyperactivity, decreased anxiety-like behavior, and impairment of working memory, which are reminiscent of symptoms observed in patients with psychiatric disorders such as schizophrenia and bipolar disorder. These results suggest that deficiency of Reelin-Dab1 signal in the dorsal forebrain is involved in the pathogenesis of some symptoms of human psychiatric disorders.

A rare arterial branch distributing to the thymus, ectopic intrathymic parathyroid, and thyroid glands which passed ventral to the right common carotid artery: a case report.

The intrathymic parathyroid has been reported that this variation might be related with the hyperthyroidism. In this study, the arterial pattern supplying the intrathymic parathyroid was examined in detail in the human cadaver (67-year-old, female, right side). The ectopic parathyroid was only detected on the right side, but not on the left side. This ectopic intrathymic parathyroid was supplied by the supernumerary arterial branch that originated from the inferior thyroid artery and passed ventral to the common carotid artery. This supernumerary branch further divided into two thin branches: (1) the one distributing the intrathymic parathyroid and the right lobe of the thyroid gland and (2) the other descending toward the thoracic cavity to supply the mediastinum organs. Other arteries supplying the thyroid gland and thymus of both sides were normal. In the surgical resection of the ectopic intrathymic parathyroid, physicians should pay attention to arteries ventral to the common carotid artery. This supernumerary branch distributing to the intrathymic parathyroid may be caused by incomplete division into the primordium for the inferior parathyroid and the primordium for the thymus on the developmental process.

Gross anatomical study on the human myocardial bridges with special reference to the spatial relationship among coronary arteries, cardiac veins, and autonomic nerves.

Coronary arteries are frequently covered by cardiac muscles. This arrangement is termed a myocardial bridge. Previous studies have shown that myocardial bridges can cause myocardial ischemic diseases or cardiac arrhythmia, but the relevant pathogenic mechanisms remain unknown. We examined 60 hearts from Japanese cadavers macroscopically to clarify the spatial relationships among coronary arteries, cardiac veins and autonomic nerves. We found 86 myocardial bridges in 47 hearts from the 60 cadavers examined (78.3%). Next, we dissected out nine hearts with myocardial bridges in detail under the operating microscope. We found no additional branches of coronary arteries on the myocardial bridge surfaces. However, the cardiac veins, which usually accompany the coronary arteries, ran independently on the myocardial bridge surfaces in the same region. Cardiac autonomic nerves comprised two rami: one was associated with the coronary artery under the myocardial bridge and the other ran on the surface of the bridge. Such spatial relationships among the coronary arteries, cardiac veins and cardiac autonomic nerves at the myocardial bridges are quite similar to those in mouse embryo hearts.

Anatomical observations of the human acromioclavicular joint.

The condition of the acromioclavicular joint (ACJ) is considered to be one factor in the etiology of shoulder impingement syndrome, but there are few supporting morphological data. Fifty-two sides of 35 cadavers were investigated macroscopically and histologically using Safranin O, Fast green, and Weigert's iron hematoxylin staining. The ACJs were classified into three major types depending on the presence or absence of the articular disk. In type 1, the articular disk divided the articular cavity completely (n = 2; 3.8%). In type 2, the articular disk was incomplete and divided the joint cavity incompletely (n = 13; 25%). Type 2 was further divided into subtypes 2a and 2b depending on the configurations of the articular facets. In type 3, no articular disk was found in the joint cavity (n = 37; 71.2%). Type 3 was further divided into subtypes 3a, 3b, and 3c depending on the configurations of the articular surfaces. Fewer than half of the ACJ specimens (22/52 or 42.3%) demonstrated an ellipsoid character in which axial rotation was limited. Histological observation revealed that the upper part of the articular disk of the ACJ comprised fibrocartilage while the lower part comprised dense connective tissue. In cases where the ACJ appears to be an ellipsoid joint, its limited axial rotation restricts posterior tilting of the scapula during arm elevation, which could contribute to shoulder impingement syndrome.

Morphological analysis of the branches of the dorsal pancreatic artery and their clinical significance.

The dorsal pancreatic artery (DP), characterized by a course that crosses behind the proximal part of the splenic vein. It is regarded as clinically important, providing essential distribution to the pancreas. However, the origin of the DP is extremely variable and therefore cannot provide a sufficient basis for identifying it. The DPs of 11 cadavers were investigated in terms of origin, course and distribution. A total of 45 branches of the DP are classified into seven types on the basis of course and distribution. One of these seven types was consistently observed among the specimens: it ran to the right, passed behind the superior mesenteric vein and anterior surface of the posterior part of the head of the pancreas, and then distributed to the uncinate process and the posterior part of the head of the pancreas. Variations in the branching pattern of the DP can be explained from the following perspective: the consistent branch (#5) is the stem of the DP, and other branches originate from it. It is advisable for surgeons to pay attention to this consistent branch of the dorsal pancreatic artery when performing a pancreaticoduodenectomy.

Dynamic changes in the gene expression of zebrafish Reelin receptors during embryogenesis and hatching period.

The brain morphology of vertebrates exhibits huge evolutionary diversity, but one of the shared morphological features unique to vertebrate brain is laminar organization of neurons. Because the Reelin signal plays important roles in the development of the laminar structures in mammalian brain, investigation of Reelin signal in lower vertebrates will give some insights into evolution of vertebrate brain morphogenesis. Although zebrafish homologues of Reelin, the ligand, and Dab1, a cytoplasmic component of the signaling pathway, have been reported, the Reelin receptor molecules of zebrafish are not reported yet. Here, we sought cDNA sequence of zebrafish homologue of the receptors, vldlr and apoer2, and examined their expression patterns by in situ hybridization. Developmental gene expression pattern of reelin, dab1, vldlr, and apoer2 in the central nervous system of zebrafish was compared, and their remarkable expression was detected in the developing laminar structures, such as the tectum and the cerebellum, and also non-laminated structures, such as the pallium. The Reelin receptors exhibited different spatial and temporal gene expression. These results suggest a possibility that duplication and subsequent functional diversity of Reelin receptors contributed to the morphological and functional evolution of vertebrate brain.

Pseudoganglion on the connecting branch between the deep branch of the lateral plantar nerve and medial plantar nerve.

The connecting branch between the deep branch of the lateral plantar nerve and medial plantar nerve often has an enlarged site. We investigated these enlarged sites of the connecting branches. We observed the 22 human feet of 20 Japanese cadavers. We investigated the connecting branch macroscopically and histologically. We found the connecting branches between the deep branch of the lateral plantar nerve and medial plantar nerve in 19 feet out of 22 feet. This connecting nerve branch was interposed between the tendon of the flexor hallucis longus and the flexor hallucis brevis, and there enlarged in the anteroposterior direction. After penetration, numbers of fascicles of this connecting branch were increased at the enlarged site. In this region, the connective tissues surrounding the nerve fascicles and vessels were more developed compared with the adjoining sides of this branch. A few fascicles at this enlarged site innervated the first metatarsophalangeal joint capsule. Other nerve fascicles arose from the connecting branch and branched off muscular branches to the flexor hallucis brevis. This branch possibly receives the physical exertion or friction during gait due to its position. Deformity and overload of the foot can cause sensory disorders of the foot, but the anatomical basis for the relationship between the deformity/overload and sensory disorders of the foot is unclear. We discussed that this connecting branch can be a potential cause of pressure neuropathies in the human foot.

Expression of strawberry notch family genes during zebrafish embryogenesis.

Our previous study suggested a possible role for Sbno1, a mouse homologue of strawberry notch gene during brain development. In this report, we cloned the zebrafish homologues of sbno, and examined their expression pattern during embryogenesis by whole-mount in situ hybridization. Zebrafish have three sbno genes: one Sbno1 homologue and two Sbno2 homologues, sbno2a and sbno2b. We observed that the expression of sbno1 and sbno2a was initially ubiquitous and gradually became predominant in the central nervous system as development progressed. The expression of sbno2b was observed in non-neural tissues in contrast to the other two genes. sbno1 and sbno2a exhibited higher expression in distinct regions within the nervous system of pharyngula-stage embryos, suggesting possible differing roles for sbno1 and sbno2a during later stages of embryogenesis. Together, the observed gene expression patterns suggest an important role of sbno-family genes during development of the vertebrate central nervous system.

The transcriptional repressor RP58 is crucial for cell-division patterning and neuronal survival in the developing cortex.

The neocortex and the hippocampus comprise several specific layers containing distinct neurons that originate from progenitors at specific development times, under the control of an adequate cell-division patterning mechanism. Although many molecules are known to regulate this cell-division patterning process, its details are not well understood. Here, we show that, in the developing cerebral cortex, the RP58 transcription repressor protein was expressed both in postmitotic glutamatergic projection neurons and in their progenitor cells, but not in GABAergic interneurons. Targeted deletion of the RP58 gene led to dysplasia of the neocortex and of the hippocampus, reduction of the number of mature cortical neurons, and defects of laminar organization, which reflect abnormal neuronal migration within the cortical plate. We demonstrate an impairment of the cell-division patterning during the late embryonic stage and an enhancement of apoptosis of the postmitotic neurons in the RP58-deficient cortex. These results suggest that RP58 controls cell division of progenitor cells and regulates the survival of postmitotic cortical neurons.

GABA modulates development of cerebellar Purkinje cell dendrites under control of endocannabinoid signaling.

Purkinje cells (PCs) are the sole projection neurons in the cerebellar cortex with highly arborized dendrites, on which they receive glutamatergic and GABAergic inputs. Whereas influences of glutamatergic inputs on dendritic development of PCs have been well studied, those of GABA remain elusive. Here we examined effects of GABAergic signaling on dendritogenesis of PCs in dissociated cerebellar cultures. Treatment with GABA(A) agonists such as muscimol altered Purkinje dendrites to longer and less branched morphology, while GABA(A) antagonists resulted in shorter dendrites. In contrast, neither a GABA(B) agonist nor antagonist had major effects on dendritic morphology. Simultaneous addition of a glutamatergic antagonist cocktail or the Trk receptor antagonist K252a did not block muscimol. Furthermore, blockade of endocannabinoid signaling by either AM251 or tetrahydrolipstatin resulted in longer and less branched dendrites similar to those treated with GABA(A) agonists suggesting upstream regulation by endocannabinoids. Notably, whereas Purkinje dendrites extended in random directions in the presence of muscimol, they oriented to coexisting GABAergic interneurons when treated with AM251. Taken together, our results postulate the hypothesis that GABA released from the cerebellar interneurons modulates dendritogenesis of PCs in an endocannabinoid-dependent manner in the developing cerebellar cortex.

Dispersion of the neurons expressing layer specific markers in the reeler brain.

Neurons with similar functions including neuronal connectivity and gene expression form discrete condensed structures within the vertebrate brain. This is exemplified within the circuitry formed by the cortical layers and the neuronal nuclei. It is well known that the Reelin protein is required for development of these neuronal structures in rodents and human, but the function of Reelin remains controversial. In this report, we used "layer-specific markers" of the cerebral cortex to carry out detailed observations of spatial distribution of the neuronal subpopulations in the brain of the Reelin deficient mouse, reeler. We observed a spatially dispersed expression of the markers in the reeler cerebral cortex. These markers are expressed also in other laminated and non-laminated structures of brain, in which we observed similar abnormal gene expression. Our observations suggest that neurons within the brain structures (such as the layers and the nuclei), which normally exhibit condensed distribution of marker expressions, loosen their segregation or scatter by a lack of Reelin.

PKH26 is an excellent retrograde and anterograde fluorescent tracer characterized by a small injection site and strong fluorescence emission.

The fluorescent dye PKH26, which binds mainly to the cell membrane, has long stability that enables the tracing of PKH26-labeled transplanted cells in host tissue. In the present study, we examined whether this fluorescent dye works as a retrograde or anterograde tracer to label neural networks within the central nervous system of adult and postnatal day 3 (P3) mice. A small injection of the dye into the medullospinal junction resulted in the retrograde labeling of corticospinal tract (CST) neurons in layer V of the sensory-motor cortex both in the adult mice and pups. Injection of the dye into the motor cortex of the P3 pups resulted in the anterograde labeling of CST fibers at a single fiber resolution level, although a similar injection of the dye into the motor cortex of the adult mice failed to stain CST fibers anterogradely. These results suggest that, while PKH26 works as a retrograde or anterograde tracer, anterograde labeling of the adult tracts can not be expected.

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.

Dorsal telencephalon-specific RA-GEF-1 knockout mice develop heterotopic cortical mass and commissural fiber defect.

Neural migration defects lead to various types of human malformations of cortical development including subcortical band heterotopia, which shows formation of a secondary cortical plate beneath the primary cortex and is typically caused by mutation of the DCX (doublecortin) gene. Subcortical band heterotopia is usually associated with mental retardation and epilepsy. We previously discovered RA-GEF-1 as a guanine nucleotide exchange factor (GEF) for Rap1 small GTPase. Here we have analysed its in-vivo role in formation of the adult cerebral cortex by using telencephalon-specific RA-GEF-1 conditional knockout (cKO) mice, generated by mating RA-GEF-1(flox/flox) mice with Emx1-cre knockin mice. RA-GEF-1 cKO mice showed severe defects in their brain structures including an ectopic cortical mass underlying a relatively normal cortex. The ectopic cortical mass lacked the normal six-layered lamination but preserved the subcortical connectivity as revealed by retrograde tracing. Further, RA-GEF-1 cKO mice exhibited a lower threshold for the induction of epileptic seizures. These phenotypes have a resemblance to those of human subcortical band heterotopia. In addition, the agenesis of anterior commissures, the dorsal hippocampus commissure, the corpus callosum and the enlargement of the lateral ventricles were observed in cKO mice. Our findings suggest a crucial function of RA-GEF-1 in neural migration.

Developmental anatomy of reeler mutant mouse.

The reeler mouse is one of the most famous spontaneously occurring mutants in the research field of neuroscience, and this mutant has been used as a model animal to understand mammalian brain development. The classical observations emphasized that laminar structures of the reeler brain are highly disrupted. Molecular cloning of Reelin, the gene responsible for reeler mutant provided insights into biochemistry of Reelin signal, and some models had been proposed to explain the function of Reelin signal in brain development. However, recent reports of reeler found that non-laminated structures in the central nervous system are also affected by the mutation, making function of Reelin signal more controversial. In this review, we summarized reported morphological and histological abnormalities throughout the central nervous system of the reeler comparing to those of the normal mouse. Based on this overview of the reeler abnormalities, we discuss possible function of Reelin signal in the neuronal migration and other morphological events in mouse development.

Unusual patch-matrix organization in the retrosplenial cortex of the reeler mouse and Shaking rat Kawasaki.

The rat granular retrosplenial cortex (GRS) is a simplified cortex, with distinct stratification and, in the uppermost layers, distinct modularity. Thalamic and cortical inputs are segregated by layers and in layer 1 colocalize, respectively, with apical dendritic bundles originating from neurons in layers 2 or 5. To further investigate this organization, we turned to reelin-deficient reeler mouse and Shaking rat Kawasaki. We found that the disrupted lamination, evident in Nissl stains in these rodents, is in fact a patch-matrix mosaic of segregated afferents and dendrites. Patches consist of thalamocortical connections, visualized by vesicular glutamate transporter 2 (VGluT2) or AChE. The surrounding matrix consists of corticocortical terminations, visualized by VGluT1 or zinc. Dendrites concentrate in the matrix or patches, depending on whether they are OCAM positive (matrix) or negative (patches). In wild-type rodents and, presumably, mutants, OCAM(+) structures originate from layer 5 neurons. By double labeling for dendrites (filled by Lucifer yellow in fixed slice) and OCAM immunofluorescence, we ascertained 2 populations in reeler: dendritic branches either preferred (putative layer 5 neurons) or avoided (putative supragranular neurons) the OCAM(+) matrix. We conclude that input-target relationships are largely preserved in the mutant GRS and that dendrite-dendrite interactions involving OCAM influence the formation of the mosaic configuration.

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 beta-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.

Rac1 in cortical projection neurons is selectively required for midline crossing of commissural axonal formation.

Rac1 is a member of Rho family GTPases and regulates multiple cellular functions through actin cytoskeleton reorganization. During cerebral corticogenesis, Rac1 has been assumed to be involved in neuronal migration, neurite formation, polarization and axonal guidance. Here we show the specific role of Rac1, regulating midline crossing of commissural axons during cortical development by using cortex-restricted Rac1-knockout mice. In the knockout mice, Rac1 was eliminated from the beginning of corticogenesis exclusively in the dorsal telencephalon where progenitors of cortical projection neurons are located. Cortical lamination was distorted only mildly in the knockout mice, being preserved with six layers of neurons. However, cortex-restricted Rac1 deletion exhibited striking agenesis of commissural axons including the corpus callosum and anterior commissure without affecting other corticofugal axons including corticospinal and corticothalamic projections. Of note, the commissural axons of the knockout mice were potent in extending their process, but failed to cross the midline. Therefore, these findings indicate that Rac1 specifically controls the midline crossing of the commissural fibers, but not axonal formation of corticospinal or corticothalamic fibers during cortical development.

Establishment of framework of the cortical area is influenced by Otx1.

Subcortical projection from layer 5 neurons is the major cortical output. A transcription factor, Otx1, which is expressed in the layer 5 subcortical projection neurons in the visual cortex, was reported to be responsible for the establishment of visual area-specific layer 5 subcortical projections by inducing the sensorimotor cortex to adopt a visual cortex identity. However, we here demonstrate that the area of corticospinal neurons shifted caudo-medially in the Otx1-null mice of which cortex is 9 tenths in size compared with that of the wild-type littermates, while the whole visual cortex did not convert to the sensorimotor cortex in the absence of Otx1. This suggests that Otx1 is not crucial for the development of visual cortex identity but for the determination of the proportion of cortical areas to the whole neocortex.

Nerve communication between the glossopharyngeal nerve, external carotid plexus and the superficial cervical ansa: human autopsy case.

The authors encountered a very rare human autopsy case in which the supernumerary branch of the glossopharyngeal nerve and a nerve branch arising from the external carotid plexus communicated with the superficial cervical ansa. This anomaly was observed on the left side of a 71-year-old male cadaver during the gross anatomical seminar at Niigata University in 2004. The nerve fascicle and fiber analyses indicated that the supernumerary branch of the glossopharyngeal nerve separated cranial to the branches to the pharyngeal constrictor muscles, carotid sinus and stylopharyngeal muscle and sent the nerve fibers to the muscular branches to the platysma and the cutaneous branches to the cervical region. Additionally, it was shown that the branch arising from the external carotid plexus sent the nerve fibers to the cutaneous branch to the cervical region. Although the external carotid plexus is primarily postganglionic sympathetic fibers originating from the superior cervical ganglion, the vagus and glossopharyngeal nerves gave off branches connecting to the plexus, and therefore it was not possible to determine the origins of this branch of the external carotid plexus. The present nerve fascicle analysis demonstrates that the supernumerary branch of the glossopharyngeal nerve, which innervated the platysma, did not share any nerve components with the branches to the pharyngeal constrictor muscles, carotid sinus and stylopharyngeal muscle, suggesting that this supernumerary branch may be categorized into the different group from these well-known branches.