Profiles of Periglomerular Cells in the Olfactory Bulb of Prokineticin Type 2 Receptor-deficient Mice.

Both prokineticin receptor 2 (pkr2) and prokineticin 2 (pk2) gene-deficient mice have hypoplasia of the main olfactory bulb (MOB). This hypoplasia has been attributed to disruption of the glomerulus that is caused by loss of afferent projection from olfactory sensory neurons (OSN), and to the impaired migration of granule cells, a type of interneuron. In the present study, we examined whether migration of the second type of interneuron, periglomerular cells (PGC), is dependent on the pkr2 expression by observing the localization of distinct subpopulations of PGC: calretinin (CR)-, calbindin (CB)- and tyrosine hydroxylase (TH)-expressing neurons. In the Pkr2-/- mice, the construction of the layered structure of the MOB was partially preserved, with the exception of the internal plexiform layer (IPL) and the glomerular layer (GL). In the outermost layer of the MOB, abundant CR- and CB-immunopositive neurons were observed in the hypoplastic olfactory bulb. In addition, although markedly decreased, TH-immunopositive neurons were also observed in the outermost cell-dense region in the Pkr2-/-. The findings suggest that the migration of PGC to the MOB, as well as the migration from the core to the surface region of the MOB, is not driven by the PK2-PKR2 system.

Multiple muscular variations including tenuissimus and tensor fasciae suralis muscles in the posterior thigh of a human case.

The posterior thigh muscles on the right side of an 81-year-old male cadaver had multiple variations, denoted muscles I-IV. Muscle I originated from the posteromedial surface of the greater trochanter and divided into two muscle bellies. These muscle bellies fused with the long head of the biceps femoris and were innervated by two branches from muscular branches of the semitendinosus and the long head of the biceps. Muscle II separated from the medial surface of the long head of the biceps in the proximal third and fused with the semitendinosus in the distal fourth. Muscle III was a biventer muscle. Its superior belly separated from the medial surface of the long head of the biceps in the distal third. The inferior belly of this muscle fused with the posterior surface of the crural fascia and was innervated by the tibial nerve. Muscle IV separated from the adductor magnus muscle, passed between the long and short heads of the biceps, fused with the inferior belly of muscle III, and was innervated by the muscular branch of the common fibular nerve to the short head of the biceps. Peeling off the epineurium of the muscular branches to the inferior belly of muscle III showed that this nerve fascicle divided from the common trunk with branches to the gastrocnemius and soleus muscles. The inferior bellies of muscle III and muscle IV were thought to be equivalent to the tensor fasciae suralis and tenuissimus muscles, respectively.

Anatomical Study of the Brachial Plexus in the Common Marmoset (Callithrix Jacchus).

To elucidate the forelimb phylogeny of primates, anatomical analysis of the brachial plexus in platyrrhines is beneficial. In the present study, six brachial plexuses and the surrounding arteries of four common marmosets were dissected. In five specimens, the brachial plexus consisted of five ventral rami from the fifth cervical nerve (C5) to the first thoracic nerve (T1). In one specimen, the ventral ramus of the fourth cervical nerve joined with the brachial plexus. In five specimens, the upper trunk was composed of C5 and the sixth cervical nerve (C6). In one specimen, the ventral division of C6 merged with the ventral branch of the middle trunk to constitute the lateral cord. The seventh cervical nerve constituted the middle trunk, and the eighth cervical nerve and T1 formed the lower trunk in all specimens. The lateral cord gave rise to the musculocutaneous nerve, and the remaining component merged with the medial cord. The confluence of the lateral and medial cords immediately bifurcated into the median and ulnar nerves. These branching patterns of the musculocutaneous, median, and ulnar nerves were consistent and similar to the human counterparts. In the dorsal division, the single posterior cord as observed in the human brachial plexus was not observed. The axillary artery did not pass between the medial and lateral roots of the median nerve, and the axillary artery bifurcated into the brachial artery and the superficial brachial artery. Anat Rec, 300:1299-1306, 2017. © 2017 Wiley Periodicals, Inc.

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.

Promotion of colorectal cancer invasion and metastasis through activation of NOTCH-DAB1-ABL-RHOGEF protein TRIO.

UNLABELLED: We have recently identified a metastasis suppressor gene for colorectal cancer: AES/Aes, which encodes an endogenous inhibitor of NOTCH signaling. When Aes is knocked out in the adenomatous epithelium of intestinal polyposis mice, their tumors become malignant, showing marked submucosal invasion and intravasation. Here, we show that one of the genes induced by NOTCH signaling in colorectal cancer is DAB1/Dab1. Genetic depletion of DAB1 suppresses cancer invasion and metastasis in the NOTCH signaling-activated mice. DAB1 is phosphorylated by ABL tyrosine kinase, which activates ABL reciprocally. Consistently, inhibition of ABL suppresses cancer invasion in mice. Furthermore, we show that one of the targets of ABL is the RAC/RHOGEF protein TRIO, and that phosphorylation at its Tyr residue 2681 (pY2681) causes RHO activation in colorectal cancer cells. Its unphosphorylatable mutation TRIO Y2681F reduces RHOGEF activity and inhibits invasion of colorectal cancer cells. Importantly, TRIO pY2681 correlates with significantly poorer prognosis of patients with colorectal cancer after surgery. SIGNIFICANCE: These results indicate that TRIO pY2681 is one of the downstream effectors of NOTCH signaling activation in colorectal cancer, and can be a prognostic marker, helping to determine the therapeutic modality of patients with colorectal cancer.

Intramuscular nerve distribution pattern in the human tibialis posterior muscle.

The human tibialis posterior muscle (TPM) has developed to maintain the foot arches for adopting bipedal locomotion. The insertion tendon of this muscle is U-shaped in a cross section, and the fibular part of the muscle whose muscle fibers originated from the fibula has a unique architecture. To understand the developmental history of the human TPM, distribution pattern of intramuscular nerves was investigated in ten sides of eight cadavers. Muscular branches entering the TPM could be classified into five types according to the distribution pattern in the muscle. The nerves innervating the part deeper to the insertion tendon ran transversely in the medial direction toward the tibia. Muscular branches innervating the fibular part ran medioinferiorly, and in four cases had communicating rami with the nerve innervating the other parts of this muscle. Muscular branches innervating the superficial layer whose muscle bundles originated from the tibia ran laterally toward the fibula. These results suggest that the fibular part might develop from the common primordium of the TPM, and the deeper layer of the muscle might expand laterally during the development to acquire a new attaching space on the interosseous membrane and fibula. The superficial layer might also develop laterally to acquire the additional attaching spaces on the fascia of the flexor digitorum longus muscle and flexor hallucis longus muscle, thus the insertion tendon might become U-shaped. With these measures, the TPM might be able to increase muscle volume to yield stronger power.

Postnatal Development of the Corticospinal Tract in the Reeler Mouse.

Corticospinal tract (CST) neurons are dislocated in the motor cortex of Reelin-deficient mouse, reeler. In the present study, we examined whether postnatal axonal growth arising from these dislocated CST neurons are normal or not with use of anterograde tracer, DiI and retrograde tracer, HRP. A single injection of DiI into the motor cortex of the normal and reeler mice was made during postnatal period and 8-24 hours later, the animals were sacrificed to examine DiI-labeled CST axons at the lower medulla and spinal cord. Both in the normal and reeler mice, CST axons arrived at the pyramidal decussation and entered into the contralateral spinal cord around on postnatal day (P) 0.5, and descend in the ventral area of the contralateral dorsal funiculus at C2 level on P2, at C8 level on P3, at the mid-thoracic level on P4, and at the upper lumbar level on P8. The similar results were also demonstrated by the retrograde labeling of CST neurons with injection of HRP into the C1 level or upper lumbar enlargement. Next, we examined CaMKIIα expression in the CST axons of the adult normal and reeler mice. CaMKIIα-immunopositive fibers were recognized throughout the CST pathway from the internal capsule to the dorsal funiculus of the spinal cord both in the normal and reeler mice. The present study has demonstrated that ectopic location of cell bodies of reeler CST neurons do not affect postnatal development of CST axons in the spinal cord.

Expression of Beta Subunit 2 of Ca²+/Calmodulin-Dependent Protein Kinase I in the Developing Rat Retina.

Expression of beta 2 subunit of Ca²+/calmodulin-dependent protein kinase I (CaMKIß2) of the rat retina during the developmental period and in the adulthood was studied immunohistochemically. The immunoreactivity of CaMKIß2 was detected in the earliest development of the primordial retina at embryological day (E) 12. The inner neuroblastic layer from which the presumptive ganglion cells are generated showed the ubiquitous CaMKIß2 immunoreactivity at E15 and persistently expressed at the same level until postnatal day (P) 0 when the inner neuroblastic layer divides into the ganglionic cell layer and the inner plexiform layer. The strong immunoreactivity was detected in the ganglion cell layer and the moderate one in the internal plexiform layer. CaMKIß2 immunoreactivities were persistantly expressed throughout the postnatal development at the same level. The low level of intensity was first found in the inner nuclear layer at P7, followed by the outer plexiform, outer nuclear and rod-cone cell layers at the age of P12, respectively. The intensities of CaMKIß2 immunoreactivities in the inner nuclear and rod-cone cell layers were gradually increased to the strong level by P18 and persisted until adulthood. The present study revealed that the expression of CaMKIß2 in the retina was detected from the earliest development until adulthood, indicating that CaMKIß2 may be required in both proliferation and differentiation of the retinal precursor cells and subsequent formation of the functional layers. In addition, CaMKIß2 immunoreactivity in the rod-cone cell layer implies that this protein may be involved in the visual signaling process.

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.

Kif14 mutation causes severe brain malformation and hypomyelination.

We describe a novel spontaneous mouse mutant, laggard (lag), characterized by a flat head, motor impairment and growth retardation. The mutation is inherited as an autosomal recessive trait, and lag/lag mice suffer from cerebellar ataxia and die before weaning. lag/lag mice exhibit a dramatic reduction in brain size and slender optic nerves. By positional cloning, we identify a splice site mutation in Kif14. Transgenic complementation with wild-type Kif14-cDNA alleviates ataxic phenotype in lag/lag mice. To further confirm that the causative gene is Kif14, we generate Kif14 knockout mice and find that all of the phenotypes of Kif14 knockout mice are similar to those of lag/lag mice. The main morphological abnormality of lag/lag mouse is severe hypomyelination in central nervous system. The lag/lag mice express an array of myelin-related genes at significantly reduced levels. The disrupted cytoarchitecture of the cerebellar and cerebral cortices appears to result from apoptotic cell death. Thus, we conclude that Kif14 is essential for the generation and maturation of late-developing structures such as the myelin sheath, cerebellar and cerebral cortices. So far, no Kif14-deficient mice or mutation in Kif14 has ever been reported and we firstly define the biological function of Kif14 in vivo. The discovery of mammalian models, laggard, has opened up horizons for researchers to add more knowledge regarding the etiology and pathology of brain malformation.

Subcortically and callosally projecting neurons are distinct neuronal pools in the motor cortex of the reeler mouse.

Subcortically projecting neurons and callosally projecting ones are distinct neuronal pools in the cerebral cortex of the rodents. However, cortical efferent neurons are known to project multiple targets transiently by plural collateral axons. These plural axons are eliminated during prenatal and postnatal development. In the cerebral cortex of the Reelin-deficient mouse, reeler, which is caused by mutation of the reelin gene, cortical efferent neurons are ectopically distributed. However, it is still unknown whether cortical efferent neurons in the reeler mouse lose surplus collateral axons or maintain them during developmental periods. If surplus collaterals of malpositioned cortical neurons are not eliminated, neurons projecting subcortically may project their axons to the contralateral hemisphere. To test this plausible hypothesis, we made double injections of two fluorescent dyes, Fast Blue and Diamidino yellow dihydrochloride into two of three regions, i.e., upper cervical cord, ventral lateral thalamic nucleus, and contralateral motor cortex of the normal and reeler mice, to label corticospinal, corticothalamic and callosal commissure neurons in the motor cortex, retrogradely. No double labeled neurons were identified in the motor cortex of the normal and reeler mice, although the distribution patterns of these cortical efferent neurons were completely different between normal and reeler mice. These findings strongly suggest that collateral elimination of cortical efferent neurons during developing periods are not affected in this mutant mouse.

Characterization of heterotopic cell clusters in the hippocampus of the rat after prenatal treatment of methylazoxymethanol acetate.

Prenatal exposure of methylazoxymethanol acetate, a DNA methylating agent, to pregnant rats on embryonic day 15 is known to produce hippocampal malformation and laminar disorganization of the cerebral cortex. However, there are few studies to demonstrate developmental processes of abnormal structures in the hippocampus. In the present study, we examined complete serial sections of rat brains on postnatal day 0 to 2, which pretreated with methylazoxymethanol acetate on embryonic day 15. At birth, massive cellular clusters were found under the white matter of the cerebral cortex and then, a part of these clusters entered into the hippocampal CA1 sector on postnatal day 2. These ectopic cellular clusters in the CA1 were immunoreactive to anti-calbindin antibody, suggesting that the origin of these cellular clusters is equivalent to that of the cortical layer II/III neurons. Next, we injected FluoroGold into the lateral septal nucleus to examine hippocampo-septal projection. FluoroGold-labeled neurons were scattered in the ectopic cellular cluster, implying that CA1 pyramidal neurons project normally to the lateral septal nucleus. In conclusion, a majority of neurons found in the ectopic cellular cluster caused by prenatal methylazoxymethanol treatment is derived from cortical neurons, and some intrinsic pyramidal neurons in the CA1 of hippocampus are scattered throughout the ectopic cellular cluster.

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.

Function of strawberry notch family genes in the zebrafish brain development.

We previously reported embryonic expression pattern of strawberry notch (sbno) family genes, suggesting involvement in brain development. However function of sbno genes in the vertebrate development has not been known yet. Utilizing zebrafish embryos, we experimentally examined function of sbno genes during brain development in this report. Knockdown experiments of sbno1 and sbno2a disrupted brain morphology, and delayed developmental alteration of gene expression. The earliest effect of loss of function of sbno genes on the zebrafish embryogenesis that we found here was downregulation of otx2 expression. Knockdown of sbno1 specifically affects regionalization along the anterior-posterior axis of the brain. These results suggest essential roles of sbno genes in vertebrate brain development.

RA-GEF-1 (Rapgef2) is essential for proper development of the midline commissures.

The cerebral hemispheres are directly connected by three major interhemispheric fibers: the corpus callosum, the anterior commissure, and the hippocampal commissure. RA-GEF-1 (also termed Rapgef2) is a guanine nucleotide exchange factor responsible for sustained activation of Rap1. We previously reported anatomical defects of the major forebrain commissures in the adult dorsal telencephalon-specific RA-GEF-1 conditional knockout (cKO) mice. In this study, we use neuroanatomical tracing and immunohistochemistry to study the formation of the commissural fibers during early postnatal development. DiI anterograde tracing reveals the inability of the callosal axons to cross the midline in cKO mice, thereby forming Probst bundles on the ipsilateral side, which is associated with the absence of the indusium griseum glia and the glial sling at the cortical midline. Wheat germ agglutinin-conjugated horseradish peroxidase retrograde tracing verifies the agenesis of the anterior commissure in cKO mice, and DiI anterograde tracing confirms the deviation of the fibers from their original tract. As for the hippocampal commissure, agenesis and hypoplasia are observed in its dorsal and ventral parts, respectively. These results indicate the essential role of RA-GEF-1 in the proper formation of the cerebral midline commissures.

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.

Cortical layer V neurons in the auditory and visual cortices of normal, reeler, and yotari mice.

Both in the Reelin-deficient reeler and Dab1-deficient yotari mice, layer V corticospinal tract neurons in the sensory-motor cortex are radially spread instead of being confined to a single cortical layer. In the present study, we examined distribution pattern of cortical layer V neurons in the visual and auditory cortices of reeler and yotari mice with the injection of HRP into the superior and inferior colliculi of the adult animals, respectively. After the injection of HRP into the superior colliculus of the normal mouse, retrogradely labeled cells were distributed in layer V of the visual cortex, while the similar injection of HRP in the reeler and yotari mice produced radial dispersion of retrograde labeling through all of the depths of the visual cortex of these mutant mice. Next, we injected HRP into the inferior colliculus of the normal, reeler and yotari mice. Retrogradely labeled neurons were distributed in layer V of the normal auditory cortex, whereas they were again radially scattered in the auditory cortex of the reeler and yotari mice. Taken together with the previous and present findings, layer V cortical efferent neurons are radially scattered in the sensory-motor, visual and auditory cortices of the reeler and yotari mice.