Epithelial development of the urinary collecting system in the human embryo.

The urinary collecting system (UCS) consists of organized ducts that collect urine from the nephrons and transport it to the ureter and bladder. Understanding the histogenesis of the UCS is critical. Thirty human embryos between the Carnegie stages (CS) 18 and 23 were selected from the Congenital Anomaly Research Center, Kyoto, Japan. Epithelia of the UCS, ureter, and bladder of each sample were randomly selected. Histological findings of the epithelia were analyzed according to the following criteria: type of epithelium, presence or absence of glycogen, percentage of migrated nuclei, percentage of cells in mitosis, and the surrounding mesenchyme. A thickened epithelium lining a narrow luminal cavity was observed in the pre-expanded pelvic specimens at CS18-CS23. At CS23, after pelvic expansion, the UCS showed a thin epithelium with a large luminal cavity mainly located on the early branches, whereas the epithelium covering the subsequent branches had medium thickness. Histological characteristics differed depending on the UCS part and sample stage. The degree of differentiation was evaluated, revealing that in CS18-CS23 pre-expanded pelvis specimens, the undifferentiated epithelium was found in the zeroth to third/fifth generation, whereas at CS23, after pelvic expansion, a differentiated epithelium covered the UCS zeroth to seventh generation. In a comparison of the urothelial epithelium between the UCS, ureter, and bladder, we found that urinary tract differentiation may be initiated in the bladder, followed by the ureter, UCS zeroth to seventh generations, and finally, UCS eighth to end generations. An understanding of the histogenesis of embryonic stage UCS can aid in the clinical management of congenital urinary tract defects and other diseases.

Regional differences in the umbilical vein and ductus venosus at different stages of normal human development.

During the fetal period, oxygenated blood from the placenta flows through the umbilical vein (UV), portal sinus, ductus venosus (DV), and inferior vena cava (IVC) to the heart. This venous route varies regionally in many aspects. Herein, we sought to characterize the venous route's morphological features and regional differences during embryonic and early-fetal periods. Twenty-nine specimens were selected for high-resolution digitized imaging; 18 embryos were chosen for histological analysis. The venous route showed a primitive, large, S-shaped curved morphology with regional narrowing and dilation at Carnegie stage (CS) 15. Regional differences in vessel-wall differentiation became apparent from approximately CS20. The vessel wall was poorly developed in most DV parts; local vessel-wall thickness at the inlet was first detected at CS20. The lumen of the venous route changed from a nonuniform shape to a relatively round and uniform morphology after CS21. During the early-fetal period, two large bends were observed around the passage of the umbilical ring and at the inlet of the liver. The length ratio of the extrahepatic UV to the total venous route increased. The sectional area gradually increased during embryonic development, whereas differences in sectional area between the DV, UV, and IVC became more pronounced in the early-fetal period. Furthermore, differences in the sectional area between the narrowest part of the DV and other hepatic veins and the transverse sinus became more pronounced. In summary, the present study described morphological, morphometric, and histological changes in the venous route throughout embryonic and early-fetal development, clarifying regional characteristics.

Histological Properties of a Chemically Fixed Human Embryo Visualized with Quantitative Susceptibility Mapping.

A chemically fixed Carnegie stage 23 (approximately 56 days of gestation) human embryo specimen was imaged using 3D spin-echo and gradient-echo sequences in a static magnetic field strength of 4.74T, and a quantitative susceptibility map was calculated using the 3D gradient-echo image. The acquired 3D microscopic images (90 µm cube voxel size) clarified the relationship between R2 (transverse relaxation rate), R2* (apparent transverse relaxation rate), and magnetic susceptibility in the heart, liver, kidney, and spinal cord. The results suggested that the R2* and magnetic susceptibility in each tissue were probably due to paramagnetic iron ions originating from erythrocytes. The large R2* (~130 s-1) and magnetic susceptibility (~0.122 ppm) in the liver were attributed to its hemopoietic function. A large magnetic susceptibility (~0.116 ppm) was also observed in the spinal cord, but we conclude that more detailed future studies are needed.

Brain Structures in a Human Embryo Imaged with MR Microscopy.

PURPOSE: To delineate brain microstructures in human embryos during the formation of the various major primordia by MR microscopy, with different contrasts appropriate for each target. METHODS: We focused mainly on the internal structures in the cerebral cortex and the accessory nerves of the brain. To find appropriate sequence parameters, we measured nuclear magnetic resonance (NMR) parameters and created kernel density plots of T1 and T2 values. We performed T1-weighted gradient echo imaging with parameters similar to those used in the previous studies. We performed T2*-weighted gradient echo imaging to delineate the target structures with the appropriate sequence parameters according to the NMR parameter and flip angle measurements. We also performed high-resolution imaging with both T1- and T2*-weighted sequences. RESULTS: T1, T2, and T2* values of the target tissues were positively correlated and shorter than those of the surrounding tissues. In T1-weighted images with a voxel size of (30 µm)3 and (20 µm)3, various organs and tissues and the agarose gel were differentiated as in previous studies, and the structure of approximately 40 µm in size was depicted, but the detailed structures within the cerebral cortex and the accessory nerves were not delineated. In T2*-weighted images with a voxel size of (30 µm)3, the layered structure within the cerebral cortex and the accessory nerves were clearly visualized. Overall, T1-weighted images provided more information than T2*-weighted images, but important internal brain structures of interest were visible only in T2*-weighted images. Therefore, it is essential to perform MR microscopy with different contrasts. CONCLUSION: We have visualized brain structures in a human embryo that had not previously been delineated by MR microscopy. We discussed pulse sequences appropriate for the structures of interest. This methodology would provide a way to visualize crucial embryological information about the anatomical structure of human embryos.

Three-dimensional imaging analysis of the developmental process of posterior meniscofemoral ligaments in rat embryos.

INTRODUCTION: The posterior meniscofemoral ligament (pMFL) of knee joint is a ligament that runs posterior to the posterior cruciate ligament (PCL) and it is known that the height of the pMFL attachment site causes meniscus avulsion. Therefore, understanding the three-dimensional (3D) structure of the pMFL attachment site is essential to better understand the pathogenesis of meniscus disorders. However, the developmental process of pMFL has not been well investigated. The purpose of this study was to analyze pMFL development in rat knee joints using 3D reconstructed images produced from episcopic fluorescence image capture (EFIC) images and examine its relationship with other knee joint components. METHODS: Knee joints of Wistar rat embryos between embryonic day (E) 16 and E21 were observed with HE stained tissues. Serial EFIC images of the hindlimbs of E17-E21 were respectively captured, from which 3D images were reconstructed and the features of pMFL structure: length and angle, were measured. Besides, the chronological volume changes and the volume ratio of the knee joint components compared to E17 were calculated to identify the differences in growth by components. RESULTS: pMFL was observed from E17 and was attached to the medial femoral condyle and lateral meniscus at all developmental stages, as in mature rats. The lack of marked variation in the attachment site and angle of the pMFL with the developmental stage indicates that the pMFL and surrounding knee joint components developed while maintaining their positional relationship from the onset of development. CONCLUSION: Current results may support to congenital etiology of meniscus disorder.

Pyramidalis muscle formation during human embryonic and early fetal periods.

The pyramidalis muscle (PM) is a paired small triangular muscle of the anterior abdominal wall; however, its physiological significance is unclear. Recent studies have failed to detect this muscle during embryonic period. Hence, the present study aimed to determine the time when PM is emerging and reveal its features using high-resolution magnetic resonance imaging. Fourteen embryos between Carnegie stage (CS)18 and CS23 and 59 fetuses (crown-rump length: 39.5-185.0 mm) were selected for this study. The PM was first detected in one of the three samples at CS20. It was detected in five of the seven samples (71.4%) between CS21 and CS23. Forty-eight samples (81.4%) at early fetal period had PMs on both the right and left sides, and 3 (5.1%) had it only on the right side. Eight samples (13.6%) had no PMs. No side-differences or sexual dimorphisms were detected. The PM length was larger than the width in most samples, although the length/width ratio varied among the samples. The PM/rectus abdominis muscle length and PM/umbilicus-pubic symphysis length ratios were almost constant, irrespective of the crown-rump length. The PM was located ventrally inferior to the rectus abdominis and closer to the medial muscle groups of the lower limb than the rectus abdominis. The present study demonstrated that PM formation occurred in the late embryonic period, and that the frequency, side differences, sex dimorphism, and spatial position of the PM in the early fetal period were similar to those in adults.

Bronchial tree of the human embryo: Examination based on a mammalian model.

The symmetry of the right and left bronchi, proposed in a previous comparative anatomical study as the basic model of the mammalian bronchial tree, was examined to determine if it applied to the embryonic human bronchial tree. Imaging data of 41 human embryo specimens at Carnegie stages (CS) 16-23 (equivalent to 6-8 weeks after fertilization) belonging to the Kyoto collection were obtained using phase-contrast X-ray computed tomography. Three-dimensional bronchial trees were then reconstructed from these images. Bronchi branching from both main bronchi were labeled as dorsal, ventral, medial, or lateral systems based on the branching position with numbering starting cranially. The length from the tracheal bifurcation to the branching point of the labeled bronchus was measured, and the right-to-left ratio of the same labeled bronchus in both lungs was calculated. In both lungs, the human embryonic bronchial tree showed symmetry with an alternating pattern of dorsal and lateral systems up to segmental bronchus B9 as the basic shape, with a more peripheral variation. This pattern is similar to that described in adult human lungs. Bronchial length increased with the CS in all labeled bronchi, whereas the right-to-left ratio was constant at approximately 1.0. The data demonstrated that the prototype of the human adult bronchial branching structure is formed and maintained in the embryonic stage. The morphology and branching position of all lobar bronchi and B6, B8, B9, and the subsegmental bronchus of B10 may be genetically determined. On the other hand, no common structures between individual embryos were found in the peripheral branches after the subsegmental bronchus of B10, suggesting that branch formation in this region is influenced more by environmental factors than by genetic factors.

Morphogenesis of the pulmonary vein and left atrial appendage in human embryos and early fetuses.

The left atrium wall has several origins, including the body, appendage, septum, atrial-ventricular canal, posterior wall, and venous component. Here, we describe the morphogenesis of left atrium based on high-resolution imaging (phase-contrast X-ray computed tomography and magnetic resonance imaging). Twenty-three human embryos and 19 fetuses were selected for this study. Three-dimensional cardiac images were reconstructed, and the pulmonary veins and left atrium, including the left atrial appendage, were evaluated morphologically and quantitatively. The positions of the pericardial reflections were used as landmarks for the border of the pericardial cavity. The common pulmonary vein was observed in three specimens at Carnegie stages 17-18. The pericardium was detected at the four pulmonary veins (left superior, left inferior, right superior, and right inferior pulmonary veins) at one specimen at Carnegie stage 18 and all larger specimens, except the four samples. Our results suggest that the position of the pericardial reflections was determined at two pulmonary veins (right and left pulmonary vein) and four pulmonary veins almost simultaneously when the dorsal mesocardial connection between the embryo and heart regressed. The magnetic resonance images and reconstructed heart cavity images confirmed that the left atrium folds were present at the junction between the body and venous component. Three-dimensional reconstruction showed that the four pulmonary veins entered the dorsal left atrium tangentially from the lateral to the medial direction. More specifically, the right pulmonary veins entered at a greater angle than the left pulmonary veins. The distance between the superior and inferior pulmonary veins was shorter than that between the left and right pulmonary veins. Three-dimensional reconstruction showed that the venous component increased proportionally with growth. No noticeable differences in discrimination between the right and left parts of the venous component emerged, while the junction between the venous component and body gradually became inconspicuous but was still recognizable by the end of the observed early fetal period. The left superior pulmonary vein had the smallest cross-sectional area and most flattened shape, whereas the other three were similar in area and shape. The left atrial appendage had a large volume in the center and extended to the periphery as a lobe-like structure. The left atrial appendage orifice increased in the area and tended to become flatter with growth. The whole left atrium volume^(1/3) increased almost proportionally with growth, parallel to the whole heart volume. This study provided a three-dimensional and quantitative description of the developmental process of the left atrium, comprising the venous component and left atrial appendage formation, from the late embryonic to the early fetal stages.

High-resolution MRI for human embryos with isotropic 10 µm resolution at 9.4 T.

Magnetic resonance (MR) microscopy of human embryos has contributed significantly to the development of human embryology. Higher-resolution MR microscopy will have obvious benefits, for example, in visualizing small structures that are blurred or lost in lower-resolution images, providing detailed information on the development and growth of various organs, and improving the accuracy of MR volumetry. However, high-resolution MR microscopy has yet to be realized because of many technical challenges. In this study, therefore, we have performed high-resolution MR microscopy for human embryos with isotropic resolutions of (12 µm)3 at full sampling and (10 µm)3 at compressed sensing, which far exceeds the resolution of previous embryonic MR studies. The hardware and the pulse sequence were improved to achieve higher spatial resolution. Line profile, signal-to-noise ratio, and histogram analysis using phantom images were performed to verify that the resolution and the voxel size were identical. Comparison with optical microscopy images of embryo specimens at the same developmental stage was performed to confirm that the microstructures were well delineated. Our results show that imaging at this high resolution effectively depicts the microstructures of human embryos. This technology is the cornerstone for constructing an unprecedented high-quality atlas that will contribute to the development of human embryology.

Coronary artery established through amniote evolution.

Coronary arteries are a critical part of the vascular system and provide nourishment to the heart. In humans, even minor defects in coronary arteries can be lethal, emphasizing their importance for survival. However, some teleosts survive without coronary arteries, suggesting that there may have been some evolutionary changes in the morphology and function of coronary arteries in the tetrapod lineage. Here, we propose that the true ventricular coronary arteries were newly established during amniote evolution through remodeling of the ancestral coronary vasculature. In mouse (Mus musculus) and Japanese quail (Coturnix japonica) embryos, the coronary arteries unique to amniotes are established by the reconstitution of transient vascular plexuses: aortic subepicardial vessels (ASVs) in the outflow tract and the primitive coronary plexus on the ventricle. In contrast, amphibians (Hyla japonica, Lithobates catesbeianus, Xenopus laevis, and Cynops pyrrhogaster) retain the ASV-like vasculature as truncal coronary arteries throughout their lives and have no primitive coronary plexus. The anatomy and development of zebrafish (Danio rerio) and chondrichthyans suggest that their hypobranchial arteries are ASV-like structures serving as the root of the coronary vasculature throughout their lives. Thus, the ventricular coronary artery of adult amniotes is a novel structure that has acquired a new remodeling process, while the ASVs, which occur transiently during embryonic development, are remnants of the ancestral coronary vessels. This evolutionary change may be related to the modification of branchial arteries, indicating considerable morphological changes underlying the physiological transition during amniote evolution.

Coronary arteries are tasked with supplying the heart with oxygenated blood and nutrients. Any blockage or developmental problem in these blood vessels can have severe and sometimes lethal consequences. Due to their importance for health, researchers have extensively studied how coronary arteries form in humans and mice; a more limited range of studies have also looked at their equivalent in zebrafish. However, little is known about these structures develop in animals such as birds, amphibians, or other groups of fish. This makes it difficult to retrace the evolutionary processes that have given rise to the coronary arteries we are familiar with in mammals. To address this knowledge gap, Mizukami et al. set out to compare blood vessel development around the heart of mammals, birds, amphibians, and fish. To do this, they performed detailed anatomical studies of blood vessel structure at different stages of development in mice as well as quail, frogs and newts, zebrafish and sharks. In both mice and quail, small arterial subepicardial vessels (or ASVs) emerged early in development around the heart; these subsequently reorganised and remodelled themselves to give rise to the 'true' coronary arteries characteristic of the mature heart. Frogs and newts also developed similar ASV-like structures; however, unlike their mammalian and bird equivalents, these vessels did not reorganise, instead being retained into adulthood. In fish, blood vessel development resembled that of amphibians, suggesting that the coronary artery-like structures seen in some fish are an 'ancestral' form of ASVs, rather than the equivalent of the mature coronary arteries in mammals and birds. This work sheds light on the evolutionary processes shaping essential structures in the heart. In the future, Mizukami et al. hope that this knowledge will help develop a greater range of experimental animal models for studying heart disease and potential treatments.

Three-dimensional morphological analysis of the human spleen and its surrounding organs during the early fetal period.

The spleen has variations in its morphology and is considered to acquire a defined shape in the third month of gestation. However, few studies have investigated spleen development during the first 3 months of fetal life. This study aimed to determine the three-dimensional (3D) morphogenesis of the spleen during the third month of gestation. In this study, 30 fetal specimens (crown-rump length [CRL]: 22-103 mm) were subjected to magnetic resonance imaging analysis. We manually segmented the spleen, stomach, and adrenal gland, reconstructed 3D models, and analyzed the volume and shape of these organs. The results showed that the variation in spleen size was large compared to that in other organs. Spleen morphology was classified into six types based on the number of splenic surfaces as follows: two-faced, three-faced, four-faced, five-faced, ovoid, and irregular. Two-faced spleens were only observed in small specimens, whereas three- and four-faced spleens were observed in larger specimens. We also revealed that the number of fetal splenic surfaces increased as CRL enlarged. Additionally, 3D models indicated that some specimens formed their splenic surfaces without contact with the adjacent organs. This suggested that the splenic surface may be caused not only by pressure from the faced organs but also by an intrinsic program. This study may provide a better understanding of the normal development of the spleen during the early fetal period, and may potentially assist future studies in investigating congenital morphological anomalies of the spleen.

Kyoto Collection in The Anatomical Record.

Human embryology began at the end of the 19th century and has developed using valuable human embryo specimens; the Carnegie and Blechshmidt Collections are famous examples. Although established after these two collections, the Kyoto Collection of Human Embryos and Fetuses has become the largest collection worldwide, and its major asset is 1044 serial tissue sections comprising 547 normal and 497 abnormal cases. Morphological changes have been the focus of analysis owing to the absence of fresh embryos in the Kyoto Collection. Furthermore, analysis methods have undergone significant changes. For example, morphometrics has been used to quantitatively analyze shape changes, although it may result in the loss of information on shape changes, which can hinder the visualization of analysis results. However, geometric morphometrics has been recently introduced to the fetal and embryonic stages to circumvent this problem. With the development of DNA analysis kits, several hundred DNA base pairs have been extracted from the Kyoto Collection of studies conducted from the 2000s to the 2010s through genetic analysis. Future technological advances are eagerly awaited.

Femoral posture during embryonic and early fetal development: An analysis using landmarks on the cartilaginous skeletons of ex vivo human specimens.

The pre-axial border medially moves between the fetal and early postnatal periods, and the foot sole can be placed on the ground. Nonetheless, the precise timeline when this posture is achieved remains poorly understood. The hip joint is the most freely movable joint in the lower limbs and largely determines the lower-limb posture. The present study aimed to establish a timeline of lower-limb development using a precise measurement of femoral posture. Magnetic resonance images of 157 human embryonic samples (Carnegie stages [CS] 19-23) and 18 fetal samples (crown rump length: 37.2-225 mm) from the Kyoto Collection were obtained. Three-dimensional coordinates of eight selected landmarks in the lower limbs and pelvis were used to calculate the femoral posture. Hip flexion was approximately 14° at CS19 and gradually increased to approximately 65° at CS23; the flexion angle ranged from 90° to 120° during the fetal period. Hip joint abduction was approximately 78° at CS19 and gradually decreased to approximately 27° at CS23; the average angle was approximately 13° during the fetal period. Lateral rotation was greater than 90° at CS19 and CS21 and decreased to approximately 65° at CS23; the average angle was approximately 43° during the fetal period. During the embryonic period, three posture parameters (namely, flexion, abduction, and lateral rotation of the hip) were linearly correlated with each other, suggesting that the femoral posture at each stage was three-dimensionally constant and exhibited gradual and smooth change according to growth. During the fetal period, these parameters varied among individuals, with no obvious trend. Our study has merits in that lengths and angles were measured on anatomical landmarks of the skeletal system. Our obtained data may contribute to understanding development from anatomical aspects and provide valuable insights for clinical application.

Three-dimensional morphogenesis of the human diaphragm during the late embryonic and early fetal period: Analysis using T1-weighted and diffusion tensor imaging.

A precise understanding of human diaphragm development is essential in fetal medicine. To our knowledge, no previous study has attempted a three-dimensional (3-D) analysis and evaluation of diaphragmatic morphogenesis and development from the embryonic to the early fetal period. This study aimed to evaluate the morphogenesis and fibrous architecture of the developing human diaphragm during the late embryonic and early fetal periods. Fifty-seven human embryos and fetuses (crown-rump length [CRL] = 8-88 mm) preserved at the Congenital Anomaly Research Center of Kyoto University and Shimane University were analyzed. 3-D morphogenesis and fiber orientation of the diaphragm were assessed using phase-contrast X-ray computed tomography, T1-weighted magnetic resonance imaging (T1W MRI), and diffusion tensor imaging (DTI). T1W MR images and DTI scans were obtained using a 7-T MR system. The diaphragm was completely closed at Carnegie stage (CS) 20 and gradually developed a dome-like shape. The diaphragm was already in contact with the heart and liver ventrally in the earliest CS16 specimen observed, and the adrenal glands dorsally at CS19 or later. In the fetal period, the diaphragm contacted the gastric fundus in samples with a CRL ≥41 mm, and the spleen in samples with a CRL ≥70 mm. The relative position of the diaphragm with reference to the vertebrae changed rapidly from CS16 to CS19. The most cranial point of the diaphragm was located between the 4th and 8th thoracic vertebrae, regardless of fetal growth, in samples with a CRL of ≥16 mm. Diaphragmatic thickness was nearly uniform (0.15-0.2 mm) across samples with a CRL of 8-41 mm. The sternal, costal, lumbar parts, and the area surrounding the esophageal hiatus thickened with growth in samples with a CRL of ≥46 mm. The thickness at the center of the diaphragm and the left and right hemidiaphragmatic domes did not increase with growth. Tractography showed that the fiber orientation of the sternal, costal, and lumbar parts became more distinct as growth progressed in CS19 or later. All fibers in the costal and lumbar regions ran toward the left and right hemidiaphragmatic domes, except for those running to the caval opening and esophageal hiatus. The fiber orientation patterns from the right and left crura surrounding the esophageal hiatus were classified into three types. Distinct fiber directions between the costal and sternal and between the costal and lumbar diaphragmatic parts were observable in samples with a CRL of ≥46 mm. Anterior costal and sternal fibers ran toward the center. Fiber tracts around the center and the left and right hemidiaphragmatic domes; between the costal and lumbar orientations; and between the costal and sternal orientations showed a tendency for decreasing fractional anisotropy values with fetal growth and showed less density than other areas. In conclusion, we used 3-D thickness assessment and DTI tractography to identify qualitative changes in the muscular and tendonous regions of the diaphragm during the embryonic and early fetal periods. This study provides information on normal human diaphragm development for the progression of fetal medicine and furthering the understanding of congenital anomalies.

Morphogenetic progression of thigh and lower leg muscles during human embryonic development.

Fetal musculoskeletal movements are first observed at approximately seven to 8 weeks of gestation. However, the separation and formation of skeletal muscles, especially the limbs, have not yet been described in detail. In this study, we elucidate the sequence of events leading to the formation of each thigh and lower leg muscle using serial sections. To observe muscle formation, 26 serial sections (50 legs) of human embryonic specimens ranging from Carnegie stages (CS) 19 to 23 were selected from the Kyoto collection stored at the Congenital Anomaly Research Center, Kyoto University Graduate School of Medicine. As a result, we show the detailed formation and separation processes of the thigh and lower leg muscles. In the thigh, sartorius and tensor fasciae latae are separated at CS19, and the individual muscles observed in adults are identified after CS21. In the lower leg, the tibialis anterior exhibits early separation at CS20, and all muscles are identified at CS22. This study enables future research into the relationship between embryonic development and the evolution of muscle action from quadrupedal to erect bipedal walking.

Tentorium cerebelli formation during human embryonic and early fetal development.

The morphologies of the fetal tentorium cerebelli (TC) and brain influence each other during development. This study aimed to analyze and more comprehensively understand the three-dimensional morphogenesis of the TC and fetal brain. We examined magnetic resonance imaging from 64 embryonic and fetal specimens (crown-rump length range, 9.2-225 mm). During the embryonic period, the lateral folds of the TC elongated to traverse the middle part of the midbrain. The TC and falx cerebri appeared separated, and no invaginations at the parieto-occipital region were observed. In the early fetal period, the cerebrum covered approximately half of the midbrain. The separation of the dural limiting layer at the parieto-occipital region widened from the posterior cerebrum to the cranial cerebellum. The lateral folds of the TC were spread between its tip, continuous with the falx cerebri, and its base plane, located between the midbrain and rostral hindbrain. Differences in the TC components' growth directions gradually diminished as the cerebrum covered the midbrain. We observed rotation of the TC at its median section according to its growth, which ceased in the middle fetal period. The brainstem and cerebellum extended inferiorly via differential growth, with the cerebrum covering them superiorly. The morphology of the TC curved to conform to the cerebellar and cerebral surfaces. Our present study suggests that factors affecting TC morphology differ between the early and middle fetal periods. Present data provided a more comprehensive view of TC formation according to developmental stage.

Rhythmic auditory stimulation during gait adaptation enhances learning aftereffects and savings by reducing common neural drives to lower limb muscles.

Rhythmic auditory stimulation (RAS) improves gait symmetry in neurological patients with asymmetric gait patterns. However, whether RAS can accelerate gait adaptation remains unclear. This study aimed to investigate whether RAS during gait adaptation can enhance learning aftereffects and savings of gait symmetries. Furthermore, we investigated the differences in the coherence of paired surface electromyographic (EMG) recordings during gait adaptation between with and without RAS. Nineteen healthy young adults were subjected to continuous treadmill gait with swing phase perturbation (adaptation period) with or without RAS (RAS or no-RAS condition) for 5 or 10 min (short- or long-time condition), without the perturbation for 5 min (de-adaptation period), and with the perturbation for another 5 min (re-adaptation period). Swing phase and step length symmetries were significantly greater in the RAS conditions than in the no-RAS conditions during the adaptation period. Learning aftereffects and savings of gait symmetries were significantly greater in the RAS conditions than in the no-RAS conditions in the early de-adaptation and re-adaptation periods, respectively. There were no significant differences in savings in the early re-adaptation period between the short- and long-time conditions in the RAS condition. EMG-EMG coherence in the rectus femoris muscle in the ß band (15-35 Hz) on the perturbed side was significantly lower during the early adaptation period in the RAS than in the no-RAS conditions. Therefore, RAS may enhance learning efficiency by reducing common neural drives from a cortical structure during gait adaptation, which could induce high savings of a learned gait pattern, even within short-time periods.NEW & NOTEWORTHY RAS during gait adaptation against swing phase perturbation enhances learning aftereffects and savings of gait symmetries. EMG-EMG coherence in the rectus femoris muscle in the ß band on the perturbed side during the swing phase was significantly lower in the RAS than in the no-RAS conditions during the early adaptation period. These results support the application of RAS as external feedback to improve gait symmetry during gait adaptation in the rehabilitation of neurological patients.

Mesothelial fusion mediates chorioallantoic membrane formation.

In amniotic vertebrates (birds, reptiles and mammals), an extraembryonic structure called the chorioallantoic membrane (CAM) functions as respiratory organ for embryonic development. The CAM is derived from fusion between two pre-existing membranes, the allantois, a hindgut diverticulum and a reservoir for metabolic waste, and the chorion which marks the embryo's external boundary. Modified CAM in eutherian mammals, including humans, gives rise to chorioallantoic placenta. Despite its importance, little is known about cellular and molecular mechanisms mediating CAM formation and maturation. In this work, using the avian model, we focused on the early phase of CAM morphogenesis when the allantois and chorion meet and initiate fusion. We report here that chicken chorioallantoic fusion takes place when the allantois reaches the size of 2.5-3.0 mm in diameter and in about 6 hours between E3.75 and E4. Electron microscopy and immunofluorescence analyses suggested that before fusion, in both the allantois and chorion, an epithelial-shaped mesothelial layer is present, which dissolves after fusion, presumably by undergoing epithelial-mesenchymal transition. The fusion process per se, however, is independent of allantoic growth, circulation, or its connection to the developing mesonephros. Mesoderm cells derived from the allantois and chorion can intermingle post-fusion, and chorionic ectoderm cells exhibit a specialized sub-apical intercellular interface, possibly to facilitate infiltration of allantois-derived vascular progenitors into the chorionic ectoderm territory for optimal oxygen transport. Finally, we investigated chorioallantoic fusion-like process in primates, with limited numbers of archived human and fresh macaque samples. We summarize the similarities and differences of CAM formation among different amniote groups and propose that mesothelial epithelial-mesenchymal transition mediates chorioallantoic fusion in most amniotic vertebrates. Further study is needed to clarify tissue morphogenesis leading to chorioallantoic fusion in primates. Elucidating molecular mechanisms regulating mesothelial integrity and epithelial-mesenchymal transition will also help understand mesothelial diseases in the adult, including mesothelioma, ovarian cancer and fibrosis. This article is part of the theme issue 'Extraembryonic tissues: exploring concepts, definitions and functions across the animal kingdom'.