Cervical Flexure Research Articles

Research Note: Injection of adenoviral CRISPR/Cas9 system targeting melanophilin gene into different sites of embryos induced regional feather color changes in posthatch quail

Poultry species is an important animal model in both avian research and the poultry industry. To advance our understanding of genetic factors and benefit both fields, a gene of interest can be genetically edited, and consequential phenotypic changes can be investigated. Injection of adenovirus containing the CRISPR/Cas9 system into avian blastoderm induced genome editing in blastodermal cells randomly, including primordial germ cells, which results in generation of whole-body knockout in the offspring of the virus-injected quail. However, to observe phenotypic and functional changes in whole-body, homozygous knockout of genes using this genome editing technology requires at least 2 generations of breeding of chimeric, and heterozygotes birds. In the current study, we developed a strategy to investigate the gene function in 1-generation by inducing regional genome editing around the injection sites with CRISPR/Cas9 adenovirus. The adenoviral CRISPR/Cas9 vector targeting the melanophilin (Mlph) gene, regulating feather pigmentation, was injected into 2 different regions of embryos, the cervical flexure of quail embryos at HH stage 13 to 15 and the tip of the upper limb bud of embryos at HH stage 22 to 24, to induce genome editing in those regions. Indel mutations in the target loci of the Mlph gene were detected by extracting genomic DNA from the embryonic tissues, and consequential phenotypes, feather color changes, were analyzed at 1 mo after hatch. Injection of the adenovirus into the cervical flexure and the tip of the upper limb bud of embryos resulted in 8 to 21% efficiency of indel mutation in the embryonic cells of the injected regions. In the posthatch quail, gray feathers were shown on their upper back and primary wing feathers, corresponding to the injection sites at embryos. Successful validation of this strategy for inducing genome editing in parts of tissues within 1-generation will accelerate studies on genetic functions with advantages of less time and cost, facilitating avian research and providing foundations for future application for the poultry industry.

Does Cell Proliferation Affect Embryonic Flexure and Torsion?

Two vital and simultaneous morphogenetic events that occur during early embryo development are the flexure and torsion (F&T) of the embryonic body axis. Both contribute to development of the fetal position, which can be observed in avian and mammalian embryos. Correct progression of F&T is crucial for the proper development of the heart and other organs. Embryonic flexure can be divided into three segments: cranial, cervical, and thoracic. In this study, we focus on cervical flexure and torsion of the embryonic body axis. Differential cell proliferation, the difference in the rate of division of cells between regions in the developing embryo, is one mechanism that could influence F&T. Goodrum & Jacobson (1981) observed thickening of the dorsal midbrain relative to the ventral midbrain during cranial flexure. Miller & White (1998) have shown using mouse embryos that differential growth could cause torsion of an axis confined to an attached sphere.We performed studies inhibiting cell proliferation. Fertile white leghorn chick eggs were incubated to approximately Hamburger and Hamilton (HH) Stage 12 or 13. At these stages, F&T have just begun. We then added aphidicolin, a mitotic inhibitor, to whole embryo culture. Embryos were observed at 0, 5, and 10 hours to see if F&T were repressed. We hypothesized that the addition of aphidicolin would inhibit progression of F&T. We developed quantitative schemes to score the extent of F&T progression from HH 12 to HH 15. Embryos were scored from 0 to 4 based on the degree of cervical flexure and were also scored from 0 to 4 based on the degree of torsion. Afterward, the difference in F&T scores of same embryos at time t=0 h and t=10 h was calculated to determine the extent of both. These scores were then averaged for multiple embryos.Compared with controls (n=46), embryos incubated in 400 µM aphidicolin (n=18) flexed and rotated at a significantly lower rate. For flexion, the mean difference in scores for control and treated embryos were 1.85 and 1.22, respectively. For torsion, the mean difference in scores for control and treated embryos were 1.62 and 1.25, respectively. These differences were statistically significant. It is noted that global application of aphidicolin did impact the overall health of embryos. However, our preliminary results indicate that differential cell proliferation could play a role in rotation and flexure progression in the developing chicken embryo.

On the Biomechanics of Cardiac S-Looping in the Chick: Insights From Modeling and Perturbation Studies.

Cardiac looping is an important embryonic developmental stage where the primitive heart tube (HT) twists into a configuration that more closely resembles the mature heart. Improper looping leads to congenital defects. Using the chick embryo as the experimental model, we study cardiac s-looping wherein the primitive ventricle, which lay superior to the atrium, now assumes its definitive position inferior to it. This process results in a heart loop that is no longer planar with the inflow and outflow tracts now lying in adjacent planes. We investigate the biomechanics of s-looping and use modeling to understand the nonlinear and time-variant morphogenetic shape changes. We developed physical and finite element models and validated the models using perturbation studies. The results from experiments and models show how force actuators such as bending of the embryonic dorsal wall (cervical flexure), rotation around the body axis (embryo torsion), and HT growth interact to produce the heart loop. Using model-based and experimental data, we present an improved hypothesis for early cardiac s-looping.

Craniometric Analysis of the Hindbrain and Craniocervical Junction of Chihuahua, Affenpinscher and Cavalier King Charles Spaniel Dogs With and Without Syringomyelia Secondary to Chiari-Like Malformation.

ObjectivesTo characterize and compare the phenotypic variables of the hindbrain and craniocervical junction associated with syringomyelia (SM) in the Chihuahua, Affenpinscher and Cavalier King Charles Spaniel (CKCS).MethodAnalysis of 273 T1-weighted mid-sagittal DICOM sequences of the hindbrain and craniocervical junction from 99 Chihuahuas, 42 Affenpinschers and 132 CKCSs. The study compared 22 morphometric features (11 lines, eight angles and three ratios) of dogs with and without SM using refined techniques based on previous studies of the Griffon Bruxellois (GB) using Discriminant Function Analysis and ANOVA with post-hoc corrections.ResultsThe analysis identified 14/22 significant traits for SM in the three dog breeds, five of which were identical to those reported for the GB and suggest inclusion of a common aetiology. One ratio, caudal fossa height to the length of the skull base extended to an imaginary point of alignment between the atlas and supraoccipital bones, was common to all three breeds (p values 0.029 to <0.001). Associated with SM were a reduced occipital crest and two acute changes in angulation i) ‘sphenoid flexure’ at the spheno-occipital synchondrosis ii) ‘cervical flexure’ at the foramen magnum allied with medulla oblongata elevation. Comparing dogs with and without SM, each breed had a unique trait: Chihuahua had a smaller angle between the dens, atlas and basioccipital bone (p value < 0.001); Affenpinschers had a smaller distance from atlas to dens (p value 0.009); CKCS had a shorter distance between the spheno-occipital synchondrosis and atlas (p value 0.007).ConclusionThe selected morphometries successfully characterised conformational changes in the brain and craniocervical junction that might form the basis of a diagnostic tool for all breeds. The severity of SM involved a spectrum of abnormalities, incurred by changes in both angulation and size that could alter neural parenchyma compliance and/or impede cerebrospinal fluid channels.

On the role of intrinsic and extrinsic forces in early cardiac S‐looping

Looping is a crucial phase during heart development when the initially straight heart tube is transformed into a shape that more closely resembles the mature heart. Although the genetic and biochemical pathways of cardiac looping have been well studied, the biophysical mechanisms that actually effect the looping process remain poorly understood. Using a combined experimental (chick embryo) and computational (finite element modeling) approach, we study the forces driving early s-looping when the primitive ventricle moves to its definitive position inferior to the common atrium. New results from our study indicate that the primitive heart has no intrinsic ability to form an s-loop and that extrinsic forces are necessary to effect early s-looping. They support previous studies that established an important role for cervical flexure in causing early cardiac s-looping. Our results also show that forces applied by the splanchnopleure cannot be ignored during early s-looping and shed light on the role of cardiac jelly. Using available experimental data and computer modeling, we successfully developed and tested a hypothesis for the force mechanisms driving s-loop formation. Forces external to the primitive heart tube are necessary in the later stages of cardiac looping. Experimental and model results support our proposed hypothesis for forces driving early s-looping.

Asymmetry of Spinal Segments Mobility in Canoeists and its Relationship with Racing Speed

The aim of this study was to determine the extent of asymmetry of spinal segment mobility in canoeists. Moreover, the relationship between this parameter and racing speed was analyzed. The study included 18 canoeists with a mean age of 16.4 years. Mobility of cervical, thoracic and lumbar spine, in sagittal, coronal and transverse planes, was measured with the aid of a tensometric electrogoniometer. The racing speed was based on results achieved during the qualifying competition for the Polish national team. Spinal mobility was measured within two days after the competition. Significant associations were observed between average racing speed and the asymmetry coefficients of the cervical (r=−0.52; p=0.03) and lumbar spinal flexure in the coronal plane (r=0.57; p=0.01). The extent of the asymmetry of the cervical spine flexure in the coronal plane should possibly be reduced, because such asymmetry exerts a negative effect on racing speed. In contrast, canoeist’s training should be oriented towards increasing the asymmetry of the lumbar spine flexure in the coronal plane. However, one should keep in mind that such an approach, although favorable in terms of race performance, could negatively affect the canoeist’s health.

Estudo microscópico e macroscópico, com enfoque radiográfico e de alizarina, no desenvolvimento embrionário e fetal de gatos domésticos (Felis catus) em diferentes idades gestacionais

O gato doméstico (Felis catus) foi nomeado por Carolus Linnaeus em seu livro Systema Naturae, em 1798. A família Felidea apresenta muita semelhança morfológica com os felinos selvagens. O estudo da embriologia do gato doméstico é de grande valia, uma vez que, é considerado um importante modelo animal quando comparado aos gatos selvagem em extinção, especialmente relacionado às pesquisas sobre biologia reprodutiva. Este trabalho objetivou análisar e comparar as fases embrionárias de quatro embriões e um feto de felinos domésticos. Nos embriões com idade gestacional estimada em 17 dias (0,5cm CR) podemos observar pela análise macroscópica a presença de dilatação rostral correspondente ao prosencéfalo, o local placóide do cristalino, a flexura cervical, os quatro arcos faríngeos com os sulcos que o dividem, a proeminência cardíaca, o indício do brotamento do membro pélvico, além da presença de somitos. Na região caudal do embrião, visualizamos a curvatura cranio-caudal, permitindo ao mesmo uma posição em formato de "C". Nos embriões com idade gestacional estimada em 22 dias (1,2cm CR), na análise macroscópica foi visualizado o prosencéfalo, vesícula óptica com pigmentação da retina, vesícula ótica, quarto ventrículo, fígado, membros torácicos e pélvicos com discreta distinção dos dígitos e vascularização superficial. Nos embriões com idade gestacional estimada em 25 dias (1,5cm CR) notamos a presença do prosencéfalo e mesencéfalo, a curvatura cervical pronunciada, vesícula óptica com forte pigmentação da retina, vesícula ótica, membros pélvicos e torácicos bem desenvolvidos, com distinção dos dígitos e fígado bem pronunciado. Os fetos com idade gestacional estimada em 52 dias (10cm CR) possuem estruturas internas e externas facilmente identificadas em animais adultos. Com relação às estruturas ósseas notamos que as mesmas não apresentam nenhuma epífise óssea formada, sendo visíveis somente as diáfises ósseas. Na análise microscópica, o embrião de idade gestacional de 19 dias (0,9cm CR) revelou a presença do rostro, cavidade oral com lábio superior e inferior, cavidade nasal, olho e a abertura do 4º ventrículo encefálico, esôfago, coração com átrio e ventrículo, pulmão, fígado, crista mesonéfrica, gônada primitiva, estômago, broto do membro torácico, coluna vertebral e a medula espinhal em formação. Esse trabalho é de grande importância para o estudo da morfologia externa e interna de gatos domésticos, principalmente no que diz respeito ao desenvolvimento ósseo e articular, considerando as alterações que podem ou não ser promovidas pelo uso de terapias medicamentosas ou celulares durante o desenvolvimento embrionário e fetal.

Embriologia e genetica dello sviluppo cerebellare

During the fourth week of development, the mesencephalic flexure and cervical flexure appear in the cranial region of the neural tube, delimiting three neural vescicles: the prosencephalon, mesencephalon and rhomboencephalon. During the fifth week, the pontine flexure forms in the roof of the rhomboencephalon, marking the division between the metencephalon and myeloencephalon, the future medulla oblongata. The dorsal part of the metencephalon (alar plate), between the mesencaphalic isthmus and the pontine flexure, will give rise to the cerebellum, whereas its ventral part (basal plate) will give rise to the pons. The alar plates, lateral to the deepening pontine fissure and the hindbrain cavity, present an intense proliferative activity with the formation of two extensions which fuse just behind the isthmus to form the cerebellar plate. During the sixth week of development, the rhombic lips appear in the posterolateral regions of this strip and after persistent proliferative activity they fuse along the median line. Median thickening of this region will give rise to the vermis, while the two lateral masses become the lateral lobes. During the seventh week, the flocculonodular lobe will form. Between the eighth and ninth weeks, the vermis expands caudally with progressive dilatation of the fourth ventricle whose Luschka and Magendie foramina are still unperforated. The cerebellar proliferation and differentiation processes are correlated to the activity of two separate germinal areas: the ependymal ventricle and the rhombic lip. From the eighth week, neuroblastic migration starts from the rhombic lip which in successive genetically controlled steps leads to the formation of the external granular layer of the cerebellar cortex. From the ventriculo-ependymal zone cells migrate to form the cerebellar nuclei destined to differentiate into Purkinje cells. The cerebellar recess starts to develop around the twelfth week with the formation of the primary fissure separating the anterior and posterior parts of the cerebellum to end with the complete formation of all cerebellar lobes at around the twenty-fourth week of development. The foliation process starts when the recess has formed and continues in the first months of life after birth. The processes of cell development and differentiation are genetically programmed and the outcome of an interaction between genetic and environmental factors. The different stages of embryonic development are thought to be controlled by sequential ordered activation of genetic clusters (homeotypic genes) which in turn encode for a series of molecules directly involved in the regulation of cell and tissue interactions. There is increasing evidence of genetic involvement in the different types of cerebellar malformation whose expression and association with other extracerebellar malformations depends on the developmental age at which the genetic change occurred.

Magnetic resonance imaging and three-dimensional reconstructions of the brain of a fetal common dolphin, Delphinus delphis.

To demonstrate the kinds of data that can be obtained non-destructively and non-invasively from preserved museum specimens using modern imaging technology the head region of a whole body fetal specimen of the common dolphin, Delphinus delphis, aged 8-9 months post-conception, was scanned using Magnetic Resonance Imaging (MRI). Series of scans were obtained in coronal, sagittal and horizontal planes. A digital three-dimensional reconstruction of the whole brain was prepared from the coronal series of scans. Sectional areas and three-dimensional volumes were obtained of the cerebral hemispheres and of the brainstem-plus-cerebellum. Neuroanatomical features identified in the scans include the major sulci of the cerebral hemispheres, well-differentiated regions of gray and white matter, the mesencephalic, pontine, and cervical flexures, the "foreshortened" appearance of the forebrain, and the large auditory inferior colliculi. These findings show that numerous features of the fetal common dolphin brain can be visualized and analyzed from MRI scans.

Interruption of cardiac output does not affect short-term growth and metabolic rate in day 3 and 4 chick embryos.

The heart beat of vertebrate embryos has been assumed to begin when convective bulk transport by blood takes over from transport by simple diffusion. To test this hypothesis, we measured eye growth, cervical flexure and rates of oxygen consumption ( V(O2)) in day 3-4 chick embryos denied cardiac output by ligation of the outflow tract and compared them with those of embryos with an intact cardiovascular system. Eye diameter, used as the index for embryonic growth, increased at a rate of approximately 4.5-5 % h(-)(1) during the observation period. There was no significant difference (P>0.1) in the rate of increase in eye diameter between control (egg opened), sham-ligated (ligature present but not tied) and ligated embryos. Similarly, the normal progression of cervical flexure was not significantly altered by ligation (P>0.1). V(O2) (ml O(2 )g(-)(1 )h(-)(1)) at 38 degrees C, measured by closed respirometry, was not significantly different (P>0.1) on day 3 in sham-ligated (14.5+/-1.9 ml O(2 )g(-)(1 )h(-)(1)) and ligated 17.6+/-1.8 ml O(2 )g(-)(1 )h(-)(1)) embryos. Similarly, on day 4, V(O2) in sham-ligated and ligated embryos was statistically the same (sham-ligated 10. 5+/-2.9 ml O(2 )g(-)(1 )h(-)(1); ligated 9.7+/-2.9 ml O(2 )g(-)(1 )h(-)(1)). Expressed as a linear function of body mass (M), V(O2) in sham-ligated embryos was described by the equation V(O2)=-0.48M+24.06 (r(2)=0.36, N=18, P<0.01), while V(O2) in ligated embryos was described by the equation V(O2)=-0.53M+23.32 (r(2)=0.38, N=16, P<0.01). The regression line describing the relationship between body mass and V(O2) for pooled sham-ligated and ligated embryos (the two populations being statistically identical) was V(O2)=-0.47M+23.24. The slope of this regression line, which was significantly different from zero (r(2)=0.30, N=34, P<0.01), was similar to slopes calculated from previous studies over the same range of body mass.Collectively, these data indicate that growth and V(O2) are not dependent upon cardiac output and the convective blood flow it generates. Thus, early chick embryos join those of the zebrafish, clawed frog and axolotl in developing a heart beat and blood flow hours or days before required for convective oxygen and nutrient transport. We speculate that angiogenesis is the most likely role for the early development of a heart beat in vertebrate embryos.

Unilateral vitelline vein ligation alters intracardiac blood flow patterns and morphogenesis in the chick embryo.

To study the role of blood flow in normal and abnormal heart development, an embryonic chicken model was developed. The effect of altered venous inflow on normal intracardiac blood flow patterns was studied by visualization of blood flow with India ink. At stage 17, India ink was injected into a capillary or small venule within a specific yolk sac region. After determination of the normal intracardiac flow pattern, the right lateral vitelline vein was ligated, and the new intracardiac flow pattern was studied. Ligation resulted in disturbance of normal intracardiac flow patterns, which was most obvious in the conotruncus. The long-term effect of these abnormal intracardiac flow patterns on the development of the heart and pharyngeal arch arteries was investigated by permanent ligation in ovo with a microclip at stage 17 and subsequent evaluation at stages 34, 37, and 45. These experiments revealed anomalies of the vascular system in 58 of the 91 ligated embryos studied. We observed intracardiac malformations consisting of subaortic ventricular septal defects (n = 52), semilunar valve anomalies (n = 19), atrioventricular anomalies (n = 7), and pharyngeal arch artery malformations (n = 32). It is concluded that abnormal intracardiac blood flow, resulting from hampered venous inflow, may result in serious intracardiac and pharyngeal arch artery malformations comparable to defects observed in embryonic chicken models subjected to neural crest ablation, cervical flexure experiments, and excessive retinoic acid treatment.

Morphological characteristics of the developing human brain during the embryonic period.

Many features of the developing nervous system are visible from external observations of intact human embryos. In this study, a photographic atlas from the 4th to the 7th week after ovulation (Carnegie stages 10-18) is provided. The neural folds began to fuse at stage 10, and the rostral and caudal neuropore were closed during stages 11 and 12, respectively. The three primary divisions of the brain were distinguishable before closing of the neural tube. The five secondary brain vesicles were formed during stages 14-15. The development of the cerebellum and cerebrum were first observed at stages 14 and 15, respectively. The mesencephalic flexure was seen at stage 12, and the cervical flexure and pontine flexure at stage 14. After stages 18-19, it became increasingly difficult to identify detailed features of the brain from the surface. Results from this study will help to correlate the characteristic findings of the developing central nervous system of human embryos from stereomicroscopical and light microscopical observations and to locate the exact parts of the developing human brain for other purposes.

Formation of the Cervical Flexure: An Experimental Study on Chick Embryos

It has been proposed that the cervical flexure of vertebrate embryos arises from the normal morphogenesis of the heart. This hypothesis is based on experiments in which the heart tube is removed or disrupted in early chick embryos. It has been reported that, in normal atmosphere, these embryos continued normal morphogenesis except for cervical flexure formation. In the present study, we performed similar experiments. In contrast to previous work, however, only one set of our heart-deprived chick embryos was reincubated in normal air. The other sets were reincubated in oxygen-enriched air. Under normoxia, heart removal resulted not only in prevention of the cervical flexure, but also in mesenchymal defects, and in a remarkable hypoplasia of the craniocervical region. Under hyperoxia, heart-deprived embryos developed no severe mesenchymal defects and the growth of the upper body portion was more normal, with the hypoplasia confined to the cranial region. The formation of the cervical flexure was now normalized. These results show that cervical flexure formation is not directly dependent on normal morphogenesis of the heart, but does depend on a sufficient oxygen supply to the cervical region. During early development, the cranio-cervical region of a chick embryo is more sensitive to circulatory failure than the trunk.

Correlation between the embryonic head flexures and cardiac development. An experimental study in chick embryos.

The aim of the present study is to examine whether the formation of the cranial and cervical flexures is involved in the process of cardiac looping, and whether looping anomalies are causally involved in the development of cardiac malformations. For this purpose, the formation of the cranial and cervical flexures was experimentally suppressed in chick embryos by introducing a straight human hair into the neural tube. In the experimental embryos, the absence of the cervical flexure, alone or in combination with a reduced cranial flexure, was always associated with anomalies in the looping of the tubular heart. The convergence of the primary distant venous and arterial ends of the heart, as well as the normal movement of the ventricular region from its original position, cranial and ventral from the cardiac inflow, to its final position caudal to the presumptive atria, was suppressed to an extent related to the degree to which the formation of the flexures was prevented. Positional immaturity of the heart loop (increased distance between its inflow and outflow, and cranio-ventral position of the ventricular region) was associated with incomplete deformations (reduced angulations) of the cardiac wall at the atrioventricular or conoventricular junctional areas. Reduced angulations were associated with the hypoplasia of the anlagen of the cardiac septa at the level of the angulation (av-cushions, conal ridges). Hypoplasia of these anlagen was followed by incomplete or absent fusion of their opposite free edges, which finally resulted in atrioventricular or ventricular septal defects. These results show that the convergence of the venous and arterial ends of the tubular heart and the caudo-dorsal movement of its ventricular region are related to the formation of the cervical flexure, and that the mesenchymal septa of the heart seem to develop in response to deformations of the embryonic heart, which are generated by the process of cardiac looping. Therefore, the positional and morphological changes of the looping heart are regarded as playing a key role in the process of normal and abnormal morphogenesis of the heart.

MR imaging of the developing human brain. Part 1. Prenatal development.

To establish a baseline of the magnetic resonance (MR) imaging appearance of the fetal brain in early stages of development, the authors undertook a study of fixed and fresh specimens of embryos and fetuses of 6-28 weeks gestational age. Images of formalin-preserved and fresh specimens were comparable in their depiction of anatomic structures. On MR images of embryos of 6 weeks gestational age, the rhombic and cervical flexures, aqueduct of Sylvius, diencephalon, cerebellum, cerebral hemisphere, and fourth ventricle could be differentiated. The optic recess and chiasm, pituitary gland, pineal recess, third ventricle, pons, olfactory lobe, corpus striatum, insula, and parietal and temporal lobes could be distinguished as early as 11 weeks gestation. Although MR imaging is impractical as a screening tool for intrauterine abnormalities, it can demonstrate the fetus in great detail and allows a more specific evaluation of fetal anatomy. With the information provided by MR imaging, it may be possible to establish guidelines for assessment of the stage of development during intrauterine life.

The mechanism of cervical flexure formation in the chick

Chick embryos, during stages 14 to 25, undergo an arching of the hindbrain and cervical neural tube that is termed cervical flexure. We have found that if the truncus arteriosus is severed during stage 12-13, the embryos survive for more than 24 h and do not show cervical flexure. The embryos have a beating heart, the expected number of somites, and often have discernible wing and leg buds. Light and electron micrographs reveal no histological abnormalities. The percentage of cells that become labeled with tritiated thymidine is close to normal, indicating that most of the cells are healthy. These results suggest that cervical flexure is related to normal morphogenesis of the heart. At stage 10, the heart is almost straight, with the prospective ventricle cranial to the prospective sinus venosus. The heart tube loops between stage 10 and stage 23, first to the right and then caudad, so that the ventricle becomes caudal to the sinus venosus. The heart undergoes these morphogenetic movements autonomously. The truncus arteriosus does not increase in length during caudal movement of the ventricle, so the cervical region is pulled into an arch. Bending of the cervical region into an arch can be prevented in intact embryos by injecting agar into the foregut, so that the foregut cannot bend. However, after about 24 h of further growth, if the axis cannot bend, the truncus begins to leak blood and the embryo dies. We conclude that cervical flexure is a response of the embryonic axis to the morphogenesis of the heart.

Central nervous system malformations induced by triamcinolone acetonide in nonhuman primates: pathogenesis.

The pathogenetic sequence for TAC-induced encephalocele is in agreement with hypotheses proposing that neural tube closure is followed by protrusion of the mesencephalon, with subsequent growth and development resulting in herniation of the cerebrum and cerebellum. This model could serve to clarify the pathogenesis of encephalocele and to stimulate further study in comparing this defect to other dysraphic states. Triamcinolone acetonide (TAC) was administered intramuscularly (10 mg/kg) to 16 pregnant rhesus monkeys (Macaca mulatta) for 5 alternate days of pregnancy, beginning on gestational day (GD) 23. Conceptuses were removed by hysterotomy at GD 35, 42, 50, or 70 and examined grossly and histologically. Length, area, and perimeter of the tectum and aqueduct area and perimeter were measured with an image analyzer. Changes in treated specimens were suggestive of forces within or ventral to the tectum resulting in dorsal protrusion, rostral-posterior stretching, and attenuation. The angle of the cephalic, pontine, and cervical flexures was also measured. The more acute angle of the cephalic flexure and less acute cervical flexure of treated specimens could represent altered orientation secondary to a mesenchymal deficiency. However, the less acute angle of the pontine flexure in treated specimens suggests an intrinsic alteration in the neural tube. This suggests that encephalocele may result from a combination of mesenchymal and neural tube abnormalities.

A description of caudal migration during growth leading to the formation of the pericardial and pleural coeloms, to caudal movement of the aortic arches, and to development of the shoulder.

Formation of the shoulder in the chick embryo results from movement of cells into the base of the wing from more cranial regions. Between stage 23 and stage 27, the lateral body wall overlying the pleural coelom is caused to move into the base of the wing because the cranial margin of the pleural coelom moves from the level of the tenth somite to the level of the 16th somite. Thus, formation of the shoulder is related to the formation of the pleural coelom. Textbook descriptions of the formation of the pleural coelom did not permit this relationship to be delineated. Therefore, investigation of the relationship between the formation of the shoulder and the formation of the pleural coelom seemed warranted. At stage 12 the coelom is lateral to the gut from the level of the second somite to the level of the 16th somite and is continuous with the extraembryonic coelom. During stage 13 the coelom obtains a ventral margin at the level of the third and fourth somites as the heart fuses with the lateral body wall to form the lateral mesocardium. Between stage 16 and stage 23, the second (cervical) flexure develops, causing the lateral mesocardium to form a ventral margin of the coelom from the level of the first to the 17th somites. Between stage 16 and stage 23, the coelom becomes ventral to the pharynx and lung to the level of the ninth somite. From stage 23 to stage 27, the lateral body wall from the cranial margin of the lung to the caudal margin of the hyoid arch moves in a caudal direction. During stage 27, the lateral body wall fuses with the lateral mesocardium to form the pleuro-pericardial membrane. The wing remains ventral to the 16th to 19th somites, causing a pileup of cells at the cranial base of the wing, the shoulder.

An immunocytochemical study of the development of central serotoninergic neurons in the chick embryo.

The development of central serotoninergic neurons in the chick embryo has been investigated immunocytochemically by utilizing an antiserum to serotonin (5-HT). Immunoreactive neurons are first detected in the brainstem on embryonic day 4 (E4, stage 23), days earlier than 5-HT systems have been detected previously by biochemical techniques. The earliest 5-HT-containing cells at E4 appear rostral to the pontine flexure, yet by E5, 5-HT neuronal groups are observed throughout the brainstem from just caudal to the mesencephalic flexure to the cervical flexure. During this and subsequent phases of development, two distinct patterns of cellular migration seem to be involved in the formation of the various 5-HT neuronal groups. One pattern involves a ventral migration of 5-HT cells, which appears dependent upon the directional guidance of midline radial processes (formed by floor plate cells) that extend across the neuroepithelium. The other pattern involves a lateral migration of cells, followed by an aggregation and rearrangement of 5-HT neurons into distinct subgroups or clusters. Through these patterns of migration most components of the 5-HT neuronal system can be recognized as early as E12, with the mature organization of the 5-HT cell groups occurring by E17. One unexpected finding was the comparatively late appearance (between E9 and E12) of 5-HT neurons in the paraventricular organ of the hypothalamus. Thus, in comparison to the initial observation of the majority of brainstem 5-HT neurons at E4 to E5, the hypothalamic 5-HT cells appear after a delay of between 5 and 7 days. Such differences illustrate the fact that neurons sharing a common neurotransmitter phenotype do not necessarily share the same developmental timetable for the expression of that particular phenotype, or they may undergo neurogenesis during considerably different periods of embryogenesis.

Embryonic head posture and palatal shelf elevation

During embryonic and fetal stages of human development, elongation and straightening of the neck occurs. Authors have suggested that the resulting change in head posture lifts the lower border of the mandible from the pericardial region, so allowing jaw movements to assist the withdrawal of the tongue from between the palatal shelves. This aids elevation of the palatal processes prior to fusion. This paper reports on histological sections of 28 mm twin embryos which demonstrate that palatal elevation can take place in the absence of elongation and straightening of the neck, demonstrated by observation of existence of cephalic, pontine and cervical neural flexure of the brain.