El miocardio ventricular desde un enfoque cuantitativo. Su relación con la longitud del embrión humano / The ventricular myocardium from a quantitative approach. It ́s relation to the length of the human embryo

Introducción: Pese al incremento en conocimientos de la morfogénesis cardiaca humana, se conoce poco sobre los detalles cuantitativos en ello. Objetivo: Describir cuantitativamente el desarrollo del miocardio ventricular compacto y no compacto y su correlación con la longitud cráneo rabadilla. Métodos: Estudio descriptivo, transversal en 18 embriones humanos de los estadios 17 al 23 de Carnegie, pertenecientes a la Embrioteca de la Universidad de Ciencias Médicas de Villa Clara. Se midió la longitud del embrión, el grosor del miocardio compacto, trabecular y total en la pared lateral de ambos ventrículos y del vértice cardiaco. Resultados: El grosor de la pared lateral del miocardio compacto aumenta en ambos ventrículos desde los estadios 17 al 23 de Carnegie, de 0,06 mm hasta 0,17 mm en el derecho y de 0,09 mm hasta 0,23 mm en el izquierdo. El grosor de la pared lateral trabeculada disminuye con el avance de los estadios, de 0,43 mm a 0,34 mm en el derecho y de 0,45 mm a 0,37 mm en el izquierdo. El grosor de la pared lateral total aumenta de 0,48 mm a 0,51 mm en el ventrículo derecho y de 0,52 mm a 0,62 mm en el izquierdo. El grosor de la pared del vértice compacto aumenta de 0,19 mm a 0,25 mm. Conclusiones: La compactación de la pared ventricular aumenta con el desarrollo; la longitud cráneo raquis se relaciona con el grosor del miocardio ventricular(AU)

Introduction: Despite the increase in knowledge of human morphogenesis, especially cardiogenesis and the processes by which the morphology of the ventricular myocardium is defined, little is known about the quantitative details in it. Objectives: To quantitatively describe the development of compact and non-compact ventricular myocardium and its correlation with cranio-rump length. Methods: descriptive, cross-sectional study in 18 human embryos from Carnegie stages 17 to 23, belonging to the Embryoteca of the Villa Clara University of Medical Sciences. The length of the embryo, the thickness of the compact, trabecular and total myocardium were measured in the lateral wall of both ventricles and the cardiac apex. Results: The thickness of the lateral wall of the compact myocardium increases in both ventricles from Carnegie stages 17 to 23, from 0.06 mm to 0.17 mm in the right and from 0.09 to 0.23 mm in the left ventricles. The thickness of the trabeculated lateral wall decreases with the advancement of the stages, from 0.43 mm to 0.34 mm in the right and from 0.45 mm to 0.37 mm in the left. The total lateral wall thickness increases from 0.48 mm to 0.51 mm in the right ventricle and from 0.52 mm to 0.62 mm in the left. The wall thickness of the compact vertex increases from 0.19 mm to 0.25 mm. Conclusions: Ventricular wall compaction increases with development; the cranio-spinal length is related to the thickness of the ventricular myocardium(AU)

Efeito do estradiol e da progesterona no desenvolvimento e na qualidade de embriões bovinos produzidos in vitro / Effect of estradiol and progesterone on development and quality of bovine embryos produced in vitro

Avaliaram-se o desenvolvimento e a qualidade de embriões bovinos, cocultivados com células epiteliais do oviduto bovino (CEOBs) expostas ou não ao estradiol e à progesterona. Os ovócitos foram maturados in vitro por 24h e, então, fertilizados utilizando-se sêmen congelado, em estufa de CO2 a 5 por cento e 38,5ºC. As CEOBs foram cultivadas em TCM-199 com ou sem estradiol (E2) (24 horas), nas mesmas condições da maturação e fertilização in vitro (MIV e FIV), e, em seguida, adicionadas aos diferentes grupos em CR2 com ou sem progesterona (P4) (G1=P4+E2); (G2=E2); (G3=P4) e (G4=controle). Após 18h da FIV, as células foram cultivadas nos diferentes sistemas. Nenhuma diferença (P>0,05) foi observada nas taxas de clivagem entre G1, G2 e G4 (53,5 por cento; 56,3 por cento; 51,7 por cento) e nos padrões de blastocistos (BLs) (29,3 por cento; 31,2 por cento, 28,7 por cento). Índices menores (P<0,05) foram obtidos no G3 para ambas as variáveis (34,5 por cento; 16,4 por cento). G1 e G2 apresentaram taxas de eclosão maiores (P<0,05) que os outros grupos (23,3 por cento; 23,2 por cento), sendo G4 (19,3 por cento) diferente de G3 (16,1 por cento). Em G1, G2 e G3, o número de células nos BLs aumentou 125,9; 128,4 e 123,6, respectivamente (P<0,05), em relação ao G4 (112,5). Conclui-se que o tratamento das CEOBs com o E2, nas primeiras 24 horas de cultivo, pode ser usado isoladamente ou em combinação com a progesterona, a fim de melhorar a qualidade de embriões bovinos produzidos in vitro.

The development and quality of bovine embryos co-cultivated with bovine oviduct epithelial cells (BOEC) supplemented or not with estradiol and progesterone were evaluated. Oocytes were matured and fertilized in vitro (MIV/FIV) (5 percentCO2/38.5ºC). The BOEC were cultivated in TCM-199 with or without estradiol (E2) (24h), in the same conditions of MIV/FIV. Presumptive zygotes were transferred to BOEC in suspension after in vitro maturation and fertilization of bovine oocytes with thawed percoll-selected sperm. The zygotes were cultivated in CR2 medium containing or not progesterone (P4) (G1=P4+E2), (G2=E2), (G3=P4), and (G4=control). No significant differences (P>0.05) were found in the cleavage rates among G1, G2, and G4 (53.5 percent, 56.3 percent, and 51.7 percent) as well as in relation to the blastocystis (BL) rates (29.3 percent, 31.2 percent, and 28.7 percent). However, significant differences (P<0.05) were observed in the G3 for both variables (34.5 percent and 16.4 percent). G1 and G2 showed hatching rates higher (P<0.05) than the other groups (23.3 percent; 23.2 percent), being G4 (19.3 percent) different from G3 (16.1 percent) (P<0.01). In the groups G1, G2, and G3, the total cell number of the BL increased (125.9, 128.4, and 123.6) (P<0.05) compared to G4 (112.5). These results demonstrate that the treatment of the BOEC with estradiol in first the 24 hours of culture can be used separately or in combination with the progesterone to improve the quality of bovine embryos produced in vitro.

Influence of climatic conditions on in vitro production of bovine embryos / Influência do clima na produção in vitro de embriões bovinos

Temperature and rainfall were analyzed daily during six years to evaluate their influence on in vitro production of bovine embryos. Weekly replications (n=480) were performed on 14,778 ovaries collected at slaughterhouses. Cumulus oocyte complexes (n=19,180) were fertilized with a pool of Bos taurus taurus semen in one incubator with 5 percent CO2. Presumable zygotes were cultured in gasified plastic bags with 5 percent CO2, 5 percent O2, and 90 percent N2. In the first year, cleavage and embryo yield were 60.3 percent and 15.6 percent, respectively, being lower (P<0.05) than in the following years. Average cleavage rates were always lower in winter (P<0.0001), thus producing less embryos. Winter climatic conditions had a negative influence on in vitro production, when cleavage and embryo yield declined, possibly because of reduced availability and growth of native pasture.

A temperatura e a precipitação pluviométrica foram analisadas diariamente, durante seis anos, para avaliar sua influência sobre a produção in vitro de embriões bovinos. As repetições semanais (n=480) foram realizadas com 14.778 ovários coletados em matadouros. Os oócitos (n=19.180) foram maturados em estufa com atmosfera com controle de temperatura e umidade saturada com 5 por cento de CO2 e, após 20h, foram fecundados com sêmen de Bos taurus taurus e mantidos sob as mesmas condições de atmosfera da maturação. Os zigotos foram cultivados em placas de quatro poços em bolsas gaseificadas com 5 por cento de CO2, 5 por cento de O2 e 90 por cento de N2, à temperatura de 39ºC e umidade saturada. No primeiro ano, a taxa clivagem (60,3 por cento) e a produção de embriões (15,6 por cento) foram inferiores (P<0,05) aos demais anos. As taxas de clivagem foram sempre menores no inverno (P<0,0001). As condições climáticas no inverno tiveram influência negativa sobre a produção in vitro de embriões bovinos e houve diminuição nos índices de clivagem e produção de blastocistos, possivelmente devido à reduzida disponibilidade e crescimento da pastagem nativa.

Desarrollo embrionario y estrategias antiluteoliticas hormonales en programas de transplante de embriones bovinos / Embryonic development and hormonal antiluteolytic strategies for embryo transfer programs in cattle

Durante el desarrollo embrionario se establecen múltiples interacciones entre hormonas,factores de crecimiento (FC) y diferente tipo de moléculas, con lo cual se genera una seriede señales que desencadenan el reconocimiento materno de preñez (RMP) entre los días 15y 17, momento en el cual se genera un “periodo crítico”, cuando el endometrio libera lahormona prostaglandina F2α (PGF2α)para causar regresión del cuerpo lúteo (CL), en el cualse sintetiza progesterona (P4), hormona encargada de favorecer un adecuado ambienteuterino para el desarrollo embrionario y el mantenimiento de la preñez. El embrión en desarrollogenera señales antiluteolíticas que bloquean la producción de la PGF2α, por lo que se podríasugerir que el mantenimiento de la preñez es dependiente de la efectividad de este bloqueo,entre otros factores; el cual es generado por la acción del Interferón tau (IFN-τ), producidoen las células mononucleares del trofoblasto embrionario. Este bloqueo garantiza la integridaddel CL y de esta forma la producción normal de P4. Por lo anterior se podría pensar que lamanipulación estratégica apoyada hormonalmente, mejora las condiciones para el efectivobloqueo de los agentes luteolíticos que tienen su mayor actividad durante el llamado “periodocrítico”, con lo cual se mejora la tasa de sobrevivencia en programas de transplante embrionarioya que se crearía un ambiente uterino adecuado para el establecimiento y normal desarrollodel embrión. Por esto mismo, últimamente se han planteado mecanismos de apoyo hormonalque causan un bloqueo apropiado en cuanto a la producción de PGF2α uterina durante losdías15 y 17 del ciclo estral.

Eye-open at birth phenotype with reduced keratinocyte motility in LGR4 null mice.

We observed a consistent eye-open at birth (EOB) phenotype in mouse pups homozygous for a leucine-rich repeat containing G-protein coupled receptor 4 (Lgr4) allele deleting the whole transmembrane domain coding region. An in vitro wound-healing scratch assay showed notably reduced keratinocyte motility in the null mice. Phalloidin staining of F-actin in the eyelid epidermis was also reduced. We also generated keratinocyte-specific Lgr4 deficient mice, circumventing the embryonic/neonatal lethality and kidney abnormalities. Most of the conditional Lgr4 knockout mice showed the EOB phenotype. Thus, Lgr4 might be a novel gene class regulating cell motility.

Fate-mapping the mammalian hindbrain: segmental origins of vestibular projection neurons assessed using rhombomere-specific Hoxa2 enhancer elements in the mouse embryo.

As a step toward generating a fate map of identified neuron populations in the mammalian hindbrain, we assessed the contributions of individual rhombomeres to the vestibular nuclear complex, a major sensorimotor area that spans the entire rhombencephalon. Transgenic mice harboring either the lacZ or the enhanced green fluorescent protein reporter genes under the transcriptional control of rhombomere-specific Hoxa2 enhancer elements were used to visualize rhombomere-derived domains. We labeled functionally identifiable vestibular projection neuron groups retrogradely with conjugated dextran-amines at successive embryonic stages and obtained developmental fate maps through direct comparison with the rhombomere-derived domains in the same embryos. The fate maps show that each vestibular neuron group derives from a unique rostrocaudal domain that is relatively stable developmentally, suggesting that anteroposterior migration is not a major contributor to the rostrocaudal patterning of the vestibular system. Most of the groups are multisegmental in origin, and each rhombomere is fated to give rise to two or more vestibular projection neuron types, in a complex pattern that is not segmentally iterated. Comparison with studies in the chicken embryo shows that the rostrocaudal patterning of identified vestibular projection neuron groups is generally well conserved between avians and mammalians but that significant species-specific differences exist in the rostrocaudal limits of particular groups. This mammalian hindbrain fate map can be used as the basis for targeting genetic manipulation to specific subpopulations of vestibular projection neurons.

Retinoic acid is required for specification of the ventral eye field and for Rathke's pouch in the avian embryo.

We have investigated the role of retinoic acid (RA) in eye development using the vitamin A deficient quail model system, which overcomes problems of retinoic acid synthesising enzyme redundancy in the embryo. In the absence of retinoic acid, the ventral optic stalk and ventral retina are missing, whereas the dorsal optic stalk and dorsal retina develop appropriately. Other ocular abnormalities observed were a thinner retina and the lack of differentiation of the lens. In an attempt to explain this, we studied the expression of various dorsally and ventrally expressed genes such as Pax2, Pax6, Tbx6, Vax2, Raldh1 and Raldh3 and noted that they were unchanged in their expression patterns. In contrast, the RA catabolising enzymes Cyp26A1 and Cyp26B1 which are known to be RA-responsive were not expressed at all in the developing eye. At much earlier stages, the expression domain of Shh in the prechordal plate was reduced, as was Nkx2.1 and we suggest a model whereby the eye field is specified according to the concentration of SHH protein that is present. We also describe another organ, Rathke's pouch which fails to develop in the absence of retinoic acid. We attribute this to the down-regulation of Bmp2, Shh and Fgf8 which are known to be involved in the induction of this structure.

Criopreservação e desenvolvimento in vitro de embriões bovinos / Cryopreservation and in vitro survival of bovine embryos

O objetivo deste trabalho foi estudar a remoção do crioprotetor, em duas ou três etapas, em embriões bovinos produzidos in vitro após a congelação em vapor de Nirtogênio. Blastocistos expandidos (1329) foram mantidos em co-cultivo (controle) ou criopreservados em 3 protocolos de congelação em vapor de nitrogênio. Os embriões foram equilibrados na solução de 10% de EG por 10 minutos e em 17%, 22% ou 28% de EG por 30 segundos. Após o envase, as palhetas foram mantidas em vapor de nitrogênio por 2 minutos e armazenadas em nitrogênio líquido. Após a descongelação, os crioprotetores foram diluídos em duas etapas, usando 0,3M de sacarose e solução isotônica ou em três etapas usando 0,3M de sacarose + 10% de EG; 0,3M de sacarose e solução isotônica. Os embriões foram co-cultivados com células da granulosa, avaliando as taxas de re-expansão após 24 horas e de eclosão após 24, 48, 72 e 96 horas. Para os grupos congelados no vapor e diluição do crioprotetor em duas etapas, as taxas de eclosão foram de 1,94; 11,88 e 6,06% para EG17, EG 22 e EG 28 respectivamente. Já para os grupos com diluição do crioprotetor em três etapas, as taxas de eclosão foram de 4,67; 9,90 e 10,78% para EG17, EG22 e EG28 respectivamente.

The aim of this study was to evaluate the dilution of cryoprotectant in 2 or steps of bovine in vitro-produced embryos after quick-freezing. A total of 1329 expanded blastocyst were kept in co-culture as control group or cryopreserved by 3 quick-freezing protocols. The embryos were exposed to 10% EG for 10 minutes then to 17%, 22% or 28% for 30 seconds. After loading, the straws were held in nitrogen vapor for 2 minutes and then plunged and stored in liquid nitrogen. After warming, cryoprotectants were diluted in two steps using 0.3 M sucrose and isotonic solution, or three steps using 0.3 M sucrose + 10% EG then 0.3 M sucrose and isotonic solution. Embryos were co-cultured on a granulosa cell monolayer and evaluated after 24 hours for re-expansion and at 24, 48, 72 and 96 hours of co-culture for hatching rates. The in vitro survival rates of embryos cryopreserved by the quick-freezing method and two-step cryoprotectant dilution were 1.94; 11.88 and 6.06%, for EG17, EG22 and EG28 groups, respectively. At the three step dilution, the in vitro survival rates were 4.67; 9.90 and 10.78% for EG17, EG22 and EG28 groups, respectively.

Expression of TGFbeta Family in the Developing Internal Ear of Rat Embryos

In order to investigate the expression patterns of the transforming growth factor (TGF)beta isoforms in the internal ear, an immunohistochemical study of rat embryos was performed. Rat embryos were taken on the 13th, 15th, 17th, and 19th day after conception and their internal ears were immunohistochemically stained against TGF beta1, beta2, and beta3. As a result, the 13-day-old embryo showed a very weak positivity to TGF beta1. After the 15th day of pregnancy, no reactivity to TGF beta1 was defected. Immunoreactivity to TGF beta2 was observed from the 15th day of pregnancy throughout the rest of the period. The ampulla of the semicircular canal and the cochlear duct showed a notably strong immunohistochemical reaction. A strong reaction to TGF beta3 was observed on the 15th day of pregnancy. However, no positive reactions were observed thereafter. A strong immunoreactivity was observed especially on the apical cytoplasms, the surfaces of the epithelial cells, and basement membranes of the cochlear duct, as well as the semicircular canals of the developing internal ear of rat embryo.

Effects of male and female sex steroids on the development of normal and the transient Froriep's dorsal root ganglia of the chick embryo.

Sex steroids can influence developmental processes and support the survival of neurons in the embryonic central nervous system. Recent studies have shown that estrogen receptors are also expressed in the peripheral nervous system, in the dorsal root ganglia (DRG) of chick embryos. However, no studies have examined the effects of sex steroids on development of embryonic DRG. In the present study, 0.2 microg, 1.0 microg, 5.0 microg 10 microg, 20 microg, 25 microg, and 40 microg doses of testosterone or estradiol were delivered to chick embryos at Hamburger and Hamilton stage 18 (E3). The actions of these doses of sex steroids on the development of the C5DRG (fifth cervical ganglion, a "normal" DRG) and C2DRG (a transient ganglion known as a "Froriep's DRG") were then evaluated by quantifying ganglionic volumes, cell number, proliferation, and apoptosis after 1 day of growth to stage 23. We found that both testosterone and estradiol promoted proliferation of cells in both normal DRG and the Froriep's ganglia. By contrast, estradiol significantly increased the number of apoptotic cells, while testosterone strongly inhibited apoptosis. These actions of sex steroids on DRG development were dose-dependent, and C5DRG and C2DRG showed different sensitivities to the applied sex steroids. In addition, the present results demonstrated that specific ER and AR inhibitors (tamoxifen and flutamide) did not influence the effects of 5 microg E2 and 5 microg T on C2 and C5DRG significantly. These results demonstrate that male and female sex steroids can modulate DRG development through an epigenetic mechanism, as had been shown for the central nervous system.

La bioetica y la dignidad humana Que hay de nuevo sobre la Ética y el Pre-embrión Humano? / Bioethics and human dignity

El reconocimiento universal a la dignidad humana no ha sido una empresa fácil de conseguir ni tampoco es comprensible para muchos contemporáneos. Las razones son muchas, pero sobresalen estas:Necesidad de un desarrollo del juicio moral que haga comprensible los valores morales universales.Desarrollo incipiente de sociedades bien organizadas.Climas políticos propios.También se necesita dilucidar “por qué las personas son dignas”, entendiendo por dignidad una cualidad transitiva que implica que alguien es merecedor de recibir un trato especial a cambio o en proporción a un mérito o condición también especial. Lo anterior nos pone a pensar sobre la siguiente afirmación: “El pre-embrión humano no es persona, pero sí el embrión. El pre-embrión humano es persona humana potencial”.Entonces...¿será el pre-embrión un ser carente de dignidad humana?.

La realidad del Embrion Humano en los primeros quince dias de vida / The reality of the Human Embryo in the first fifteen days of life

El embrión temprano no es un simple tejido homogéneo e indiferenciado. El cigoto (o fase unicelular del individuo) se constituye, a partir del material heredado de los progenitores, como una célula con organización polarizada y con una propiedad peculiar que la distingue de cualquier otra célula: contiene el plano de las primeras divisiones celulares y se organiza en una unidad vital, tanto en sus estructuras espaciales como en sus funciones. Es un organismo en su fase inicial más sencilla y no una mera célula. El concepto de preembrión (aplicado al embrión preimplantatorio), como una fase del desarrollo en que no ha alcanzado el carácter de individuo de la especie, por la posibilidad de dar origen por división a gemelos monocigóticos, carece de fundamento biológico. Un embrión no se parte en dos mitades porque es asimétrico y las diversas células que lo componen son diferentes entre sí, desde el estado de dos células. El estatus del embrión preimplantatorio (generado naturalmente o creado in vitro) es el mismo: individuo de la especie humana. In vitro disminuye drásticamente la capacidad de un correcto desarrollo, en simbiosis armonizada con la madre; pero no significa menor humanidad, sino que siendo un ser humano, se le ha generado y situado en unas circunstancias en las que la capacidad de seguir viviendo está limitada

Zebrafish foxi1 mediates otic placode formation and jaw development.

The otic placode is a transient embryonic structure that gives rise to the inner ear. Although inductive signals for otic placode formation have been characterized, less is known about the molecules that respond to these signals within otic primordia. Here, we identify a mutation in zebrafish, hearsay, which disrupts the initiation of placode formation. We show that hearsay disrupts foxi1, a forkhead domain-containing gene, which is expressed in otic precursor cells before placodes become visible; foxi1 appears to be the earliest marker known for the otic anlage. We provide evidence that foxi1 regulates expression of pax8, indicating a very early role for this gene in placode formation. In addition, foxi1 is expressed in the developing branchial arches, and jaw formation is disrupted in hearsay mutant embryos.

Influence of the embryonic preplate on the organization of the cerebral cortex: a targeted ablation model.

Transgenic mice were generated to permit the targeted ablation of cortical preplate cells at the time they are born. In these mice, the 1.3 kb golli promoter of the myelin basic protein gene was used to drive the herpes simplex virus thymidine kinase (TK) transgene in cortical preplate cells. Heterozygous transgenic pairs were bred, and pregnant dams were treated with ganciclovir at embryonic days 11-12 to ablate preplate cells at the time the preplate was forming. This paradigm exposed control (TK-) and experimental (TK+) littermates to exactly the same conditions. Embryological ablation of preplate cells led to an early disruption of the radial glial framework and subplate structure in the developing cortex and dramatically altered the cellular lamination and connectivity of the cortical plate. The disturbed radial glial network contributed to an impaired radial migration of neurons into the cortical plate from the ventricular zone. The cortical plate became dyslaminated, and there was a substantial reduction in short- and long-range cortical projections within the cortex and to subcortical regions. Cell death within the cortical plate and the proliferative zones was substantially increased in the ablated animals. After birth, a cortical lesion developed, which became exacerbated with the secondary onset of hydrocephaly in the second postnatal week. The results underscore the critical importance of the preplate in cortex formation, mediated through its guidance of the formation of radial glial scaffolding, subsequent neuronal migration into the incipient cortical plate, and the final arrangement of its vertical organization and cellular connectivity.

El estatuto moral del embrión / Embryo¹s moral status

No disponible

No disponible

Embryogenomics: developmental biology meets genomics.

Fundamental questions in developmental biology are: what genes are expressed, where and when they are expressed, what is the level of expression and how are these programs changed by the functional and structural alteration of genes? These questions have been addressed by studying one gene at a time, but a new research field that handles many genes in parallel is emerging. The methodology is at the interface of large-scale genomics approaches and developmental biology. Genomics needs developmental biology because one of the goals of genomics--collection and analysis of all genes in an organism--cannot be completed without working on embryonic tissues in which many genes are uniquely expressed. However, developmental biology needs genomics--the high-throughput approaches of genomics generate information about genes and pathways that can give an integrated view of complex processes. This article discusses these new approaches and their applications to mammalian developmental biology.

Reconstrucción tridimensional de la mano de un embrión humano de 43 días: evaluación experimental, teoría y metodología / Three-dimensional reconstruction of the hand of a human embryo of 43 days: experimental evaluation, theory and methodology

El avance tecnológico en la práctica obstétrica y particularmente en el campo de la ultrasonografía ha dejado en evidencia el impacto que la visualización del dinámico desarrollo de la gestación ha tenido, en el conocimiento de los fenomenos que ocurren durante la vida intrauterina. En esta perspectiva y dada la demanda creciente de información, particularmente del desarrollo embrionario, la reconstrucción tridimensional virtual desde secciones bidimensionales se presenta como una nueva opci¢n y una herramienta util al conocimiento del desarrollo de estructuras complejas durante la morfogénesis. En este trabajo se presentan material, métodos, y resultados experimentales que se obtuvieron al desarrollar un sistema computacional interactivo para la reconstrucción tridimensional de la mano de un embrión humano de 43 días a partir de cortes seriados. Se describen los componentes principales del ambiente virtual, las principales técnicas de generación de actores tridimensionales y en especial la técnica Marching Cubes empleada para la generación de superficie de la piel y de la estructura esquelética de la mano. Se discute el problema de registración de imagenes, describiendo la técnica de registraci¢n de puntos rígidos empleada y se presenta un método alternativo de registración secundaria para solucionar problemas derivados por la distancia existente entre los puntos fiduciales y la muestra en estudio. Se presentan además, los resultados obtenidos de la aplicación de la técnica Data Cutting e interpolación trilineal para la obtención de cortes virtuales sagitales, axiales y coronales. Sobre la base de los resultados obtenidos, se proponen futuras aplicaciones incorporando técnicas de textura y de visualización en función del tiempo. Se enfatiza la importancia de la interdisciplina en el logro de estos objetivos

Generation of a novel functional neuronal circuit in Hoxa1 mutant mice.

Early organization of the vertebrate brainstem is characterized by cellular segmentation into compartments, the rhombomeres, which follow a metameric pattern of neuronal development. Expression of the homeobox genes of the Hox family precedes rhombomere formation, and analysis of mouse Hox mutations revealed that they play an important role in the establishment of rhombomere-specific neuronal patterns. However, segmentation is a transient feature, and a dramatic reconfiguration of neurons and synapses takes place during fetal and postnatal stages. Thus, it is not clear whether the early rhombomeric pattern of Hox expression has any influence on the establishment of the neuronal circuitry of the mature brainstem. The Hoxa1 gene is the earliest Hox gene expressed in the developing hindbrain. Moreover, it is rapidly downregulated. Previous analysis of mouse Hoxa1(-/-) mutants has focused on early alterations of hindbrain segmentation and patterning. Here, we show that ectopic neuronal groups in the hindbrain of Hoxa1(-/-) mice establish a supernumerary neuronal circuit that escapes apoptosis and becomes functional postnatally. This system develops from mutant rhombomere 3 (r3)-r4 levels, includes an ectopic group of progenitors with r2 identity, and integrates the rhythm-generating network controlling respiration at birth. This is the first demonstration that changes in Hox expression patterns allow the selection of novel neuronal circuits regulating vital adaptive behaviors. The implications for the evolution of brainstem neural networks are discussed.

Morphogenesis of prechordal plate and notochord requires intact Eph/ephrin B signaling.

Eph receptors and their ligands, the ephrins, mediate cell-to-cell signals implicated in the regulation of cell migration processes during development. We report the molecular cloning and tissue distribution of zebrafish transmembrane ephrins that represent all known members of the mammalian class B ephrin family. The degree of homology among predicted ephrin B sequences suggests that, similar to their mammalian counterparts, zebrafish B-ephrins can also bind promiscuously to several Eph receptors. The dynamic expression patterns for each zebrafish B-ephrin support the idea that these ligands are confined to interact with their receptors at the borders of their complementary expression domains. Zebrafish B-ephrins are expressed as early as 30% epiboly and during gastrula stages: in the germ ring, shield, prechordal plate, and notochord. Ectopic overexpression of dominant-negative soluble ephrin B constructs yields reproducible defects in the morphology of the notochord and prechordal plate by the end of gastrulation. Notably disruption of Eph/ephrin B signaling does not completely destroy structures examined, suggesting that cell fate specification is not altered. Thus abnormal morphogenesis of the prechordal plate and the notochord is likely a consequence of a cell movement defect. Our observations suggest Eph/ephrin B signaling plays an essential role in regulating cell movements during gastrulation.