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Critical conditions for pattern formation and in vitro tubulogenesis driven by cellular traction fields.

In vitro angiogenesis assays have shown that tubulogenesis of endothelial cells within biogels, like collagen or fibrin gels, only appears for a critical range of experimental parameter values. These experiments have enabled us to develop and validate a theoretical model in which mechanical interactions of endothelial cells with extracellular matrix influence both active cell migration--haptotaxis--and cellular traction forces. Depending on the number of cells, cell motility and biogel rheological properties, various 2D endothelial patterns can be generated, from non-connected stripe patterns to fully connected networks, which mimic the spatial organization of capillary structures. The model quantitatively and qualitatively reproduces the range of critical values of cell densities and fibrin concentrations for which these cell networks are experimentally observed. We illustrate how cell motility is associated to the self-enhancement of the local traction fields exerted within the biogel in order to produce a pre-patterning of this matrix and subsequent formation of tubular structures, above critical thresholds corresponding to bifurcation points of the mathematical model. The dynamics of this morphogenetic process is discussed in the light of videomicroscopy time lapse sequences of endothelial cells (EAhy926 line) in fibrin gels. Our modeling approach also explains how the progressive appearance and morphology of the cellular networks are modified by gradients of extracellular matrix thickness.

Biomechanical Phenomena↗

EEG correlation and impulse activity of neuronal populations of individual structures of the cat brain during elaboration and reproduction of motor and alimentary reactions in instrumental conditioned reflexes.

Stable changes in EEG and spike activity of neuronal populations in different brain formations were studied on models of instrumental conditioned reflexes: motor and motor alimentary. A depencence has been established of the EEG amplitude-frequency parameters in the motor and striate cortical zones and the hippocampus on definite changes of unit spike activity in these areas. Simultaneous recording of the EEG and the spike activity of neuronal populations helps to elucidate the neurophysiological nature of individual rhythms of bio-electrical activity. Learned animals exhibit a stable reproduction of the spatial-temporal EEG patterns and motor alimentary reactions when automatic presentation of reinforcement is changed over to an arbitrary one.

Animals↗

Growth movements during prenatal development of human facial morphology.

After formation of the primary palate, human facial morphology develops rapidly and by 10-12 weeks pc the face has characteristics that appear typically human. The objective of this study was to review major growth movements and developmental changes in craniofacial tissues between 7 and 12 weeks pc. During this period (20 - 80 mm CR length), the upper and lower facial regions grow forward rapidly to achieve relationships to the cranial base that are similar to those present later prenatally and postnatally. Initial ossification of facial bones is present but the primary cartilages form the continuous craniofacial skeleton through the entire period. Rapid directional growth of the cartilaginous components between 7 and 10 weeks appears to be important to development of the typically human facial appearance prior to the formation of the continuous bony skeleton. The effects of altered primary cartilage growth on bony skeletal patterns were examined in experimental animal studies in which embryonic rats were exposed to teratogens. Reduction in the length of Meckel's cartilage or alteration in the shape of the cartilage was found to affect the size and shape of the bony mandible that developed later. Therefore, growth movements of the chondrocranium and Meckel's cartilage appear to play an important role in spatial relocation of developing facial bones during formation of craniofacial morphology. The results suggest that significant alterations in growth during this period when the primary cartilages form the continuous skeleton may produce significant irreversible effects on later prenatal and postnatal craniofacial morphology.

Animals↗

3-Dimensional anatomic relationship of serotonergic and muscarinic receptor binding in the pontine reticular formation of the human infant brainstem.

Cholinergic and serotonergic neurons of the rostral pontine reticular formation have been implicated by animal studies in the modulation of sleep and waking. To define better the spatial relationships between muscarinic and serotonergic receptor binding in the rostral human brainstem, we used 3-dimensional computer reconstructions of serial autoradiographs generated with radioligands to muscarinic and serotonergic receptors. Receptor binding was assessed in a series of 9 human infants, and 3-dimensional reconstructions were performed in a representative infant at 53 postconceptional weeks. The computer reconstructions demonstrated a 3-dimensional distinct pattern in the rostral pontine reticular formation, with high (3H)lysergic acid diethylamide binding to serotonin receptors in the median raphe nucleus flanked by paramedian bands of high (3H)quinuclidinyl benzilate binding to muscarinic cholinergic receptors in the medial nucleus pontis oralis. Based upon comparisons to animal data, we suggest that the muscarinic-serotonergic pattern of receptor binding in the rostral pontine reticular formation represents part of the neurochemical organization of the circuitry involved in the modulation of rapid eye movement (REM) sleep in humans.

Autoradiography↗

The egg came first, of course! Anterior-posterior pattern formation in Drosophila embryogenesis and oogenesis.

The anterior-posterior body pattern of the Drosophila embryo is initiated through the action of maternal gene products. In particular, three groups of maternally acting genes (the anterior, posterior and terminal groups) have been shown to direct the synthesis and spatial restriction of the three major organizing activities in the egg. The initial spatial localizations of the maternal organizing activities are established during oogenesis. After fertilization these activities regulate zygotic gene activity along the anterior-posterior axis of the egg.

Animals↗

Expression of insulin-like growth factor and binding protein genes during nephrogenesis.

To study the role of insulin-like growth factors (IGFs) and their binding proteins (IGFBPs) in human nephrogenesis, we examined the temporal and spatial pattern of expression of these genes using in situ hybridization. The uninduced metanephric blastema (MB) expressed abundant IGF-II mRNA. With induction by the ureteric duct (UD), the aggregated MB additionally expressed IGFBP-2 and IGFBP-4 mRNAs. The mature UD expressed IGFBP-3 mRNA while the ampulla in contact with the MB lacked IGFBP-3 mRNA and expressed IGFBP-2 exclusively. Upon formation of the S-shape nephron, IGFBP-2 mRNA was expressed in the committed glomerular and epithelial cells which also expressed IGF-II and IGFBP-4, and the mesenchyme of the vascular cleft expressed IGFBP-5 mRNA. In the maturing glomerulus, the glomerular epithelial cells expressed IGF-II mRNA together with IGFBP-2 and IGFBP-4 mRNAs, while IGFBP-5 mRNA was localized to the mesangium and supporting mesenchyme. As the proximal tubule was formed the epithelium expressed less of IGFBP-2 mRNA and more of IGFBP-4 mRNA. The renal mesenchyme in the cortex and medulla expressed abundant IGF-II mRNA, and lower levels of IGFBP-4 and -5 mRNAs. The epithelium of the collecting ducts and pelvicalyceal system expressed abundant IGFBP-3. In contrast, IGF-I, IGFBP-1, and IGFBP-6 mRNAs were expressed at low levels. The specific temporal and spatial pattern of expression of IGFBP genes on the background of abundant IGF-II gene expression suggests that the IGFBP peptides, as modulators of IGF action, are expressed locally at specific points of nephrogenesis to interact with IGF-II to regulate mesenchymal induction, renal epithelial cell commitment, differentiation and growth.

Carrier Proteins↗

The neuropharmacological and neurochemical basis of place learning in the Morris water maze.

The Morris water maze (MWM) offers several advantages over other methods of studying the neurochemical basis of learning and memory, particularly with respect to its ability to dissociate deficits in memory formation from deficits in sensory, motor, motivational and retrieval processes. The contributions of nearly all of the major neurotransmitter systems have been investigated and consistent patterns have emerged. Normal function in glutamatergic and cholinergic systems is necessary for spatial learning, as blockade of NMDA receptors and cholinergic hypofunction prevents spatial learning but does not impair recall. Peptides such as adrenal and sex hormones and somatostatin may also be necessary for spatial learning. In contrast, activity in either GABAergic or opioidergic systems impairs spatial learning, though by quite different means. GABAergic activity prevents memory function, whereas opioidergic activity reduces motivation. Normal monoaminergic activity is necessary for normal performance in the MWM, but not for spatial learning per se. However, noradrenergic and serotonergic systems may enhance cholinergic-mediated mnemonic processes. Further research into the relative contributions of different receptor subtypes as well as interactions between neurochemical systems should provide significant advances in our understanding of the neural basis of learning and memory in mammals.

Animals↗

Synchronization of vectorial noise-sustained structures.

The synchronization of vectorial, noise-sustained structures in nonlinear optical systems is discussed. In particular, the analysis is made for nondegenerate optical parametric oscillators with walk off. The interplay between walk off and noise fluctuations leads to the formation of noise-sustained transverse patterns in both the signal and idler fields. Despite the fact that both patterns are stochastic macroscopic structures driven by independent sources of noise, their correlation grows with time, finally leading to a spatially distributed time synchronization of noise-sustained structures. A physical explanation of this phenomenon is found by analyzing the linear instability process and the existence of exact nonlinear solutions that show the same correlation.

Journal Article↗

The regulation of Dictyostelium development by transmembrane signalling.

Dictyostelium discoideum has a well characterized life cycle where unicellular growth and multicellular development are separated events. Development is dependent upon signal transduction mediated by cell surface, cAMP receptor/G protein linkages. Secreted cAMP acts extracellularly as a primary signal and chemoattractant. There are 4 genes for the distinct cAMP receptor subtypes, CAR1, CAR2, CAR3 and CAR4. These subtypes are expressed with temporally and spatially specific patterns and cells carrying null mutations for each gene have distinct developmental phenotypes. These results indicate an essential role for cAMP signalling throughout Dictyostelium development to regulate such diverse pathways as cell motility, aggregation (multicellularity), cytodifferentiation, pattern formation and cell type-specific gene expression.

Animals↗

Shoot and floral meristem maintenance in arabidopsis.

The shoot apical meristem (SAM) of higher plants functions as a site of continuous organogenesis within which a small pool of pluripotent stem cells replenishes the cells incorporated into lateral organs. This article summarizes recent results demonstrating that the fate of stem cells in Arabidopsis shoot and floral meristems is controlled by overlapping spatial and temporal signaling systems. Stem cell maintenance is an active process requiring constant communication between neighboring groups of SAM cells. Information flows via a ligand-receptor signal transduction pathway, resulting in the formation of a spatial feedback loop that stabilizes the size of the stem cell population. Termination of stem cell activity during flower development is achieved by a temporal feedback loop involving both stem cell maintenance genes and flower patterning genes. These investigations are providing exciting insights into the components and activities of the stem cell regulatory pathway and into the interaction of this pathway with molecular mechanisms that control floral patterning.

Arabidopsis↗

Printing patterns of biospecifically-adsorbed protein.

The advancement of elastomeric patterning techniques in recent years has significantly enhanced our ability to spatially control biomaterial surface chemistry at the micrometre level. The application of this technology to the patterning of biomolecules onto solid surfaces has created many potential applications including the development of advanced biosensors, combinatorial library screening and the formation of tissue engineering templates. In this paper, we describe the direct patterning of protein by microcontact printing. An important consideration for the fabrication of protein micropatterns intended for these applications is the nature of the protein immobilization to a substrate. To date, the patterning of proteins by direct microcontact printing (microCP) has relied on the non-covalent adsorption to a substrate. Ideally, the proteins need to be firmly anchored onto a surface without adversely effecting their activity. Here, the high affinity avidin-biotin receptor-ligand interaction has been exploited to form arrays of avidin molecules onto a polymeric substrate expressing biotin moieties. This has created a generic technique by which any biotinylated species can be subsequently immobilized into defined patterns. Utilizing atomic force microscopy (AFM), the patterned surfaces have been characterized to molecular resolution. The micropatterned sample supported cell adhesion when biotin-(G)11-GRGDS was bound to the avidin bearing arrays.

3T3 Cells↗

Glutamatergically induced pattern of Ca2+ driving potential as a mechanism of postsynaptic plasticity.

Simulation studies were performed in a model of neuronal dendrite with Na+ and K+ channels and with ionotropic and metabotropic glutamate receptors. The ionotropic receptors were either N-methyl-D-aspartate (NMDA)-sensitive, voltage-dependent, and permeable to Ca2+, Na+, and K+, or non-NMDA-sensitive, voltage-independent, and permeable to Na+ and K+. The metabotropic receptors provided a catalytic effect on Ca2+-induced Ca2+ release from intracellular stores. Local intracellular concentration [Ca2+]i in the cytoplasm was changed because of exchange with the stores, axial diffusion, and transmembrane inward passive and outward pump fluxes. Tonic activation of ionotropic and metabotropic receptors in a particular range of intensities triggered the formation of spatially periodic [Ca2+]i hot and cold bands arising from an initial uniform state. The period and width of the bands were smaller at higher levels of tonic NMDA activation and higher metabotropically controlled rates of Ca2+-induced Ca2+ release. The bandwidths also depended on the dendrite diameter, the specific membrane, and cytoplasm resistivity. This activity-induced pattern led to long-term, spatially inhomogeneous change in local excitatory postsynaptic potentials (EPSPs) of NMDA synapses phasically activated with the same presynaptic intensity. The phasic EPSPs were potentiated if the synapse occurred in the hot band.

Animals↗

Collagenase-3 (MMP-13) and integral membrane protein 2a (Itm2a) are marker genes of chondrogenic/osteoblastic cells in bone formation: sequential temporal, and spatial expression of Itm2a, alkaline phosphatase, MMP-13, and osteocalcin in the mouse.

Endochondral bone formation requires the action of cells of the chondrocytic and osteoblastic lineage, which undergo continuous differentiation during this process. To identify subpopulations of resting, proliferating, and hypertrophic chondrocytes and osteoblasts involved in bone formation, we have identified here two novel marker genes present in endochondral and intramembranous ossification. Using Northern blot analysis and in situ hybridization on parallel sections of murine embryos and bones of newborn mice we compared the expression pattern of the recently cloned Itm2a and MMP-13 (collagenase-3) genes with that of established marker genes for bone formation, such as alkaline phosphatase (ALP), osteocalcin (OC), and collagen type X, during endochondral and intramembranous ossification. During embryonic development expression of Itm2a and ALP was detectable at midgestation (11.5 days postcoitum [dpc]) and increased up to 16.5 dpc. MMP-13 and OC expression started at 14.5 dpc and 16.5 dpc, respectively. This temporal expression was reflected in the spatial distribution of these markers in the growth plate of long bones. In areas undergoing endochondral ossification Itm2a expression was found in chondrocytes of the resting and the proliferating zones. Expression of ALP and MMP-13 are mutually exclusive: ALP transcripts were found only in collagen type X positive hypertrophic chondrocytes of the upper zone. MMP-13 expression was restricted to chondrocytes of the lower zone of hypertrophic cartilage also expressing collagen type X. In osteoblasts involved in endochondral and intramembranous ossification Itm2a was not present. ALP, MMP-13, and OC were mutually exclusively expressed in these cells suggesting a differentiation-dependent sequential expression of ALP, MMP-13, and OC. The identification of the continuum of sequential expression of Itm2a, ALP, MMP-13, and OC will now allow us to establish a series of marker genes that are highly suitable to characterize bone cells during chondrocytic and osteoblastic differentiation in vivo.

Alkaline Phosphatase↗

Extracellular matrix modifications in the interdigital spaces of the chick embryo leg bud during the formation of ectopic digits.

In previous studies we have observed that the interdigital mesenchyme of the chick leg bud, in the stages preceding the onset of cell death, retains a significant regulatory potential, forming ectopic extra digits under a variety of surgical manipulations. Most evidence suggests that interdigital extra digits are caused by the abolition of local antichondrogenic effects operating in the interdigital spaces under normal conditions rather than by modifications of the signalling mechanisms accounting for the normal patterning of the digits in early stages of development. The interdigital spaces exhibit a complex scaffold of extracellular matrix with well-defined domains of spatial distribution of type I and type VI collagens, tenascin, fibronectin, laminin and elastic matrix components that have been proposed to play a role in the establishment of the non-chondrogenic fate of the interdigital tissue in situ. In an attempt to analyze this possible role of the interdigital extracellular matrix (ECM), in the present work we have studied changes in the pattern of ECM distribution associated with the formation of extra digits. Extra digits were induced by making a T-cut in the third interdigital space of the leg but of stage 29 HH chick embryos. Subsequent modifications of the ECM were detected immunohistochemically in whole-mount specimens using laser confocal microscopy. Our results reveal that in the first hours after the operation, changes in the ECM apparently related to the healing of the wound cause a significant reorganization of the normal ECM scaffold of the interdigit. In addition, chondrogenesis of the interdigital tissue is preceded by disappearance of elastin fibers in the interdigital mesenchyme subjacent to the wound and by an intense deposition of tenascin. Tenascin deposition and loss of the elastin fibrillar scaffold were also observed preceding chondrogenesis in fragments of interdigital tissue explanted to culture conditions. The significance of these observations in relation to the establishment of the skeletal elements of the autopodium is discussed.

Animals↗

A model of spatial map formation in the hippocampus of the rat.

Using experimental facts about long-term potentiation (LTP) and hippocampal place cells, we model how a spatial map of the environment can be created in the rat hippocampus. Sequential firing of place cells during exploration induces, in the model, a pattern of LTP between place cells that shifts the location coded by their ensemble activity away from the actual location of the animal. These shifts provide a navigational map that, in a simulation of the Morris maze, can guide the animal toward its goal. The model demonstrates how behaviorally generated modifications of synaptic strengths can be read out to affect subsequent behavior. Our results also suggest a way that navigational maps can be constructed from experimental recordings of hippocampal place cells.

Action Potentials↗

Immunocytochemical detection and spatial distribution of myosin light-chain kinase in preimplantation mouse embryos.

As a follow-up to our previous study on the role of myosin light-chain kinase (MLCK), a Ca2+/calmodulin-dependent enzyme, in the development of preimplantation mouse embryos, we examined the presence and pattern of distribution of MLCK during preimplantation development of the mouse by whole-mount, indirect immunocytochemistry and by Western blotting, using a monoclonal antibody against MLCK. At all stages of preimplantation development, the nucleus was brightly stained with an unstained region around the nucleus, and regions near the cell membrane were also brightly stained. Using the optical sectioning capability of the confocal laser scanning microscope, we found that, up to the eight-cell stage, the regions of cell contact were mostly unstained, but along with the process of compaction, cell contact regions showed a clear staining pattern along with clearing of the cytoplasm. During formation of the blastocyst, a ring of immunofluorescence was found at the margin of the blastocoel. In the blastocyst, cells of the inner cell mass were less immunofluorescent than trophectoderm cells. These staining results appear to be due to specific immunoreaction between MLCK and the antibody, because the staining patterns were abolished when the antibody was preabsorbed by MLCK purified from chicken gizzard smooth muscle. In Western blotting of blastocysts, we found a band at 130 kD. We also show by immunoblotting and immunohistochemistry of various mouse tissues that the antibody used in this study has cross-reactivity to MLCK of various muscle and non-muscle tissues of the mouse. The presence and spatial distribution of MLCK at various stages of preimplantation development of the mouse suggest that it could play a crucial role in the regulation of the contractile events involved in the initial differentiation that occurs during formation of the mouse blastocyst.

Animals↗

Remodeling of myocardial sleeve and gap junctions in canine superior vena cava after rapid pacing.

OBJECTIVE: We studied the response of the superior vena cava (SVC) myocardial sleeve to atrial fibrillation (AF). METHODS AND RESULTS: We examined adult male dogs without pacing (N=6) and after rapid atrial pacing (600 bpm) for 2 weeks (P2w; N=5) and 6-8 weeks (P6-8w; N=5). After pacing, the sleeve was increased in thickness (non-paced vs. either paced group, both P<0.05). This was associated with an increase in proliferative activity, which was higher in the P2w than the P6-8w animals (P<0.05). In addition, collagen content increased, and the component cardiomyocytes become more unevenly oriented and shorter and narrower in shape (non-paced vs. either paced group, both P<0.05). Pacing had different effects on connexin40 (Cx40) and Cx43 gap junctions. There was a 98% increase in Cx43 signal in P2w, and a 74% increase in P6-8w animals (non-paced vs. each paced group, both P<0.05). In contrast, Cx40 signal decreased 47% in P2w but increased 44% in P6-8w animals (non-paced vs. each paced group, both P<0.05). CONCLUSIONS: Rapid atrial pacing results in a specific pattern of remodeling of the canine SVC sleeve, including changes in size and shape, spatial orientation, and gap junction expression profile of the component cardiomyocytes. These changes may co-operatively affect the electrical properties and contribute to the formation and maintenance of the arrhythmogenic substrate of AF.

Animals↗

Prediction of microdamage formation using a mineral-collagen composite model of bone.

Age-related changes in bone quality are mainly manifested in the reduced toughness. Since the post-yield deformation of bone is realized through microdamage formation (e.g., microcracking and diffuse damage), it is necessary to understand the mechanism of microdamage formation in bone in order to elucidate underlying mechanisms of age-related bone fractures. In this study, a two-dimensional shear lag model was developed to predict stress concentration fields around an initial crack in a mineral-collagen composite. In this model, non-linear elasticity was assumed for the collagen phase, and linear elasticity for the mineral. Based on the pattern of the stress concentration fields, the condition for microdamage formation was discussed. The results of our analyses indicate that: (1) an initial crack formed in mineral phase may cause stress concentration in the adjacent mineral layers; (2) the pattern of stress concentration fields depends not only on the spatial but also mechanical properties of the collagen and mineral phases; (3) the pattern of the stress concentration fields could determine either coalescence or scattering of nano cracks around the initial crack.

Animals↗