Detection of Dynamic Spatiotemporal Response to Periodic Chemical Stimulation in a Xenopus Embryonic Tissue

Embryonic development is guided by a complex and integrated set of stimuli that results in collective system-wide organization that is both time and space regulated. These regulatory interactions result in the emergence of highly functional units, which are correlated to frequency-modulated stimulation profiles. We have determined the dynamic response of vertebrate embryonic tissues to highly controlled, time-varying localized chemical stimulation using a microfluidic system with feedback control. Our approach has enabled localized spatiotemporal manipulation of the steroid hormone dexamethasone (DEX) in Animal Cap (AC) tissues isolated from gastrulating Xenopus embryos. Using this approach we investigated cell-scale responses to precisely controlled stimulation by tracking the redistribution of a GFP-tagged DEX-reporter constructed from the human glucocorticoid receptor (GR). We exposed defined regions of a single AC explant to different stimulation conditions—continuous stimulation, periodic stimulation, and no stimulation. We observed collective behavior of the GR transport into the nucleus was first-order. Furthermore, the dynamic response was well-modeled by a first-order differential equation with a single time derivative. The model predicted that responses to periodic stimulations closely matched the results of the frequency-based experiments. We find that stimulation with localized bursts versus continuous stimulation can result in highly distinct responses. This finding is critical as controlled space and time exposure to growth factors is a hallmark of complex processes in embryonic development. These complex responses to cellular signaling and transport machinery were similar to emergent behaviors in other complex systems, suggesting that even within a complex embryonic tissue, the overall system can converge toward a predictive first-order response.

[1]  William C. Messner,et al.  Probing Cellular Dynamics with a Chemical Signal Generator , 2009, PloS one.

[2]  R. Jove,et al.  Regulation of Glucocorticoid Receptor Function through Assembly of a Receptor‐Heat Shock Protein Complex a , 1993, Annals of the New York Academy of Sciences.

[3]  R. Keller,et al.  Induction of neuronal differentiation by planar signals in Xenopus embryos , 1993, Developmental dynamics : an official publication of the American Association of Anatomists.

[4]  G. Hager,et al.  Visualization of glucocorticoid receptor translocation and intranuclear organization in living cells with a green fluorescent protein chimera. , 1996, Proceedings of the National Academy of Sciences of the United States of America.

[5]  W. Messner,et al.  Pressure-driven spatiotemporal control of the laminar flow interface in a microfluidic network. , 2007, Lab on a chip.

[6]  D. Beebe,et al.  Controlled microfluidic interfaces , 2005, Nature.

[7]  George C. Verghese,et al.  Operating Regimes of Signaling Cycles: Statics, Dynamics, and Noise Filtering , 2007, PLoS Comput. Biol..

[8]  Y. Saka,et al.  Understanding how morphogens work , 2008, Philosophical Transactions of the Royal Society B: Biological Sciences.

[9]  M. Horb,et al.  Germ layers to organs: using Xenopus to study "later" development. , 2006, Seminars in cell & developmental biology.

[10]  Brandon Kuczenski,et al.  Modulation of fluidic resistance and capacitance for long-term, high-speed feedback control of a microfluidic interface. , 2009, Lab on a chip.

[11]  Walter Kauzmann,et al.  The Structure and Properties of Water , 1969 .

[12]  L. Davidson,et al.  Experimental control of excitable embryonic tissues: three stimuli induce rapid epithelial contraction. , 2010, Experimental cell research.

[13]  Steven Vogel,et al.  Comparative Biomechanics: Life's Physical World , 2003 .

[14]  K. Umesono,et al.  Localization, trafficking, and temperature-dependence of the Aequorea green fluorescent protein in cultured vertebrate cells. , 1995, Proceedings of the National Academy of Sciences of the United States of America.

[15]  S Pennathur,et al.  Flow control in microfluidics: are the workhorse flows adequate? , 2008, Lab on a chip.

[16]  H. Korswagen,et al.  Wnt Gradient Formation Requires Retromer Function in Wnt-Producing Cells , 2006, Science.

[17]  B. Gumbiner,et al.  Paraxial protocadherin mediates cell sorting and tissue morphogenesis by regulating C-cadherin adhesion activity , 2006, The Journal of cell biology.

[18]  Simplification of Mathematical Models of Chemical Reaction Systems , 1998 .

[19]  J. Slack,et al.  Origin of Stem Cells in Organogenesis , 2008, Science.

[20]  James C. Smith.  Forming and interpreting gradients in the early Xenopus embryo. , 2009, Cold Spring Harbor perspectives in biology.

[21]  M. Richardson.  Diffusible gradients are out - an interview with Lewis Wolpert , 2009 .

[22]  Rustem F. Ismagilov,et al.  Dynamics of Drosophila embryonic patterning network perturbed in space and time using microfluidics , 2005, Nature.

[23]  G. Malacinski,et al.  Amphibian in vitro heart induction: a simple and reliable model for the study of vertebrate cardiac development. , 2003, The International journal of developmental biology.

[24]  M. Asashima,et al.  In vitro induction systems for analyses of amphibian organogenesis and body patterning. , 2001, The International journal of developmental biology.

[25]  M. Berridge,et al.  Calcium signalling: dynamics, homeostasis and remodelling , 2003, Nature reviews. Molecular cell biology.

[26]  L. V. Belousov,et al.  Scott F. Gilbert—Developmental Biology, 2010, Sinauer Associates, Inc., Sunderland, MA Ninth Edition , 2011, Russian Journal of Developmental Biology.

[27]  Y. Kalaidzidis,et al.  Kinetics of Morphogen Gradient Formation , 2007, Science.

[28]  I. Macara,et al.  Evidence using a green fluorescent protein-glucocorticoid receptor chimera that the Ran/TC4 GTPase mediates an essential function independent of nuclear protein import , 1996, The Journal of cell biology.

[29]  K. Kroll,et al.  Transgenic Xenopus embryos from sperm nuclear transplantations reveal FGF signaling requirements during gastrulation. , 1996, Development.

[30]  M. Mavrovouniotis,et al.  Simplification of Mathematical Models of Chemical Reaction Systems. , 1998, Chemical reviews.

[31]  Kerem Pekkan,et al.  Three-dimensional chemical profile manipulation using two-dimensional autonomous microfluidic control. , 2010, Journal of the American Chemical Society.

[32]  Lance A Davidson,et al.  Patterning and tissue movements in a novel explant preparation of the marginal zone of Xenopus laevis. , 2004, Gene expression patterns : GEP.

[33]  A. Brivanlou,et al.  Scaling of BMP gradients in Xenopus embryos , 2009, Nature.

[34]  D. Melton,et al.  Vertebrate embryonic induction: mesodermal and neural patterning. , 1994, Science.

[35]  Scott E Fraser,et al.  Convergent extension: the molecular control of polarized cell movement during embryonic development. , 2002, Developmental cell.

[36]  R. Keller,et al.  Cell rearrangement during gastrulation of Xenopus: direct observation of cultured explants. , 1991, Development.

[37]  H. Sive,et al.  Efficient hormone-inducible protein function in Xenopus laevis. , 1995, Developmental biology.

[38]  K Ribbeck,et al.  Kinetic analysis of translocation through nuclear pore complexes , 2001, The EMBO journal.

[39]  L. Wolpert.  Diffusible gradients are out - an interview with Lewis Wolpert. Interviewed by Richardson, Michael K. , 2009, The International journal of developmental biology.

[40]  Marek Mlodzik,et al.  Planar cell polarization: do the same mechanisms regulate Drosophila tissue polarity and vertebrate gastrulation? , 2002, Trends in genetics : TIG.

[41]  L. Wolpert.  Developmental Biology , 1968, Nature.

[42]  David H. Sharp,et al.  Dynamic control of positional information in the early Drosophila embryo , 2004, Nature.

[43]  M. Freeman.  Feedback control of intercellular signalling in development , 2000, Nature.

[44]  M. Elowitz,et al.  Frequency-modulated nuclear localization bursts coordinate gene regulation , 2008, Nature.

[45]  G. Whitesides,et al.  Polymer microstructures formed by moulding in capillaries , 1995, Nature.

[46]  B. Grzybowski,et al.  Nanoparticle-based solution deposition of gold films supporting bioresistant SAMs. , 2009, Langmuir : the ACS journal of surfaces and colloids.

[47]  Paul Dungel,et al.  Distribution of [3H]Dexamethasone in Rat Subcutaneous Tissue after Delivery from Osmotic Pumps , 2006, Biotechnology progress.

[48]  Julian Lewis,et al.  From Signals to Patterns: Space, Time, and Mathematics in Developmental Biology , 2008, Science.

[49]  B. Kay.  Xenopus laevis: Practical uses in cell and molecular biology. Injections of oocytes and embryos. , 1991, Methods in cell biology.