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Vitro Environment (vitro + environment)
Selected AbstractsDevelopment of glutamate receptors in auditory neurons from long-term organotypic cultures of the embryonic chick hindbrainEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 2 2009Carmen Diaz Abstract We used long-range organotypic cultures of auditory nuclei in the chick hindbrain to test the development of glutamate receptor activity in auditory neurons growing in a tissue environment that includes early deprivation of peripheral glutamatergic input, subsequent to removal of the otocyst. Cultures started at embryonic day (E)5, and lasted from 6 h to 15 days. Neuronal migration, clustering and axonal extension from the nucleus magnocellularis (NM) to the nucleus laminaris (NL) partially resembled events in vivo. However, the distinctive laminar organization of the NL was not observed. Glutamate receptor (GluR) activity was tested with optical recordings of intracellular Ca2+ in the NM. ,-Amino-3-hydroxy-5-methyl-4-isoxazoleproprionic acid (AMPA)/kainate receptors had Ca2+ responses with a time course similar to that in control slices. Peak amplitude, however, was significantly lower. N -methyl- d -aspartate (NMDA)-mediated Ca2+ responses were higher in 2-day cultures (E5 + 2d) than in E7 explant controls, returning later to control values. Metabotropic GluRs did not elicit Ca2+ responses at standard agonist doses. Blocking NMDA or AMPA/kainate receptors with specific antagonists for 10 days in culture did not limit neuronal survival. Blocking metabotropic GluRs resulted in complete neuronal loss. Thus, ionotropic GluRs are not required for NM neuronal survival. However, their activity during development is affected when neurons grow in an in vitro environment that includes prevention of arrival of peripheral glutamatergic input. [source] How does polyspermy happen in mammalian oocytes?MICROSCOPY RESEARCH AND TECHNIQUE, Issue 4 2003Wei-Hua Wang Abstract Polyspermy is one of the most commonly observed abnormal types of fertilization in mammalian oocytes. In vitro fertilization (IVF) provides approaches to study the mechanisms by which oocytes block polyspermic fertilization. Accumulated data indicate that oocyte, sperm and insemination conditions are all related to the occurrence of polyspermic fertilization. A high proportion of immature and aged oocytes showed polyspermy as compared with mature oocytes. Preincubation of oocytes and/or sperm with oviductal epithelial cells or collected oviductal fluid before IVF reduces polyspermic penetration. Recently, it was found that an abnormal zona pellucida is one of main causes of polyspermy in human eggs. A high proportion of polyspermy has resulted from the use of a high concentration of capacitated spermatozoa at the site of fertilization, irrespective of in the in vivo or in vitro environment. Oviductal secretions or oviductal epithelial cells themselves can regulate the number of spermatozoa reaching or binding to the zona pellucida thus reducing multiple sperm penetration. Suboptimal in vitro conditions, such as supplementations in IVF media, pH, and temperature during IVF, also induce polyspermic fertilization in some mammals. Species-specific differences are present regarding the relationship between insemination conditions and polyspermy. Microsc. Res. Tech. 61:335,341, 2003. © 2003 Wiley-Liss, Inc. [source] How microspores transform into haploid embryos: changes associated with embryogenesis induction and microspore-derived embryogenesisPHYSIOLOGIA PLANTARUM, Issue 1 2008José M. Seguí-Simarro Microspore embryogenesis is the most powerful androgenic pathway to produce haploid and doubled haploid plants. To deviate a microspore toward embryogenesis, a number of factors, different for each species, must concur at the same time and place. Once induced, the microspore undergoes numerous changes at different levels, from overall morphology to gene expression. Induction of microspore embryogenesis not only implies the expression of an embryogenic program, but also a stress-related cellular response and a repression of the gametophytic program to revert the microspore to a totipotent status. In this review, we compile the most recent advances in the understanding of the changes undergone by the induced microspore to readapt to the new developmental scenario. We devote special attention to the efforts made to uncover changes in the transcriptome of the induced microspore and microspore-derived embryo (MDE). Finally, we discuss the influence that an in vitro environment exerts over the MDE, as compared with its zygotic counterpart. [source] Cell deposition system based on laser guidanceBIOTECHNOLOGY JOURNAL, Issue 9 2006Russell K. Pirlo Abstract We have designed a laser cell deposition system that employs the phenomenon of laser guidance to place single cells at specific points in a variety of in vitro environments. Here, we describe the components of the system: the laser optics, the deposition chamber, the microinjection cell feeding system and our custom system control software application. We discuss the requirements and challenges involved in laser guidance of cells and how our present system overcomes these challenges. We demonstrate that the patterning system is accurate within one micrometer by repeatedly depositing polymer microspheres and measuring their position. We demonstrate its ability to create highly defined living patterns of cells by creating a defined pattern of neurons with neurite extensions displaying normal function. We found that the positional accuracy of our system is smaller than the variations in cell size and pattern disruptions that occur from normal cell movement during substrate adhesion. The laser cell deposition system is a potentially useful tool that can be used to achieve site- and time-specific placement of an individual cell in a cell culture for the systematic investigation of cell-cell and cell-extracellular matrix interactions. [source] | ||||||||||