Three-dimensional analysis of intracellular organelles by cell shape difference with photo-micropatterning

Ultrastructural analyses of cytoplasmic changes in Saccharomyces cerevisiae X2180-1A (MATa) that had been treated with alpha factor were performed by using the freeze-substitution fixation method. After alpha factor treatment, cells exhibited a pointed projection, which is a unique pattern of oriented cell surface growth. The relationship between projection formation and intracellular organelles was examined using serial thin sections and computer-aided three-dimensional reconstructions. Using these analyses membrane vesicles and other organelles were detected, and studies on their dynamic structural reorganization became feasible. Production of membrane vesicles (average 65 nm in diameter) was induced upon exposure of the cells to alpha factor before projection emergence. The total number of membrane vesicles increased at the early stage and decreased at the late stage of projection formation. Three-dimensional analysis indicated that the vesicles were at first dispersed throughout the cell, then accumulated at the site where the projection formed. Morphological changes and multiplication of the Golgi body were seen during the process of projection formation. Other intracellular organelles (nucleus, vacuole, rough endoplasmic reticulum and mitochondria) were also rearranged, showing a polar organization of the cytoplasm during projection formation.

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In situ investigation of the primary recrystallization of alpha titanium in HVEM

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100110313 ◽

1978 ◽

Vol 36 (1) ◽

pp. 634-635

Author(s):

S. Naka ◽

R. Penelle ◽

R. Valle

Keyword(s):

Dimensional Analysis ◽

Texture Evolution ◽

Cold Work ◽

Three Dimensional ◽

Primary Recrystallization ◽

Thin Foils ◽

In Situ Investigation ◽

Grain Growth Model ◽

Alpha Titanium

The in situ experimentation technique in HVEM seems to be particularly suitable to clarify the processes involved in recrystallization. The material under investigation was unidirectionally cold-rolled titanium of commercial purity. The problem was approached in two different ways. The three-dimensional analysis of textures was used to describe the texture evolution during the primary recrystallization. Observations of bulk-annealed specimens or thin foils annealed in the microscope were also made in order to provide information concerning the mechanisms involved in the formation of new grains. In contrast to the already published work on titanium, this investigation takes into consideration different values of the cold-work ratio, the temperature and the annealing time.Two different models are commonly used to explain the recrystallization textures i.e. the selective grain growth model (Beck) or the oriented nucleation model (Burgers). The three-dimensional analysis of both the rolling and recrystallization textures was performed to identify the mechanismsl involved in the recrystallization of titanium.

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Three-Dimensional Subcellular Organization of Hepatocytes in Intact Liver: Simultaneous Visualization of Cytoskeletal and Membrane Markers by Confocal Microscopy

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100163903 ◽

1996 ◽

Vol 54 ◽

pp. 288-289

Author(s):

Greg V. Martin ◽

Ann L. Hubbard

Keyword(s):

Three Dimensional ◽

Integral Membrane Proteins ◽

Polarized Secretion ◽

Microscope Images ◽

Computer Aided ◽

Intracellular Organelles ◽

Cytoskeletal Elements ◽

Simultaneous Visualization

The microtubule (MT) cytoskeleton is necessary for many of the polarized functions of hepatocytes. Among the functions dependent on the MT-based cytoskeleton are polarized secretion of proteins, delivery of endocytosed material to lysosomes, and transcytosis of integral plasma membrane (PM) proteins. Although microtubules have been shown to be crucial to the establishment and maintenance of functional and structural polarization in the hepatocyte, little is known about the architecture of the hepatocyte MT cytoskeleton in vivo, particularly with regard to its relationship to PM domains and membranous organelles. Using an in situ extraction technique that preserves both microtubules and cellular membranes, we have developed a protocol for immunofluorescent co-localization of cytoskeletal elements and integral membrane proteins within 20 µm cryosections of fixed rat liver. Computer-aided 3D reconstruction of multi-spectral confocal microscope images was used to visualize the spatial relationships among the MT cytoskeleton, PM domains and intracellular organelles.

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