Chloroplast ultrastructure in leaves ofUrtica dioica L. analyzed after high-pressure freezing and freeze-substitution and compared with conventional fixation followed by room temperature dehydration

Abstract

In this article, we report on the adaptation of high-pressure freezing and freeze-substitution (HPF-FS) for ultrastructural analysis of leaf tissue with special emphasis on chloroplasts. To replace the gas in the intercellular spaces, a mixture of water and methanol (MeOH) was employed. We compared three different supplements for FS--osmiumtetroxide, uranyl acetate, and safranin--with regard to the preservation of the ultrastructure of chloroplasts and other cellular compartments. The results show that (i) replacement of air within intercellular spaces by 8% (v/v) MeOH has no influence on the ultrastructure of the chloroplasts, (ii) undulation of membranes frequently observed after conventional preparation of specimens does not occur during chemical fixation but during room temperature dehydration, and (iii) uranyl acetate or osmium tetroxide employed during FS are not superior over safranin.

Architecture of Thylakoid Membrane Networks

“…It is therefore necessary to replace intercellular gases with an uncompressible liquid, or "space filler" under light vacuum prior to HPF. Usually hexadecane is used (Michel et al, 1991), but up to 20% dextran (Al-Amoudi et al, 2004 or 8% methanol in water (Pfeiffer and Krupinska, 2005) can also be utilized (see also review by McDonald, 2007).…”

Photosynthesis and ultrastructure of photosynthetic apparatus in tomato leaves under elevated temperature

“…It is usually considered that the changes of photosynthetic characteristics are consistent with the changes of chloroplast ultrastructure. For example, chloroplast membranes are ruptured and general loss of structural integrity follows (Pfeiffer et al 2005). Heat-induced changes of chlorophyll (Chl) fluorescence and the damage of the photosynthetic apparatus were reported (Xu et al 2006).…”

Amelioration of heat stress during reproductive stage in rice by melatonin

“…In contrast melatonin significantly increased heat induced reduction in quantum yield by 21.3% (0.64) in PB1121 and negligible in NL44. Heat stress disintegrates thylakoid membrane structure by disturbing lipid bilayer (Pfeiffer and Krupinska 2005) and oxidizes proteins of oxygen-evolving complex (OEC) (Dias and Lidon, 2009;Rexroth et al 2011), electron transport chain and dissociates the light-harvesting complex (Tang et al 2007, Xue et al 2011 which leads to decreased photosynthesis rate. Our result suggests that NL44 has acquired inbuilt mechanism overcome heat stress induced damage but PB1121 lacks this, which was supplemented by melatonin treatment and stabilized PSII activity.…”