Sublethal damage during cryopreservation of rainbow trout sperm

Although cellular damage during cryopreservation of freshwater fish spermatozoa has been reported in several studies, there is a lack of correlation between this damage and the fertility rates of eggs using postthawed milt. The apparent lack of such correlation may be due to other undetected sublethal cryodamage, which could affect the cell functionality and viability. This may be extremely important for freshwater fish spermatozoa whose ability to fertilize the egg requires dilution in water or hypoosmotic solutions, an hazardous environment for the cells. This study tested the change in cell permeability during cryopreservation, using Hoechst 33258 to assess cell permeability. The permeability of spermatozoa at different times after dilution in several hypoosmotic media were investigated. In the first trial, fresh semen, sperm diluted in freezing media (CPT), and freeze/thawed semen were studied. Three CPT were tested (Me2SO, DMA, and methanol). In the second trial, the addition of egg yolk as a membrane stabilizer was investigated. Samples were frozen at -20 degreesC/min in a programmable cooler and thawed in a 25 degreesC water bath. Dilution in the CPTs slightly increased the susceptibility of cells to damage but freezing/thawing caused a dramatic increase in the fragility of cells, which were killed in a few seconds after their contact with the hypoosmotic solutions. Egg yolk provided a significant protection to the membrane, allowing the cells a greater and more prolonged survival in the fertilization media. Samples frozen with Me2SO displayed the best results. These results are consistent with the achieved fertility rates that demonstrated sublethal cryodamage in the function of the sperm membrane that was not detected by standard procedures. Copyright 1998 Academic Press.     

The role of facilitated diffusion of oxygen in tissue hypoxia

The evidence that cytochrome P-450 can act as a tissue oxygen carrier is outlined, and a new experimental approach to the measurement of the fraction of oxygen carried this way is described. This fraction is increased when cytochrome P-450 concentration is increased, which occurs on exposure of the experimental animal to hypoxia. This appears to be a new mechanism of tissue acclimation to hypoxia.Presented at the Seventh International Biometeorological Congress, 17–23 August 1975, College Park, Maryland, USA.     

Use of an encapsulated fluorescent probe to measure intracellular PO2

The objectives of this investigation were to produce a reliable, senstive probe to measure intracellular P_O2 with a high degree of resolution and to apply this technique to biological systems. A fluorescent molecule, pyrene dissolved in paraffin oil, was encapsulated in polyacrylamide to form a probe of nanometer dimensions. The quantitative and microscopic oxygen values were determined by analyzing the quenching of the fluorescence of the probe by oxygen, as displayed on a television monitor by a silicon-intensified-target camera. The nanocapsules had a sensitivity of approximately 1 mm P_O2, a spatial resolution of 0.5 μm, and a temporal resolution of milliseconds. Calibrated nanocapsules within nonrespiring Amoeba proteus responded to ambient partial pressures of oxygen. At two different ambient partial pressures, nanocapsules engulfed by respiring amoebas indicated an intracellular P_O2 28 mm Hg less than extracellular P_O2. The capsules retained their sensitivity to oxygen for at least 8 months.     

The Design and Analysis of Twin Animal Experiments

The experimental design used in twin studies on veal calves at the University of Guelph during the years 1965–1968 combined split trials with uniformity trials on each treatment. Each replicate consisted of four twin pairs. Both members of one pair selected at random were put on treatment 1, both members of another pair were put on treatment 2, and the remaining two pairs were “split” with one twin from each pair randomly assigned to each treatment. There were a number of replicates of this basic design over the course of the experiment. The objective of this paper is: (i) to determine the feasibility of an overall analysis of the given design, (ii) to determine an accurate method for assessing treatment effects, and (iii) to find an efficient method for detecting the presence of treatment Xpair interaction.