Visualising the premature brain using 17.6 Tesla magnetic resonance imaging

The aim of this study was to evaluate the ability of a 17.6 Tesla magnetic resonance (MR) microscope to determine external and internal structures and three-dimensional (3D) volume rendering of premature bovine brain tissue. Two bovine embryos (Carnegie-stages 16 and 21) were examined. 3D magnetic resonance imaging (MRI) was performed with a high field MR-scanner at a field strength of 17.6 Tesla. Images with isotropic nominal resolutions up to 39.1 μm were acquired. The MR images corresponded very well with histological slices. 3D virtual models of the embryonic brain were easily produced in a relatively short time and the high field scanner provided highly detailed images of formalin fixed brain tissue. Manual segmentation and automatic volume rendering is a valuable tool for the fast generation of 3D brain models and, to some degree, can replace conventional techniques in comparative embryology.     

Influence of Staining Method on the Values of Avian Sperm Head Morphometric Variables

Computer‐assisted systems for the assessment of sperm morphometry (ASMA systems) have been used successfully with several mammalian species. Unfortunately, they have so far been of little use for assessing bird semen, a consequence of the filiform shape of avian spermatozoa. This study compares two staining techniques (Hemacolor^® and aniline blue staining) for the morphometric analysis of rooster and red‐legged partridge spermatozoa as part of a computer‐assisted light microscopy method. For both species, Hemacolor^® staining provided a significantly higher percentage of measurable cells (93.7 ± 11.7% in roosters and 71.9 ± 15.3% in red‐legged partridges). Hemacolor^® also showed greater repeatability (lower coefficients of variation) for length and area in roosters' sperm and for width in the case of red‐legged partridge's sperm. In the roosters, the Hemacolor^® technique returned significantly (p < 0.05) larger sperm head width and area values than did the aniline blue technique, while the latter resulted in greater sperm head length values (p < 0.05). In the red‐legged partridge, no differences were seen in the results for sperm head width and area provided by the two techniques, but aniline blue staining was associated with longer length measurements. In conclusion, the morphometric values recorded differed depending on the staining method and species. However, the Hemacolor^® technique might be deemed the more appropriate for computerized sperm assessment systems as it provides larger percentages of measureable cells and shows greater repeatability.     

Comparison of Membrane‐Permeant Fluorescent Probes for Sperm Viability Assessment in the Ram

This study was designed to compare the effectiveness and usability of four permeant fluorochromes (CFDA; SYBR‐14; Hoechst‐33342; and acridine orange), combined with propidium iodide to assess sperm membrane integrity. Three different experiments were conducted. The first trial was designed to study the optimal dye concentration and minimum incubation time required to achieve optimum fluorescence intensities and contrast for each fluorochrome combination using ram fresh semen samples. Both SYBR‐14 and acridine orange allowed a direct assessment of sperm membrane integrity, without the need of incubating samples, whereas a minimum of 4 and 6 min of incubation at 37°C was necessary to achieve optimum fluorescence intensities in the CFDA and Hoechst groups, respectively. In the second trial, fresh semen samples were mixed with different volumes of membrane‐affected sperm (semen treated with three cycles of freezing to ?20°C and thawing at room temperature) to produce semen samples with known proportions of damaged spermatozoa. The results were compared with the theoretical values predicted on the basis of the estimations made on fresh and frozen samples. The proportions of damaged sperm in each sample determined using the four fluorochrome combinations agreed with the predicted theoretical values, with the acridine orange/propidium iodide providing the best adjustment. The third experiment was performed to compare the results of sperm membrane integrity using the four fluorochrome combinations. The proportions of plasmalemma‐intact sperm determined by acridine orange and SYBR‐14 were greater (p < 0.0001) than the proportions of intact sperm determined by CFDA and Hoechst stains. It was concluded that the most efficient combinations to be used in ram sperm were AO/PI and SYBR/PI because it allowed a direct assessment of sperm viability without the need to incubate samples and obtaining reliable results.     

In vitro Capacitation and Acrosome Reaction of Dog Spermatozoa can be Feasibly Attained in a Defined Medium Without Glucose

Incubation of dog spermatozoa in a medium without glucose and in the presence of lactate and pyruvate (l-CCM) for 4 h at 38.5 degrees C in a 5% CO(2) atmosphere induced in vitro capacitation of these cells. This was verified after the combined specific capacitation-like changes in percentages of viability and altered acrosomes, motility characteristics, sperm location of reactivity against Pisum sativum, Arachis hypogaea and Helix pomatia lectins and the tyrosine phosphorylation pattern. Furthermore, a feasible acrosome reaction (AR) was induced when spermatozoa incubated in l-CCM for 4 h were further co-incubated for 1 h with canine oocytes. This was demonstrated by AR-like changes in percentages of viability, altered acrosomes, motility characteristics and sperm location of reactivity against P. sativum, A. hypogaea and H. pomatia lectins. All these results clearly indicate that in vitro capacitation, and subsequent AR, can be feasibly achieved without the presence of sugars. This ability can be related to the specific characteristics of energy-metabolism regulation reported in dog spermatozoa.     

Local Effect of the Corpus Luteum on Ovarian Follicular Functional and Morphological Features in the Goat

Ovaries of unilaterally ovulated goats (n=21) were used to study follicular morphological features (proportion and degree of atresia, oestradiol production in vitro and progesterone production of granulosa cells in culture). Follicles were dissected out and classified as small (1–2 mm), medium (2–4 mm) and large (>4 mm). Morphological and physiological features were compared in each size class between ovaries bearing and not bearing corpora lutea (CLO and NCLO, respectively). Within the same size class, there was no difference in proportion or in degree of atresia, between CLO and NCLO. A significant effect of follicular size on oestradiol production in vitro was detected, but no effect of the corpus luteum was found. Finally, progesterone production of granulosa cells in culture was significantly higher in CLO than in NCLO after 24 h (p < 0.05) and also after 90 h (p < 0.01) of culture. This higher progesterone production by CLO granulosa cells in culture could be explained by local influence of the corpus luteum stimulating the steroidogenic activity but not aromatase activity. Further studies are needed to clarify possible factors and pathways for this local effect of the corpus luteum upon follicular physiology.     

Genetic control of some polymeric glutenin fractions in hexaploid wheat (Triticum aestivum)

Reciprocal crosses were made between seven different hexaploid wheat genotypes. Hybrid kernels and their parents were used to determine the amount of polymeric glutenin fractions by size-exclusion high-performance liquid chromotography (SE-HPLC) analysis. Quantitative aspects of the genetic control for various glutenin fractions were investigated through diallel cross analysis. The association between the potential effect on hexaploid wheat quality of various allelic types and the quantitative expression of the polymeric glutenin fractions was confirmed. Significant average heterosis effects were demonstrated for insoluble glutenin (pFi), total soluble and insoluble glutenin (pF1 + pF2 + pFi) and the ratio of soluble to insoluble glutenin, (pF1 + pF2)/pFi. Some genotypes showed significant positive or negative combining abilities and general reciprocal effects for the glutenin fractions studied. For gluten quality, ‘Qualital’ was the best combiner for determining an optimal glutenin composition (high values for pFi and pF1 + pF2 + pFi and low values for pF1/pF2 and (pF1 + pF2)/pFi, respectively). These results should be of great interest in breeding programmes aimed at improving hexaploid wheat quality.     

Impacts of Irrigation with Water Containing Heavy Metals on Soil and Groundwater – A Simulation Study

This research work intended to study the impacts of irrigation water containing various levels of copper, lead, and zinc on adsorption capacity of soil packed in 4′′ plastic columns and obtained from two locations around the city of Nablus: Salem (A) and Deir Sharaf (B). Results of simulation experiments showed an increase in the copper, lead, and zinc concentrations in soil and in leachate with increasing the amount of metal in irrigation water. Copper, lead, and zinc concentrations increased also with soil depth and duration of application. The results also indicate that the self purification of both soils was highly affected by physical factors, i.e. the intermittent application of irrigation water to the soils in the columns caused soil wetting and drying cycles whichresulted in the formation of cracks in shrinked soils specially in the top half of the columns. Crack formation is common in such clay soils due to the climatic conditions (Mediterranean type: dry summers and wet winters) and type of clay minerals in the soil. Thus, short circuiting of water through cracks results in moving contaminants fast and deep in the soil profile.