Optimizing Electrical Activation of Porcine Oocytes by Adjusting Pre‐ and Post‐Activation Mannitol Exposure Times

Modifying electrical activation conditions have been used to improve in vitro embryo production and development in pigs. However, there is insufficient information about correlations of porcine embryo development with oocyte pre‐ and post‐activation conditions. The purpose of this study was to compare the developmental rates of porcine oocytes subjected to different mannitol exposure times, either pre‐ or post‐electrical activation, and to elucidate the reason for the optimal mannitol exposure time. Mannitol exposure times around activation were adjusted as 0, 1, 2 or 3 min. Blastocyst development were checked on day 7. Exposure of oocytes to mannitol for 1 or 2 min before electrical activation produced significantly higher blastocyst rates than exposure for 0 or 3 min. There was no significant difference in blastocyst rates when activated oocytes were exposed to mannitol for 0, 1, 2 or 3 min after electrical activation. While exposure of oocytes to mannitol for 1 min pre‐ and 3 min post‐activation showed significantly higher blastocyst development than 0 min pre‐ and 0 min post‐activation. It also showed higher maintenance of normal oocyte morphology than exposure for 0 min pre‐ and 0 min post‐activation. In conclusion, exposure of oocytes to mannitol for 1 min pre‐ and 3 min post‐activation seems to be optimal for producing higher in vitro blastocyst development of porcine parthenogenetic embryos. The higher blastocyst development is correlated with higher maintenance of normal morphology in oocytes exposed to mannitol for 1 min pre‐ and 3 min post‐activation.     

Comparison of Conventional Freezing and Vitrification with Dimethylformamide and Ethylene Glycol for Cryopreservation of Ovine Embryos

The aim of this work was to evaluate the efficiency of the cryoprotectants dimethylformamide and ethylene glycol for cryopreservation of ovine embryos using vitrification and conventional freezing. The recovered embryos were distributed randomly in three treatment groups: Gr. 1: conventional freezing (n = 44), Gr. 2: vitrification with ethylene glycol (n = 39) and Gr. 3: vitrification with dimethylformamide (n = 38). Quality of fresh embryos in control group as well as of frozen and vitrified embryos was examined by three methodologies: staining with propidium iodide and Hoechst 33258 and evaluation under fluorescent microscopy, evaluation of re‐expansion and hatching rates after culture, and determination of apoptotic index with TUNEL technique. It was established that re‐expansion rate in all treatment groups was similar. In the same time, hatching rates were higher in Gr. 1 (40.5%) and Gr. 2 (35.3%) in comparison with Gr. 3 (15.5%, p < 0.05). The number of dead cells in vitrified embryos of Gr. 2 and Gr. 3 was higher (42.6 ± 26.2 and 63.2 ± 34.65, respectively) in comparison with Gr. 1 (conventional freezing, 10.1 ± 8.5, p < 0.05). Embryos vitrified with dimethylformamide included the same quality of apoptotic cells that Gr. 1 (conventional freezing) and fresh embryos. In conclusion, the dimethylformamide and ethylene glycol used as cryoprotectant to vitrify ovine embryos, in the concentrations and exposition time tested in this work, were not as efficient as the conventional freezing for cryopreservation of ovine embryos Thus, the conventional freezing with ethylene glycol was the most efficient method to cryopreserve ovine embryos in comparison with vitrification.     

Landscape services: the concept and its practical relevance

Recently, in addition to the popular concept of “ecosystem services” (ES), the term “landscape services” (LS) has come into use. We are examining the question of whether a stronger focus on LS would be useful, particularly with regard to case studies carried out in Germany. Important reasons for introducing the term LS include the prominent role of spatial aspects, the reference to landscape elements and the landscape character, and the relevance of LS for landscape planning. We found no strong arguments for replacing the concept of ES by LS; however, we do prefer a situation-related use of both concepts. We propose the following definition: Landscape services are the contributions of landscapes and landscape elements to human well-being.     

Quantification of Whole Blood Short-chain Fatty Acids by Gas Chromatographic Determination of Plasma 2-chloroethyl Derivatives and Correction for Dilution Space in Erythrocytes

A precise, accurate and stable method for quantification of short-chain fatty acids (SCFA) in ovine whole blood is presented and validated. The method is based on esterification of plasma SCFA by reaction with chloroethyl chloroformate in a water/acetonitrile/2-chloroethanol solution and gas chromatographic (GC) analysis of the derivatives. Whole blood concentrations of SCFA could be obtained by correcting plasma concentrations for a 45% dilution space of SCFA in the erythrocyte fraction of the blood. The recovery of SCFA in plasma and whole blood was 96-100% independent of the haematocrit value when compared with water standards. The method avoided carry-over from sample to sample, contrary to earlier published methods. The average intra-assay and inter-assay coefficient of variation for repeated measurement of SCFA content in plasma samples was 2.5% and 3.1%, respectively. The derivative was found to be suitable for a precise and accurate determination of the ¹³     

Proton nuclear magnetic resonance spectroscopy based investigation on propylene glycol toxicosis in a Holstein cow

Background

It is unknown which metabolites are responsible for propylene glycol (PG)-induced toxicosis, and a better understanding of the underlying mechanisms explaining incidences of abnormal behaviour of dairy cows fed PG is therefore needed.

Methods

The study included three cows of which one developed PG toxicosis. In order to investigate how the metabolism of PG differed in the cow developing toxicosis, proton nuclear magnetic resonance (NMR) spectroscopy was applied on ruminal fluids and blood plasma samples obtained before and after feeding with PG.

Results

PG toxicosis was characterized by dyspnea and ruminal atony upon intake of concentrate containing PG. The oxygen saturation of arterial blood haemoglobin and the oxygen pressure in arterial blood decreased along with the appearance of the clinical symptoms. NMR revealed differences in plasma and ruminal content of several metabolites between the cow responding abnormally to PG and the two control cows.

Conclusion

It is concluded that PG-toxicosis is likely caused by pulmonary vasoconstriction, but no unusual metabolites directly related to induction of this condition could be detected in the plasma or the ruminal fluid.