Alteration of the lipid phase transition during mouse embryos freezing after in vitro culture with linoleic acid.

Lipids significantly affect embryo cryopreservation in some mammalian species depending on the cell lipidome quantity and composition. One of the ways to study the relationship between lipid content and cryotolerance of cells is to study the effect of lipidome modification on laboratory mice. The objective of this research was to study how in vitro culture of mouse embryos with linoleic acid (LA) will affect lipid phase transition (LPT) during cooling and subsequent embryo development after cryopreservation. Embryos obtained in vivo at the 2-cell stage were cultured with 200 µM LA for 46 h up to the morula/blastocyst stage. Thereafter, one portion of embryos was slowly frozen to reveal the effect of LA on survival after cryopreservation, another portion was used to characterize the lipid composition and to determine the temperature of the LPT onset. Confocal fluorescence microscopy of Nile Red stained embryos showed a significant increase in lipid content of the LA treated group compared to the controls. Raman measurements showed that the onset of LPT in LA treated embryos is lower than in untreated ones: -5 °C vs +2 °C. However, these changes in the LPT onset did not affect the survival rates of embryos after cryopreservation. In summary, in vitro culture with LA changes the biophysical characteristics of embryos' lipidome and is realized in lower LPT onset, but this does not affect embryo survival after cryopreservation.

Deuterated stearic acid uptake and accumulation in lipid droplets of cat oocytes.

Fatty acid uptake and accumulation in lipid droplets are essential processes of lipid metabolism. Oocyte in vitro culture in media enriched with fatty acid is used to modify the lipid content and composition, aiming to study the consequences of obesity and enhance cell cryotolerance. We applied Raman spectroscopy and deuterium labeling approach to quantify stearic acid uptake and investigate its incorporation within oocytes. Our data suggest that deuterium labeling does not affect oocyte maturation rates. The efficiency of deuterated stearic acid (dSA) uptake was shown to decrease with the increase of its concentration in culture medium and the duration of in vitro culture. The molar ratio between dSA and bovine serum albumin has no significant effect on the dSA uptake for 200 µM but modifies concentration dependence of the lipid uptake. dSA accumulates in all the lipid droplets inside oocytes. Different lipid droplets within the same oocyte exhibit different concentrations of dSA. The scatter in the dSA concentration in lipid droplets decreases with the culture time. Using dSA as an example, we provide a comprehensive description of how fatty acid concentration, its molar ratio versus bovine serum albumin, and culture time affect the uptake of the fatty acids in oocytes. Raman microspectroscopy of deuterium-labeled fatty acids is a nondestructive tool providing information about fatty acid uptake and heterogeneity of their accumulation between lipid droplets within the single oocyte.

[Effects of a high-fat diet on the lipid profile of oocytes in mice]. / Влияние диеты с повышенным содержанием жира на липидный профиль ооцитов мышей.

There are evidences that obese women exhibit a detrimental oocyte quality. However, it remains unclear how this change is associated with obesity, indirectly - or directly through a change in the content and/or composition of lipids in oocytes. The aim of this work was to study effects of a high-fat diet applied to female donor mice on the amount and qualitative composition of lipids of immature and in vivo matured oocytes. A high-fat diet caused larger body weight in female mice compared with the control ( p < 0.001; 44.77 ± 1.46 and 35.22 ± 1.57, respectively), and increased the blood levels of cholesterol ( p < 0.05; 2.06 ± 0.10 and 1.78 ± 0.10, respectively) and triglycerides ( p < 0.05; 2.13 ± 0.23 and 1.49 ± 0.21, respectively). At the same time, this diet does not affect the level of unsaturation of lipids in immature (0.207 ± 0.004 in the experiment and 0.206 ± 0.002 in the control) and matured oocytes (0.212 ± 0.005 in the experiment and 0.211 ± 0.003 in the control). Total lipid content increased during in vivo maturation of mouse oocytes. The amount of lipids was greater in mature oocytes in the experimental group compared to the control ( p < 0.01; 8.15 ± 0.37 and 5.83 ± 0.14, respectively). An increase in intracellular lipid amount during oocyte maturation was revealed both after a standard diet ( p < 0.05; 4.72 ± 0.48 and 5.83 ± 0.14, respectively) and after a fat-rich diet ( p < 0.001; 3.45 ± 0.62 and 8.15 ± 0.37, respectively). Thus, during in vivo oocyte maturation in mice the content of intracellular lipids enhanced, the high-fat diet aggravated this dynamics of lipid increase during in vivo maturation of oocytes.

Features of emotional and social behavioral phenotypes of calsyntenin2 knockout mice.

Calsyntenin-2 (Clstn2) is the synaptic protein that belongs to the super family of cadherins, playing an important role in learning and memory. We recently reported that Clstn2 knockout mice (Clstn2-KO) have a deficit of GABAergic interneurons coupled with hyperactivity and deficient spatial memory. Given, that impaired functioning of GABA receptors is linked to several psychopathologies, including anxiety and autism, we sought to further characterize Clstn2-KO mice with respect to emotional and social behavior. Clstn2-KO males and females were tested in the elevated plus-maze (EPM), open field (OF), forced swim test, social affiliation and recognition test, social transmission of food preference (STFP), dyadic social interactions and marble burying test. Clstn2-KO mice demonstrated high exploration and hyperactivity in the dimly lit EPM that affect anxiety parameters. In contrast, in a more adverse situation in the OF have increased emotionality in Clstn2-KO males, not females. Assessment of hyperactivity for prolong period in the OF showed that Clstn2-KO animals were able to decline their hyperactivity, but their ambulation still remained higher than in WT littermates. Additionally, Clstn2-KO mice expressed stereotyped behavior. Strikingly, analysis of social behavior identified deficient social motivation and social recognition only in Clstn2-KO males, but not in females. Further analysis of social communication in the STFP and direct observation of agonistic interactions confirmed the reduced social behavior in Clstn2-KO males. Altogether, current results showed Clstn2 gene and sex interactions on socio-emotional performance in mice, suggesting a possible role of calsyntenin2 in psychopathological mechanisms of autism.