Differing sperm ability to penetrate the oocyte in vivo and in vitro as revealed using colloidal preparations.

The penetration ability of boar (Sus scrofa domestica) spermatozoa exposed to viscous preparations under in vivo and in vitro fertilization conditions has been examined. Experiments involving induced ovulation in prepubertal animals and surgical insemination directly into the oviduct isthmus revealed an advantage of colloidal preparations. Based on within-animal comparisons, the incidence of penetration was 100% using both spermatozoa suspended in a viscous preparation of plant extracts and spermatozoa suspended in a control medium. However, percentages of monospermy were 22.2% in 54 oocytes inseminated with the control suspension compared with 62.5% in 48 oocytes inseminated with the colloidal preparation. An in vitro study involving 355 oocytes from slaughterhouse ovaries inseminated with in vitro-capacitated spermatozoa gave similar percentages of penetrated oocytes for both the control and colloidal suspensions. In this case, however, the percentage of monospermy was 32.7% in the control group compared with 10.6% for spermatozoa suspended in the colloidal preparation. Higher mean numbers of sperm inside the oocytes and higher numbers of sperm bound to the zona pellucida were also observed with the colloidal suspensions. In vitro motility and viability for spermatozoa in the colloidal suspensions were enhanced compared with that of the control group. Lower sperm membrane lipid disorder and reactive oxygen species generation were also observed in the viscous solution. These findings suggest that viscous fluids can enhance the ability of sperm to move, bind, and penetrate the oocyte in vitro, although this influence may be masked in vivo due to the already high viscosity in the oviductal fluid close to the time of ovulation.

Sodium pentobarbital versus alpha-chloralose anesthesia. Experimental production of substantially different slopes in the transmural CP/ATP ratios within the left ventricle of the canine myocardium.

BACKGROUND: Transmural analyses of the creatine phosphate (CP)/ATP ratio in various lamina of the canine myocardium have previously revealed significant variations in the CP/ATP ratio, with the subendocardial layer displaying a decreased ratio relative to the subepicardial layer. Without exception, these results were obtained under sodium pentobarbital anesthesia. These findings have been interpreted to imply that the normal endocardium may be operating in the oxygen-limited domain or that there are transmurally varying set points for the regulation of oxidative phosphorylation. METHODS AND RESULTS: In this work, we examine the effect of the anesthetic regimen on the transmural CP/ATP ratio within the left ventricular wall of the canine myocardium using spatially localized 31P-nuclear magnetic resonance (NMR) and an open-chest model. Two anesthetics were compared, alpha-chloralose and sodium pentobarbital. Under sodium pentobarbital, the CP/ATP ratio ranged from 1.92 +/- 0.06 to 2.51 +/- 0.08 from endocardium to epicardium, resulting in a transmural slope in the CP/ATP ratio of 0.149 +/- 0.047 (n = 22). Under alpha-chloralose, CP/ATP ratios ranged from 2.18 +/- 0.05 to 2.32 +/- 0.06, with a transmural slope of 0.035 +/- 0.018 (n = 38). Thus, the transmural slope in CP/ATP ratio was nearly four times greater with sodium pentobarbital than with alpha-chloralose, and the difference in these slopes was statistically significant (P = .029). No difference was observed in average CP/ATP obtained from the entire wall with either anesthetic. CONCLUSIONS: These results demonstrate that the transmural trend in CP/ATP ratio previously reported in the myocardium is likely to be a direct reflection of the sodium pentobarbital anesthetic regimen, not truly reflecting the trend in the normal unanesthetized animal. Moreover, since the transmural variation in CP/ATP ratio was greatly reduced with alpha-chloralose, it appears unlikely that the endocardium in the normal unanesthetized heart is operating in the oxygen-limited domain. These results also point to the importance of the anesthetic regimen in biochemical analysis, indicate the necessity of increased caution in directly translating results obtained under anesthesia, and demonstrate the unique power of in vivo NMR to extract such subtle biochemical information.

Transmural saturation transfer analysis of the creatine kinase system in the mammalian heart.

31P NMR spatial localization and saturation transfer techniques were combined to enable the transmural measurement of the forward creatine kinase (CK) rate (ATP:creatine N-phosphotransferase, EC 2.7.3.2.) in the in vivo canine myocardium. Five epicardial towards endocardial regions of the left ventricle (LV) were simultaneously examined using spatially localized voxels. Although intraleft ventricular CP/ATP ratios were constant, the pseudo first order rate constant (k') and the forward creatine kinase rate (Rf) displayed a 61% variation across the LV wall. Because CK levels and calculated [ADP], [CP] and pH are transmurally invariant in the normal left ventricle, the observed changes in the Rf could not be explained by changes in the absolute levels of these substrates and of creatine kinase. In addition, because myocardial oxygen consumption rates are known to be higher in the endocardium, these results imply that forward creatine kinase rates are not directly related to oxidative phosphorylation rates.