Summary. Spermatozoa from the corpus epididymidis of boars and rams showed head-to-head agglutination when diluted. The occurrence of agglutination coincided with the appearance of motility but preceded the ability to bind to zona-free hamster eggs. Head-to-head agglutination was inhibited by the addition of caudal epididymal plasma. A protein acting as an antagglutinin on spermatozoa from the corpus epididymidis was extracted from cauda epididymal plasma and partly purified.
J. L. Dacheux, M. Paquignon and Y. Combarnous
J-L. Gatti, C. Chevrier, M. Paquignon and J-L. Dacheux
Internal pH and motility of testicular, epididymal and ejaculated ram and boar spermatozoa were studied as a function of external ionic composition. Internal pH was estimated by the amine distribution method and motility was characterized by percentage of cells that were motile and flagellar beat frequency. Upon dilution in media at different external pH values, internal pH of boar and ram spermatozoa changed rapidly towards the external pH. High external concentrations of Na+ or K+ had no effect on the rate of equilibration and only a slight effect on the final internal pH value, ruling out a role of Na+–H+ or K+–H+ exchange mechanisms in this process. In both species, a linear relationship was observed between internal and external pH but equilibration was incomplete suggesting that there is a complex regulatory mechanism. This result was unaffected by epididymal maturation and ejaculation. Ram and boar testicular spermatozoa showed no increase in movement after dilution, suggesting that simple changes in internal pH are not a sufficient trigger for motility. At high external pH, internal pH increased and motility of epididymal boar spermatozoa was initiated. Motility of ejaculated boar spermatozoa, and epididymal and ejaculated ram spermatozoa was less dependent upon external pH and affected only very slightly by the internal pH changes. K+ or Na+ had almost no effect on motility just after dilution. After 1 h of incubation, movement decreased. Maintenance of motility in sodium or potassium showed a sharp external pH optimum. Media without Na+ and K+ allowed a better conservation of motility at external pH > 8 for ram epididymal and ejaculated spermatozoa and at external pH > 6 for boar ejaculated spermatozoa.
J. L. Courtens, H. Ekwall, M. Paquignon and L. Plöen
Summary. Boar semen was analysed by electron microscopy coupled to image analysis and X-ray energy dispersive spectroscopy, during the usual process for freezing and thawing in field conditions. Freeze–substitution and freeze–quenching permitted recording of real or potential intracellular ice before, during, and after freezing. Heads and flagella displayed two different osmotic properties before freezing. Heads were dehydrated progressively before and during freezing, while flagella were hydrated before freezing and were only dehydrated during freezing. All parts of the thawed cells were rehydrated. Ice crystal damage was mostly present in frozen mitochondria and axonemes and the acrosomes were strongly affected by thawing. The total amounts of Na, Cl, Ca, K, Mg, and Zn per cell were only elevated in frozen and thawed midpieces while the heads were permeable both to water and elements at that time.
Keywords: boar; spermatozoa; freezing; water; element concentrations; electron microscopy; image analysis; X-ray spectrophotometry
T. O'Shea, J.-L. Dacheux and M. Paquignon
Summary. Boar spermatozoa incorporated more [14C]glycerol into lipid when incubated with 200 mm- than with 25 mm-glycerol. Measurements were made of the metabolism of spermatozoa while they were being prepared for frozen storage. [14C]Glucose was converted to CO2 and lipid while the cells were cooling to 15°C. Glycerol was added at 15°C and during further cooling to 5°C glucose metabolism was greatly reduced but [14C]glycerol was converted to CO2 and lipid. Under aerobic conditions spermatozoa accumulated lactate while cooling from 30 to 15°C and from 15 to 5°C. With essentially anaerobic conditions, although more lactate was accumulated this occurred only while the cells were cooling from 30 to 15°C, and no further accumulation could be detected during cooling from 15 to 5°C. When boar spermatozoa were incubated at 37°C after storage in liquid nitrogen, metabolism of glycerol was greater than metabolism of glucose. It is suggested that this preferential use of glycerol during cooling and after storage may be one facet of its cryoprotective function. After storage, boar spermatozoa incorporated relatively less [14C]stearic and [14C]palmitic acids into phospholipids (especially phosphatidyl choline) than did freshly collected cells. Caffeine stimulated the oxygen uptake of freshly collected and thawed cells.
J. L. Dacheux, T. O'Shea and M. Paquignon
Summary. Spermatozoa were collected from the rete testis of conscious boars, from the cauda epididymidis by retro-flushing, and by ejaculation. Testicular spermatozoa showed no progressive motility, and that of ejaculated was greater than that of epididymal spermatozoa. Glycolysis and respiration of testicular spermatozoa, while lower than that of the more mature cells, were only slightly affected by the incubation conditions. Epididymal spermatozoa converted 83% of the glucose they utilized to CO2 or lactate, but testicular cells converted only 35% to these metabolites. Synthesis of lipid was greatest by testicular spermatozoa. With the more mature cells hyperosmolar conditions depressed CO2 production, but increased lactate production, and these changes were greater for ejaculated than for epididymal spermatozoa. Glycolysis plus respiration of these cells was related to their motility. These results were interpreted as showing increasing motility, glycolysis and respiration with maturation, but also decreased synthetic capacity and increased sensitivity to the environment.
C. Jeulin, J. C. Soufir, J. Marson, M. Paquignon and J. L. Dacheux
Summary. In the epididymal fluid of boars, the concentration of carnitine (nmol/mg protein) began to increase from 20 in the distal caput, then rose progressively to 700 in the distal cauda. By contrast, the carnitine content of spermatozoa only started to increase in the proximal cauda where the concentration of carnitine in the fluid was 200–300 nmol/mg protein then gradually increased in spermatozoa from more distal sites. The increase in the acetylcarnitine content of spermatozoa paralleled that of the carnitine amount and represented 50% of the total carnitine (carnitine + acetylcarnitine). We conclude that the acetylcarnitine content of epididymal spermatozoa may be used as a marker of maturation.