Ionic composition and osmolality of paddlefish (Polyodon spathula, Acipenseriformes) seminal fluid

Changes in ionic composition as Na⁺,K⁺, Ca²⁺ and Mg²⁺, osmolality inseminal fluid, percentage of motile spermatozoaand velocity were investigated in response toCPP and different dosage of LHRHa. The lowestvelocity of sperm was observed after use CPPtreatment. The velocity of spermatozoa,significant main effect of the treatment(P < 0.0001) and the time of sperm collection(P < 0.0104) were evaluated. The osmolality ofseminal fluid was different betweenexperimental groups of LHRHa (48.0–62.7mOsmol.kg^–1) and CPP (33.0–46.3mOsmol.kg^–1) treatments. The osmolalitywas significantly higher on the first day andone-half, then declined on day three, rangingfrom 33.0 to 62.7 mOsmol.kg^–1. Analysisof variance showed significant main effects ofthe treatment (P < 0.0001) and the time ofsperm collection (P < 0.0002) on the osmolalityof seminal fluid. The level of Na⁺ andK⁺ ion was different between experimentalgroups of LHRHa and CPP treatment. The highestconcentration of 11.11 mmol.l^–1 wasobserved at Na⁺ ion. Then theconcentrations declined on the level 1.56, 0.52and 0.36 mmol.l^–1 for K⁺, Ca²⁺and Mg²⁺ ions, respectively. There werehighly positive correlations between osmolalityof seminal fluid and dosage of LHRHa treatment(r = 0.84), velocity of spermatozoa andosmolality of seminal fluid (r = 0.57) andosmolality of seminal fluid and Na⁺concentration at seminal fluid (r = 0.70).Injection with LHRHa increased quality of spermas velocity of sperm, level of Na⁺,K⁺ and osmolality at seminal fluidcompared to CPP treatments.     

Kurokura Solution as Immobilizing Medium for Spermatozoa of Tench (Tinca tinca L.)

This study tested KUROKURA solution (Kurokura et al., 1984, Aquaculture 37, 267–274) and its modifications (by increasing NaCl content to 160, 180 and 200 mM) on immobilizing properties for sampling and short-term preservation of potential motility of tench spermatozoa. The immobilizing solution is used because, when collected, the sperm of most samples is contaminated by urine, causing spermatozoa to be of poor quality, with low motilities and velocities (almost 0), thus resulting in a worsened fertilization and hatching rate. Sperm was sampled with a syringe containing an immobilizing solution (IS), allowing an IS:sperm ratio of 2:1, under aerobic conditions at 0–4°C. This sperm solution was stored for 10 h and untreated sperm was collected prior to fertilization as a control. Spermatozoa quality was evaluated for the cell motility and velocity parameters and also for fertilization ability and hatching rate. Results obtained for tench sperm motility, velocity, fertilization and hatching rate showed that only sperm collected in the various immobilizing solutions can be successfully used for artificial insemination and preservation after 10 h at 0–4°C. The best immobilizing solution was found to be KUROKURA 180 (180 mM NaCl, 2.68 mM KCl, 1.36 mM CaCl₂· 2H₂O and 2.38 mM NaHCO₃), giving a fertility and hatching rate of 41%, with no change in rates after 10 h storage of sperm. Control sperm without immobilizing solution showed a fertilization and hatching rate of only 6–7%.     

The antioxidant system of sterlet seminal fluid in testes and Wolffian ducts

Oxidative stress is a possible source of spermatozoa function deterioration. Seminal fluid (SF) protects spermatozoa against reactive oxygen species (ROS) attack during development in testes and transit through the reproductive tract. Spermatozoa curvilinear velocity and percent of motile cells as well as changes in thiobarbituric acid-reactive substance (TBARS) content, superoxide dismutase, and catalase activity, and uric acid concentration in SF were evaluated in sterlet sperm collected from testes 24 h after hormone induction of spermiation and from Wolffian ducts at 12, 24, 36, and 60 h after hormone injection (HI). While testicular spermatozoa motility was not initiated in activating medium, Wolffian duct sperm showed low motility at 12 h, significant increase at 24 and 36 h, and decrease at 60 h. Testicular SF was characterized by the highest level of TBARS and activity of studied enzymes compared with SF from Wolffian duct sperm at 24 h post-HI. In fluid from Wolffian duct sperm, a significant increase in TBARS content was shown at 36–60 h post-HI. In contrast to testicular SF, in SF from Wolffian duct sperm, this increase was not counterbalanced by changes in the studied variables of antioxidant system. This may be the source of the observed decrease in spermatozoa motility parameters 60 h post-HI. The results may confirm a dual role of ROS in fish sperm physiology. The data with respect to decrease in sturgeon spermatozoa motility parameters at 60 h post-HI should be taken into account in artificial sturgeon propagation.     

Metallothioneins and detoxification. Is the use of detoxication protein for mts a language abuse?

Our own results and a literature review led us to reconsider the detoxifying function of MTs in living organisms. Despite the fact that many authors have observed a synthesis or a level increase of MTs as a response to toxic metal uptake, arguments exist which tend us to give to MTs a strict zinc homeostasic function. Many experiments have been conducted using non-natural routes of exposure and/or concentrations far from those observed even in heavily polluted environments. Zinc is the only metal for which a primary induction has been established. Correlations between zinc and MTs levels are frequently observed, specially at the early stages of life. Our knowledge about metallic cluster structure and our experimental results about inter-metallic competition for binding-sites on the apoprotein, support the idea of substitution processes instead of de novo synthesis in most cases of contamination, leading frequently to acclimation.     

A 40 years journey with fish spermatozoa as companions as I personally experienced it

When, in the 1980s, I became interested in the spermatology of fish under the light microscope, active spermatozoa were only visible thanks to their head presenting a sort of “tremor.” This situation was quite frustrating given the lack of possible information regarding the motor part called flagellum. We decided to apply simple technologies, including photography. Due to the high speed of the moving fish flagellum, the microscope illumination used a pulsed light strobe combined with a dark field microscope to record the flagellum image despite its small diameter (< 0.5 μm). Then came high-speed cinematographic microscopy up to 200 fps, as well as video cameras. At the end of the 1990s, an automatic moving object video tracking system began to be commercialized (CASA) with main advantages such as (a) a large number of cells tracked, which greatly improves statistics, (b) computer assistance allowing an automatic analysis that provides many motility parameters. Nevertheless, CASA systems are still unable to provide information about fish sperm flagella that move fast. During the 1990s, analog video camera technologies allowed acquisition of flagellum images with high resolution for detailed analysis. Since the 2000s, the use of high-speed video cameras allows the acquisition of images at a much higher resolution and frequency, up to 10,000 frames per second. Since it became possible to visualize the flagella in motion, a noble function was added to that of a propeller: that of a rudder with what a spermatozoon responds to specific signals delivered by the egg for its guidance. In the future, one can wish that an automatic flagella movement analyzer will become functional. This brief anthology puts forward the large amount of progress accomplished during past 40-year period about spermatozoa movement analysis, especially in fish.

     

The role of Ca2+ and Na+ membrane transport in brook trout (Salvelinus fontinalis) spermatozoa motility

The role of environmental ion composition and osmolality in Ca²⁺ signaled activation was assessed in spermatozoa of brook trout Salvelinus fontinalis. Milt from ten mature males was obtained by abdominal massage. Spermatozoa motility was evaluated in 0, 100, and 300 mOsm/kg NaCl or sucrose solutions, buffered by 10 mM Tris–HCl pH 8.5. For investigation of spermatozoa reaction to external Ca²⁺ concentration, 2 mM ethylene glycol tetraacetic acid (EGTA) was added to the activation media as a calcium ions chelator. For investigation of the effect of external Na⁺ concentration in conditions of low external Ca²⁺, 100 µM amiloride was added to the EGTA-containing solutions as a Na⁺ transport blocker. Low motility was observed in sucrose (Na⁺ free) solutions containing 2 mM EGTA but not in Na⁺ solutions containing 2 mM EGTA. Addition of amiloride led to significantly increased motility (P < 0.05) compared with sucrose (Na⁺ free) solutions containing 2 mM EGTA. We conclude that Na⁺ transport in Ca²⁺-free solutions plays a regulatory role in brook trout spermatozoa activation. The influence of competitive Na⁺ and Ca²⁺ transport on the control of spermatozoa activation requires further study with respect to its application for improvement of artificial activation and storage media.