Download Hadiseh Dadras: The in vitro effect of various temperatures on

Hadiseh Dadras: The in vitro effect of various temperatures
on motility, membrane lipid composition, morphology and antioxidant enzymes activity of
fish spermatozoon
Sperm motility in many fish species is influenced by several external factors such as
temperature, but, so far, temperature has been very rarely assessed in studies of
fertilization. It has also been proven that motility duration and velocity of spermatozoa
depend on temperature of the activation medium. Assuming that fish spermatozoa contain
finite energy stores, the existence of temperature mediated sperm longevity and a trade-off
between sperm activity and longevity clearly indicate that sperm swimming activity, as any
metabolic activity, will be directly influenced by temperature. Therefore, if changes in water
temperature affect the swimming velocity of sperm, such an effect would directly translate
into corresponding changes in fertilization rate.
In addition, temperature is known to influence the phase disposition of lipids in cell
membranes. Change in temperature shows an impact on membranes, particularly on the
formation of lipid clusters that break down the lipid bilayer selective permeability.
Motility characteristics are directly related to spermatozoon morphology and ultrastructure.
Spermatozoa morphology and ultrastructure can also be an indicator of male fertility when
fish are exposed to different environmental factors. Fish spermatozoa demonstrate extreme
sensitivity to minute changes in temperature of surrounding media.
In fish sperm, the primary enzymes for detoxification of reactive oxygen species (ROS) are
catalase (CAT) and superoxide dismutase (SOD); hence their activity will reveal oxidative
status. Investigation of oxidative stress indicators during motility at different temperatures
can provide information on the effects of temperature on spermatozoon metabolism and
help determine whether spermatozoon motility and fertilization capacity are influenced by
water temperature.
During the present study, sperm of six carp (Cyprinus carpio) males will be used in separate
experiments. The dry sperm will be diluted with activation medium. Sperm motility
parameters will be recorded using a CCD video camera mounted on an inverted microscope
equipped with a cooling stage. Sperm Motility parameters will be assessed by computer
assissted computer analysis (CASA) system. Cholesterol will measure with the CHOD PAP Kit
(Biolabo, France). Total phospholipid (PL) will be quantified by the procedure developed by
Rouser et al. (1970). Major lipid classes will be separated according to (Olsen and
Henderson, 1989), using the total lipid fraction with a ADC 2 system (Camag Switzerland)
using first a mobile phase consisting of Hexan, Diethylether and acetic acid, (85,15, 2);
(v/v/v). The activity of SOD and CAT will be measured by a spectrophotometer equipped
with a thermo-regulated cuvette holder. The fresh, unfixed material dropped onto
microscopic copper grids will be used for the observation in ESEM (Quanta 200 FEG).