Correlation between chemical composition of seminal plasma and sperm motility characteristics of Prussian carp ( Carassius gibelio )

Abstract

The objectives of the present study were to determine the relationships between chemicalscompositions of seminal plasma with sperm motility traits in Prussian carp, Carassius gibelio (Bloch,1782). There were significant positive correlations between sperm movment duration and Ca+2 of semen.Also, a significant positive relationship was found between percentage of motile spermatozoa and Ca+2 ofsemen. On the other hand, Na+, Cl- and pH correlated negatively with sperm movment duration.Understanding of such correlations can be useful to evaluation of sperm quality and make media(extender) for dilution of semen and improving sperm motility parameters of Prussian carp.

Full text

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AACL Bioflux, 2010, Volume 3, Issue 3.
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AACL BIOFLUX
Aquaculture, Aquarium, Conservation & Legislation
International Journal of the Bioflux Society

Correlation between chemical composition of
seminal plasma and sperm motility characteristics
of Prussian carp (Carassius gibelio)
M. Mehdi Taati, Bahareh Mehrad, Ali Shabani, and Amin Golpour

1Department of Fishery, Gorgan University of Agricultural Sciences and Natural
Resources, Gorgan, Iran. Corresponding author: M. Taati, Taati.Mehdi@gmail.com


Abstract. The objectives of the present study were to determine the relationships between chemicals
compositions of seminal plasma with sperm motility traits in Prussian carp, Carassius gibelio (Bloch,
1782). There were significant positive correlations between sperm movment duration and Ca+2 of semen.
Also, a significant positive relationship was found between percentage of motile spermatozoa and Ca+2 of
semen. On the other hand, Na+, Cl- and pH correlated negatively with sperm movment duration.
Understanding of such correlations can be useful to evaluation of sperm quality and make media
(extender) for dilution of semen and improving sperm motility parameters of Prussian carp.
Key Words: sperm motility trait, chemical composition, seminal plasma, Prussian carp.

ﺎﺑ ﯽﻣرﭘﺳا لﺎﻧﯾﻣﺳ ﻊﯾﺎﻣ تﺎﻔﺻ ﯽﺗﮐرﺣ مرﭘﺳا زﻣرﻗ ﯽھﺎﻣ)
Carassius gibelio (Bloch, 1782
رد دوﺟوﻣ ﻊﯾﺎﻣ لﺎﻧﯾﻣﺳ دﺷ هدھﺎﺷﻣ . ر ﮫﮐ دﺷ صﺧﺷﻣ ،نﯾﻧﭼﻣھ یرادﺎﻧﻌﻣ تﺑﺛﻣ ﮫطﺑا
و رﻠﮐ ،مﯾدﺳ ،رﮕﯾد یوﺳ pH تدﻣ لوط ﺎﺑ ﯽﻔﻧﻣ طﺎﺑﺗرا تﮐرﺣمرﭘﺳا دﻧﺗﺷاد . تﺧﺎﻧﺷ نﯾﻧﭼ
ﺎﭘ دوﺑﮭﺑ و ﯽﻧﻣ ﻊﯾﺎﻣ یﺎھرﺗﻣار ﯽﺗﮐرﺣزﻣرﻗ ﯽھﺎﻣ مرﭘﺳا دﺷﺎﺑ دﯾﻔﻣ.
زﻣرﻗ ﯽھﺎﻣ


Introduction. Carassius complex can be considered as the most well known fish in
history (Balon 2004). Today there is no other ornamental fish so popular and easy to
obtain as goldfish. Due to its easy availability and hardiness, goldfish became one of the
most commonly used laboratory animals. Many scientific studies, especially in the field of
physiology, used Carassius sp. as animal model (Rylkova et al 2010). Besides, Carassius
complex has an economical relevance (e.g. Carassius gibelio, sport fishing and/or
pisciculture of subsistence in many countries).
The use of high quality gametes from captive fish broodstock is of great
importance for ensuring the production of valuable offspring for aquaculture (Kjørsvik et
al 1990; Bromage & Roberts 1995). Sperm quality of male broodstock affects the
production of healthy larvae. Seminal plasma produced by the sperm duct provides an
ionic environment that maintains the viability of spermatozoa after their release from the
testes (Ciereszko 2008). Various factors can affect on sperm motility such as pH,
temperature, ions and osmolality (Alavi & Cosson 2006). Because, high quality of semen
is important to the fisheries industry and laboratory research, the biochemical
composition of teleost semen has been studied by many researchers over the years
(Piironen & Hyvarinen 1983; Billard & Menezo 1984). Some parameters such as
spermatocrit, sperm density, fertilization capacity, pH, osmolality and seminal plasma
composition are used to evaluation of sperm quality (Billard et al 1995). The seminal
plasma analysis includes inorganic constituents (Na+, K+, Ca2+, Mg2+) involved in the
process of inhibition or activation of sperm motility (Morisawa et al 1983; Morisawa
1985). Relationship between seminal plasma composition and sperm motility have been
documented in some species; Atlantic salmon, Salmo salar (Hwang & Idler 1969),
Common carp, Cyprinus carpio (Kruger et al 1984), bleak, Alburnus alburnus
(Lahnsteiner et al 1996), Rainbow trout, Oncorhynchus mykiss (Lahnsteiner et al 1998),
دﯾﮑﭼه : ﮫﻌﻟﺎطﻣ
تﻓرﯾذﭘ مﺎﺟﻧا .
دﺻرد نﺎﯾﻣ
دﻧاوﺗ ﯽﻣ
یدﯾﻠﮐ تﺎﻣﻠﮐ:
فدھ ﺎﺑرﺿﺎﺣ
ﯽﻧﻌﻣ تﺑﺛﻣ ﯽﮕﺗﺳﺑﻣھ
مرﭘﺳا کرﺣﺗ
تﮭﺟ رد
ﺑﯾﮐرﺗ نﯾﺑ طﺎﺑﺗرا نﯾﯾﻌﺗ
ی تدﻣ لوط نﯾﺑ کرﺣﺗ
دراد دوﺟو ﯽﻣرﭘﺳا یﺎﻣﺳﻼﭘ مﯾﺳﻠﮐ نازﯾﻣ و
و مرﭘﺳا تﯾﻔﯾﮐ ﯽﺑﺎﯾزرا ردﻧﺗﺳﮐا تﺧﺎﺳ ﺎھ یارﺑندرﮐ قﯾﻗر
،لﺎﻧﯾﻣﺳ یﺎﻣﺳﻼﭘ ،ﯽﯾﺎﯾﻣﯾﺷ تﺎﺑﯾﮐرﺗ ،مرﭘﺳا ﯽﺗﮐرﺣ یﺎھ ﯽﮔژﯾو
تﺎ ﯽﯾﺎﯾﻣﯾﺷ
راد مﯾﺳﻠﮐ و مرﭘﺳا
.زا
رو طﺑاﯽ

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Persian sturgeon, Acipenser persicus (Alavi et al 2004), Chinook salmon, Oncorhynchus
tshawytscha (Rosengrave et al 2009). Fish spermatozoa are immotile in seminal fluid
because of its chemical properties. In this paper, value of chemical parameters of semen
including the inorganic composition (Na+, K+, Ca2+, Mg2+ Cl−), organic composition
(protein, glucose and cholesterol) and pH and their relationships with sperm motility
characteristics (percentage and duration of motility) were investigated in Prussian carp.

Material and Method. The experiment was carried out at the aquaculture Center of
Gorgan University, Iran. Males were captured from reared hatchery at Nahar Khoran,
Gorgan, during the spawning season of Prussian carp. To stimulate fish for spawning we
injected intraperitoneally: 0.5 ml kg-1 b.w. Ovaprim (sGnRHa+dompridon). Milt samples
were collected during the 2010 spawning season from 30 sexually mature two-year-old
male Prussian carps (TL: 60.9 ± 5.3 cm, TW: 1572.7 ± 177.9 g). Semen samples were
collected by massage from the anterior portion of the testis towards the genital papilla.
Care was taken to avoid contamination of the semen with water, mucus, blood cells,
faeces or urine. To analyse the ionic composition of seminal plasma, the semen was
separated from the seminal plasma by centrifugation (Eppendorf AG, Hamburg,
Germany) and the supernatant was separated and stored frozen at -20 °C until the time
of analysis. The pH of seminal plasma was immediately determined using a laboratory pH
meter (pH meter, Iran 762). Two mineral (Ca+2 and Mg+2) and three biochemical
parameters (total protein, glucose and cholesterol) of the seminal plasma were measured
by spectrophotometric method (S2000-UV/VIS England). The concentration of Na+ and
K+ were determined with flame photometer (Jenway PFP, England) (standard kits from
Parsazmoon, Tehran, Iran).
Sperm motility analysis. Sperm motility triggered directly in activation medium
0.3% NaCl at ratio 1:1000 and immediately recorded by a videocamera (Panasonic
wv.cp240 Japan) coupled with dark field microscope (Leica USA). The duration of sperm
motility was measured immediately after initiation of sperm activation until 100 %
spermatozoa were immotile and expressed as sperm movment duration. Percentages of
motile spermatozoa after activation (%) were measured. Only forward moving sperm
were judged motile, those simply vibrating or turning on their axes was considered
immotile (Aas et al 1991).
Statistical analysis. The relationship between composition of the seminal fluid
and sperm motility characteristic (sperm movment duration) was tested using the
bivariate correlation coefficients of Pearson. Then, the Linear and non-linear regression
models were investigated using regression fits. The sperm movment duration was used
as dependent and the parameters of seminal fluid as independent variables.

Results and Discussion. The maximum, minimum, and mean of the percentage (7s
after activation) and duration of motility, ion composition and pH of the seminal fluid of
Prussian carp have been shown in Table 1.
Significant positive relationships were detected for the percentage of motile
spermatozoa vs. Ca and also, the duration of motility vs. Ca+2 of semen (Table 2). A
negative relationship was recorded between sperm movment duration and Na+, Cl- and
pH (see Table 2).
On the other hand, no relationship was found between metabolites of composition
of seminal plasma (glucose, total protein and cholesterol) and sperm movment duration
and percentage of motile spermatozoa. A negative correlation between percentage of
motile spermatozoa and K+ was recorded. Significant positive correlation between Ca+2
and sperm movment duration is shown in (Figure 1).
This is the first study that shows chemical composition of seminal plasma and its
relationship with sperm motility characteristics in Prussian carp. The differences in the
concentrations of inorganic components of semen samples of Prussian carp were
observed.

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Table 1
Maximum, minimum, and the mean of seminal plasma parameters
and motility characteristics of sperm in Prussian carp

Table 2
Relationships between the chemical composition of seminal plasma
and sperm motility traits in Prussian carp




Figure 1. Relationship between the Ca+2 and sperm movment duration in Prussian carp
(independent variable: Ca+2, dependent variable: sperm movment duration).
Seminal plasma parameters Maximum Minimum Mean SD
Sperm movment duration (s)
Percentage of motile spermatozoa
48.46
84
18.81
70
33.63
79
4.03
3
Na (mM.L )

K (mM.L )

Cl- (meqL)
173.18 99.69 101.59 21.29
32.18 20.22 26.20 3.84
163.56 122.8 143.18 11.77
Ca (mM.L )

Mg (mM.L )

pH

Glucose (mM l−1)
0.98 0.34 0.66 0.17
2.02 1.04 1.53 0.28
8.9 8.6 8.75 0.10
0.24 0.144 0.192 0.041
Total protein (gdl-1) 0.05 0.037 0.043 0.004
Cholestrol (mM l−1) 0. 15 0.016 0.083 0.037
Variables Seminal
plasma
parameters

Na (mM.L )

Cl- (meqL)

pH

Ca (mM.L )


K (mM.L )

Bivariate
coefficient


- 0.443

- 0.557

- 0.503

- 0.602
Regression function

R square

P value


y = - 0.059x + 7.338

y = - 0.025x + 8.384

y = - 36.27 + 0.085x + 6.553

y = 0.011x + 0.318

0.413

0.078

0.036

0.36

≤ 0.05

≤ 0.05

≤ 0.05

≤ 0.05


≤ 0.05

Sperm movment
duration (Sec)


Percentage of
motile spermatozoa

- 0.707

y = - 0.642x + 73.82

0.49

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These differences may appear due to the differences in secretary activity of the spermatic
duct epithelium of individuals since the formation of the seminal plasma in fish (inorganic
as well as organic compounds) is a secretion process of the spermatic duct epithelium
(Marshal 1986; Marshal et al 1989; Lahnsteiner et al 1994). In Prussian carp, significant
positive relationships were observed between the duration of motlity and percentage of
motile spermatozoa with Ca+2 of semen respectively. In agreement with our results,
similar relationships were found in other fish, for example, percentage of motile
spermatozoa vs. Ca+2 in brown trout (Hajirezaee et al 2010). In contrast to our results,
several authors were found that there was a negative relationship between the factors
mentioned above; such results are that of Perez et al (2003) for European eel. Several
studies have shown that presence of the organic and inorganic components supports the
viability of spermatozoa (Morisawa et al 1983; Piironen & Hyvarinen 1983; Stoss 1983;
Lahnsteiner et al 1994; Ciereszko et al 2000). In this regard, interactions of ions present
in the seminal plasma with the sperm membrane do influence the membrane potential
and represent a mechanism of inhibition of spermatozoa in the seminal plasma or sperm
duct, allowing the maintenance of the potential of motility before release to the
surrounding medium (Ciereszko et al 2000). These allow the maintenance of the
potential of motility before release to the surrounding medium. In the present study, we
found negative relationship between sperm movment duration and Na, Cl and pH of
semen, whereas, some positive relationships were observed by several authors. For
example, sperm motility vs. Na , K , pH in rainbow trout (Lahnsteiner et al 1998), sperm
motility, sperm movment duration vs. Na , Mg, Cl, spermatozoa motility vs. Na , K , pH in
Alburnus alburnus (Lahnsteiner et al 1996) and duration of motility vs. pH in the rainbow
trout and chum salmon (Oncorhynchus keta) (Morisawa & Morisawa 1986; Morisawa &
Morisawa 1988). But Rosengrave et al (2009) was not observed statistically significant
correlations between sperm motility traits and compositions of seminal plasma. Also,
Alavi et al (2004) observed no statistically significant correlations between seminal
plasma composition and sperm motility traits for Acipenser persicus. In our study, a
negative relationship was detected between percentage of motile spermatozoa and K+ of
semen. Kusa (1950) demonstrated that a high potassium concentration in the seminal
plasma of chum salmon (O. keta) inhibited sperm motility resulting in a decrease in
fertilization success. In contrast to our results, Hajirezaee et al (2010) found a positive
relationship between percentage of motile spermatozoa in brown trout. The effect of
spermatozoa sensitivity to K+ in the seminal fluid may also vary through the reproductive
season (Alavi & Cosson 2006). There appears to be considerable inter- and intra-specific
variability in the ionic composition of seminal plasma in fish (Alavi & Cosson 2006). This
may imply that different ions and ion concentrations are involved in regulating and
initiating sperm motility for different fish species (Billard & Cosson 1992; Scott & Baynes
1980).

Conclusions. Highly significant relationships between percentage of motile spermatozoa
with Ca+2 and also the sperm movment duration and Ca+2 suggest that this parameter
could be considered the most important seminal plasma characteristics influencing the
potential of motility of Prussian carp spermatozoa before sperm ejaculation. Thus, the
Ca+2 and sperm movment duration of semen could be main indicators for evaluating of
semen quality in Prussian carp.

Acknowledgements. We are grateful to the staff of the central laboratory in Gorgan
University. The authors would like to thank the staff of the fish Aquaculture station,
Gorgan, Iran.

References

Aas G. H., Refstie T., Gjerde B., 1991 Evaluation of milt quality of Atlantic salmon. J
Aquacul 95:125-132.

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AACL Bioflux, 2010, Volume 3, Issue 3.
http://www.bioflux.com.ro/aacl
237
Alavi S. M. H., Cosson J., Karami M., Abdolahi H., Mojazi A., 2004 Chemical composition
and osmolality of seminal plasma Acipenser persicus, their physiological
relationship with sperm motility. J Aquaculture international 35:1238-1243.
Alavi S. M. H., Cosson J., 2006 Sperm motility in fishes: effect of ions and osmolality. a
review. J Cell Biol Inter 30:1-14.
Balon E. K., 2004 About the oldest domesticates among fishes. J Fish Biol 65:1–27.
Billard R., Menezo Y., 1984 The amino acids composition of rainbow trout seminal fluid
and blood plasma: a comparison with carp. J Aquacul 41:255-258.
Billard R., Cosson M., 1992 Some problems related to the assessment of sperm motility
in freshwater fish. J Exp Biol 261:122–131.
Billard R., Cosson J., Crim L. W., Suquet M., 1995 Sperm physiology and quality. In:
Brood Stock Management and Egg and Larval Quality, Bromage N. R., Roberts R.
J. (eds), Blackwell Science, Oxford, England pp 25-52.
Bromage N. R., Roberts R. J. (eds), 1995 Broodstock Management and Egg and Larval
Quality, Blackwell Science Ltd., Oxford 424 pp.
Ciereszko A., Glogowski J., Dabrowski K., 2000 Biochemical characteristics of seminal
plasma and spermatozoa of freshwater fishes. In: Cryopreservation in Aquatic
Species, Tiersch T. R., Mazik P. M. (eds). J World Aquaculture Society, Baton
Rouge, USA pp 20-48.
Ciereszko A., 2008 Chemical composition of seminal plasma and its physiological
relationship with sperm motility, fertilizing capacity and cryopreservation success
in fish. In: Fish Spermatology, Alavi S. M. H., Cosson J., Coward R., Rafiee G.
(Eds.), Alpha Science Ltd: Oxford. pp. 215–240.
Hajirezaee S., Mojazi Amiri B., Mirvaghefi A. R., 2010 Relationships between the chemical
properties of seminal fluid and the sperm motility characteristics of Caspian brown
trout, Salmo trutta caspius (A critically endangered Salmonid fish). Research
Journal of Fisheries and Hydrobiology 5(1):27-31.
Hwang P. C., Idler D. R., 1969 A study of major cations, osmotic pressure, and pH in
seminal components of Atlantic salmon. J Fish Res Board Can 26:413–419.
Kjørsvik E., Mangor-Jensen A., Holmefjord I., 1990 Egg quality in fishes. In: Blaxter, J. H.
S., Southward A. J. (eds.), J Adv Mar Biol 26:71–113.
Kruger J. C., Smit G. L., Van Vuren J. J. J., Ferreira J. T., 1984 Some chemical and
physical characteristics of the semen of Cyprinus carpio and Oreochromis
mossambicus. J Fish Biol 24:263-272.
Kusa M., 1950 Physiological analysis of fertilization in the egg of the salmon,
Onchorynchus keta. I. Why are the eggs not fertilized in isotonic Ringer solution. J
Annot Zool Jpn 24:22–28.
Lahnsteiner F., Patzner R. A., Weismann T., 1994 The testicular main duct and the
spermatic duct in cyprinid fishes I. Morphology, fine structure and histochemistry.
J Fish Biol 44:937-951.
Lahnsteiner F., Berger B., Weismann T., Patzner R. A., 1996 Motility of spermatozoa of
Alburnus alburnus (Cyprinidae) and its relationship to seminal plasma composition
and sperm metabolism. J Fish Physiol Biochem 15:167-179.
Lahnsteiner F., Berger B., Weismann T., Patzner R. A., 1998 Determination of semen
quality of the rainbow trout by sperm motility, seminal plasma parameters and
spermatozoa metabolism. J Aquacul 163:163-181.
Morisawa M. K. H., Suzuki S., Shimizu S., Yasuda K., Morisawa M., 1983 Effect of
osmolality and potassium on motility of spermatozoa from freshwater cyprinid
fishes. J Experimental Zoology 107:95-103.
Morisawa M., 1985 Initiation mechanism of sperm motility at spawning in teleosts. J
Zoological Science 2:605-615.
Morisawa S., Morisawa M., 1986 Acquisition of potential for motility in rainbow truot and
chum salmon. J Exp Biol 126:89-96.
Morisawa S., Morisawa M., 1986 Acquisition of potential for motility in rainbow truot and
chum salmon. J Exp Biol 126:89-96.
Morisawa S., Morisawa M., 1988 Induction of potential for sperm motility by bicarbonate
and pH in rainbow trout and chum salmon. J Exp Biol 136:13-22.