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Changes of Sperm Quality Parameters in Caspian Roach (Rutilus Rutilus Caspicus) During Spawning Migration

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Changes of Sperm Quality Parameters in Caspian Roach (Rutilus Rutilus Caspicus) During Spawning Migration Czech J. Anim. Sci., 58, 2013 (3): 117–124 Original Paper Changes of sperm quality parameters in Caspian roach (Rutilus rutilus caspicus) during spawning migration A. Golpour1, M. Akhoundian2, H. Khara3, M. Rahbar1, H. Dadras3 1Young Researchers Club, Islamic Azad University, Lahijan, Iran 2Khorramshahr Marine Science and Technology University, Khorramshahr, Iran 3Department of Fishery and Aquaculture, Faculty of Natural Resources, Islamic Azad University, Lahijan, Iran ABSTRACT: In this study, changes of pH, ionic (Na+, K+, Ca2+, and Mg2+), biochemical (total protein, glucose, and cholesterol) compositions of seminal plasma, sperm motility traits (percentage of motile spermatozoa and sperm movement duration), and sperm production characteristics (sperm volume, spermatocrit, and sperm density) were studied in Caspian roach, Rutilus rutilus caspicus, during spawning migration. Sperm of 10 males was collected three times during the spawning migration (in February, March, and April). The results showed that sperm motility parameters (percentage of motile spermatozoa and sperm movement duration) changed sig- nificantly (P < 0.05) during the reproductive season, but sperm density, spermatocrit, and sperm volume did not show significant differences during spawning migration. Analyses performed at each sampling time (February, March, and April) showed significant differences (P < 0.05) in calcium, magnesium, potassium, and cholesterol, whereas there were no significant changes in Na+, pH, total protein, glucose, and cholesterol. Keywords: spawning season; spermatological parameters; biochemical parameters; semen; roach The Caspian roach (Rutilus rutilus caspicus) 1984; Piironen, 1985; Koldras et al., 1996; Rideout belonging to the family of Cyprinidae has been et al., 2004). Spermatogenesis in fish is regulated considered in fishery in the southern part of the by endogenous (reproductive endocrine system) Caspian Sea (Coad, 1980; Abdoli, 2000). Due to and environmental (thermoperiod or photoperiod) good taste and culinary customs of local people stimuli (Crim, 1982). The physiological changes there has been a great demand for this fish. The that occur after transfer are controlled by endo- population of the Caspian roach has been de- crinological system, which regulates spermato- creased due to overfishing and deterioration of its genesis and spermiation (Billard et al., 1995). In spawning ground and it is therefore considered a freshwater species, matured spermatozoa need threatened species (Kiabi et al., 1999). The roach a hypo-osmotic shock for triggering initiation is a migratory fish which enters the border river of sperm motility (Morisawa et al., 1983). The Atrak and other Iranian rivers (e.g. the Gharesoo duration of sperm motility is usually very short and the Gorganrood) for spawning. The repro- (a few minutes in freshwater species) and could duction migration in the southern Caspian Sea be influenced by external environmental factors starts from January–February and continues until such as pH, temperature, ions, and osmolality April. The fish travel about 70–80 km upriver (Alavi and Cosson, 2006). Seminal plasma char- (Petr, 1987). Most migration takes place at water acteristics (such as osmolality, pH, ionic, and bio- temperature of 10–12°C. Gamete quality in spe- chemical compounds) and Adenosine triphosphate cies with annual spawning cycles varies during (ATP) content of sperm are known as parameters the spawning season (Buyukhatipoglu and Holtz, that influence sperm motility (Alavi and Cosson, 117 Original Paper Czech J. Anim. Sci., 58, 2013 (3): 117–124 2006). In cyprinids, because of high osmolality, 10 mature males was collected by pressing gentile spermatozoa are immotile in the seminal plasma of the abdomen at each sampling time (male mean (Morisawa et al., 1983). Also, osmolality of seminal weight ± SE: 52.57 ± 9.8 g, 56.78 ± 11.4 g, and plasma differs significantly during reproductive 51.12 ± 6.5 g at each sampling time respectively). season (Alavi et al., 2008a). The parameters like Attention was taken to avoid contamination of seminal plasma composition, spermatozoa con- sperm with blood, urine, and faeces. Sperm was centration, spermatocrit, and sperm motility traits kept on ice and immediately transported to the have been used for evaluating the quality of sperm laboratory for analyses. According to procedure of (Alavi and Cosson, 2006). Assessment of sperm Bozkurt et al. (2008) sperm motility was triggered quality based on these parameters provides us directly in activation medium of 0.3% NaCl at ratio with applied approaches to improve methods for 1 : 1000 and immediately recorded with a 3 CCD artificial reproduction by developing immobiliz- video camera Panasonic WV-CP240 (Panasonic ing or activating media for fertilization (Rodina Corp., Osaka, Japan) mounted on a dark-field et al., 2004). Studying the effects of these factors microscope (Leica Camera, Allendale, USA). The on sperm quality can help establish good activa- duration of sperm motility was measured imme- tion and/or immobilizing media for improving diately after initiation of sperm activation until either artificial fertilization or cryopreservation 100% of the spermatozoa were immotile. The per- protocols. It has been already shown the sperm centage of motility was defined as the percentage quality parameters change during the spawning of progressively motile spermatozoa within each season in carp (Cyprinus carpio), tilapia (Oreo- activated sample. Progressively motile spermatozoa chromis mossambicus) (Kruger et al., 1984), rain- were defined as actively swimming in a forward bow trout (Onchorhynchus mykiss) (Munkittrick motion. Only forward moving sperm was judged and Moccia, 1987), and common barbel (Barbus motile and sperm cells that vibrated at the place barbus) (Alavi et al., 2008a). The changes could were considered motile. Observations were made be in terms of spermatozoa concentration, sperm within 2 h of semen collection. The spermatocrit volume, seminal plasma composition, pH or sperm was defined as the ratio of volume of white packed motility parameters such as percentage of motile material to the total volume of semen × 100 (Ru- spermatozoa and sperm velocity (Billard et al., rangwa et al., 2004). Microhaematocrit capillary 1995; Alavi et al., 2008a). Sperm characteristics tubes (75 mm in length, 1.1–1.2 mm in diameter) also differ among species especially in terms of were filled with semen and the end of each tube seminal plasma composition (Ciereszko, 2008) or was sealed with clay. The capillary tubes were cen- sperm motility (Alavi et al., 2008a). These differ- trifuged at 3000 g for 8 min (Sigma, Washington, ences should be considered for developing artificial USA). Measurements were taken in triplicate for fertilization or sperm cryopreservation (Billard et each sample, and the average of the three meas- al., 1995; Linhart et al., 2003). In this study, we urements was used for the results. Spermatozoa determined pH, ionic (K+, Na+, Ca2+, and Mg2+), density was calculated using the haemocytometer biochemical (total protein, glucose, and choles- method. With this aim, semen was diluted with terol) composition of the seminal plasma, sperm a saline solution (0.07 NaCl) (Alavi et al., 2009) production characteristics (spermatocrit, sperm and then a droplet (1 µl) of diluted semen was density, and sperm volume), and sperm motility placed on a Thomas haemocytometer slide (depth parameters (sperm movement duration and per- 0.1 mm) with a coverslip and counted using light centage of motile spermatozoa) during spawning microscopy. After a few minutes (to allow sperm migration in the Caspian roach. sedimentation), the number of spermatozoa was counted at magnification 200× and expressed as spermatozoa × 109/ml. Sperm volume was meas- MATERIAL AND METHODS ured in graduated tubes and expressed in ml. All experiments were performed in triplicate at room Brood fish were caught in the Gorganrood River temperature (20–22°C). To analyze the chemi- (36°44' to 37°49'N latitude and 54°42' to 56°28'E cal components of seminal fluid, the semen was longitude) three times during the spawning season centrifuged (Eppendorf AG, Hamburg, Germany) – at its beginning (February 10th), in the middle at 4000 g for 8 min and then seminal plasma was (March 3rd), and at the end (April 5th). Sperm of collected. Plasma was centrifuged twice to avoid 118 Czech J. Anim. Sci., 58, 2013 (3): 117–124 Original Paper ) b S 100 b 100 ( ac n bc a o i t 80 80 a ur a d t 60 60 n e m 40 e 40 v o Motile spermMotile (%) m 20 20 m r e p 0 0 S February March April February March April Sampling dates Sampling dates Figure 1. Changes in sperm movement duration of Cas- Figure 2. Changes in percentage of motile spermatozoa pian roach during spawning period. Different superscripts of Caspian roach during spawning period. Different su- indicate means that were significantly different (P < 0.05, perscripts indicate means that were significantly different n = 30 males) (P < 0.05, n = 30 males) possible contamination with spermatozoa. The Mg2+, total protein, glucose, and cholesterol during seminal plasma pH was determined immediately spawning season. Before data analysis by ANOVA, before freezing using a Micro pH meter 762 (Saba Duncan test was used for normality of data distri- Co., Tehran, Iran) and then samples were frozen bution and homogeneity of variance. The Duncan at –20°C until analysis. Two minerals (Ca2+ and test was used for comparison between means at a Mg2+) and three
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