Three‐Dimensional Study of the Terminal Portion in Sprague‐Dawley Rat Ejaculatory Ducts

In mammals, a pair of ejaculatory ducts exists in the urethra at the seminal colliculus. The detailed anatomical structures of the distal end of the ejaculatory ducts of Sprague‐Dawley rats were investigated by the computer‐assisted three‐dimensional reconstruction analysis using light‐microscopic serial sections. A three‐dimensional reconstruction revealed that in adult rats, the ejaculatory sinus pair consists of two parts: the cranial section – a compartment region composed of a fusion of the ampullary gland duct and the seminal vesicle duct, and the caudal section – a grooved region composed of a long slitlike ejaculatory ostium that extends into the urethra on both sides of the seminal colliculus. But the sphincter structure was not observed. The long axis of the compartment region was approximately 58 μm in length, and that of the groove region was approximately 495 μm. Although many epithelial glands ducts were distributed throughout the ejaculatory sinuses, the prostate and coagulation gland ducts did not open in these sinuses. The urethra was composed of transitional epithelium, while the ejaculatory sinuses were composed of single to stratified cuboidal epithelium. The ejaculatory ducts continued to the ejaculatory ostium in male adult Sprague‐Dawley rat were composed of the seminal vesicle ducts received the ampullary gland ducts.     

Ovarian Structure and Oogenesis of Catfish Pimelodella vittata (Lütken, 1874) (Siluriformes,Heptapteridae)

The morphology of the ovaries and oogenesis of Pimelodella vittata were studied using anatomical and histological techniques to provide information of its reproductive biology. Eighty adult females were captured trimonthly during the period November 2005 to October 2006. The ovaries are paired, saculiform organs, which are coated with tunica albuginea and contain ovigerous lamellae, where the oocytes develop before being released into the ovarian lumen and following the ovarian duct until reaching the genital papilla. Oogenesis was divided into stages based on the alterations to the nucleus, ooplasm and surrounding follicular layers. Oogonia form groups from the germinal epithelium have asynchronous development and differentiate into initial perinucleolar oocytes. The formation of the zona pellucida is initiated in the advanced perinucleolar oocytes reaching a thickness of 1.46 ± 0.58 μm in the vitellogenic oocytes. The follicular cells are squamous in perinucleolar oocytes, become cubical in the pre‐vitellogenic oocytes and prismatic in the vitellogenic oocytes with a height of 11.20 ± 4.74 μm. The histochemical reactions indicate that zona pellucida, cortical alveoli and yolk globules contain neutral glycoproteins and the follicular cells contain neutral glycoproteins in association with carboxylated and sulphated glycoconjugates. Statistical analyses showed significant differences in the diameter of the oocytes and follicular cells height as oocytes matured. This study represents the first data about the ovarian structure and oogenesis of this species.     

Anatomy and Histology of the Male Reproductive Tract and Spermatogenesis Fine Structure in the Lesser Anteater (Tamandua tetradactyla,Myrmecophagidae, Xenarthra): Morphological Evidences of Reproductive Functions

The anatomy and histology of the male genital tract of the lesser anteater were studied. Fine details of spermatozoa regarding their genesis and morphology were also studied in six adult specimens. The testes lie in the pelvic cavity. The deferent duct emerges from the epididymis and opens into the ejaculatory duct, which drains into the membranous urethra. Accessory glands (prostate, seminal vesicle and bulbourethral gland) are histologically similar to those described in other mammals. The short penis presents an urethral orifice, while the corpus spongiosum becomes thinner at the end indicating the absence of a histologically defined glans. The seminiferous epithelium shows: (1) Sertoli cells with deep nuclear indentations, (2) spermatogonia with crusty‐like chromatin, (3) spermatocytes at different stages of maturation and (4) three morphologically distinct stages of spermatid differentiation according to nuclear shape, acrosome development and chromatin condensation. Sperm heads appear oval. The length of the spermatozoa averages 67.33 ± 1.60 μm. Two specimens with inactive spermatogenesis were azoospermic. Their testes and epididymis presented sizes smaller than those with active spermatogenesis. These studies together with others in anteaters may contribute to successful breeding in conservation programmes.     

Retracted: Study of enzyme activities and protein content of beluga (Huso huso) semen before and after cryopreservation

Knowledge gained regarding the biochemical processes that occur during sperm collection, processing and freezing‐thawing might improve current sperm cryopreservation techniques. In our present study, we determined the effect of cryopreservation on the total protein concentration (TP) and the activities of certain enzymes in semen samples from the beluga (Huso huso). The TP content of the seminal plasma of fresh semen was 0.47 ± 0.026 g/l, and the TP after cryopreservation was 1.86 ± 0.6 g/l. The activities of acid phosphatase (0.82 ± 0.042 U/l), lactate dehydrogenase (234.4 ± 19.4 U/l), arylsulfatase (143.1 ± 32.5 U/l) and β‐N‐acetylglucosaminidase (58.39 ± 4.14 U/l) in the seminal plasma of fresh semen were significantly lower than those in the supernatant of frozen‐thawed semen samples (7.43 ± 0.64, 3224.6 ± 167.2, 422.6 ± 21.3 and 90.2 ± 5.37 U/l respectively). These parameters may be useful as biomarkers for estimating damage to the cell membrane of spermatozoa caused by freezing‐thawing.     

Assessment of age-related morphological changes in the testes of post-hatch light ecotype Nigerian indigenous chicken

The age-related morphological changes of the testes in light ecotype Nigerian indigenous chicken were evaluated in this study using gross anatomical, histological and histomorphometric techniques. The results showed that the testes of 3- to 9-month-old birds were light pink while testes of sexually mature chicken were creamy white in colour. The left and right testicular weight, length, diameter, circumference and the organosomatic indices increased significantly (p < .05) with increasing age across the groups. Although the mean tubular diameter and epithelial height of the left and right seminiferous tubules increased significantly (p < .05) with age, the tubular diameter, epithelial height and luminal diameter did not vary significantly (p > .05) between the left and right testes of all the groups. The one-cell layer thick germinal epithelium of the left testes at 3 to 6 months old showed islands of cell proliferation that contained spermatogonia and spermatocytes. At 6 to 9 months, the left testes exhibited numerous early spermatids with occasional occurrence of late stage spermatids while the right testes showed scanty early stage spermatids. At 12 to 18 months, the germinal epithelia of both left and right testes were characterized by the presence of Sertoli cells, spermatogonia, primary spermatocytes, numerous early and late stage spermatids as well as spermatozoa. In conclusion, the morphological features highlighted in the present study show that at pre-pubertal periods, the left testes may develop faster than the right testes. However, both left and right testes may participate actively in the production of spermatozoa during the post-pubertal life.     

Structure of Vomeronasal Organ (Jacobson organ) in Male Camelus Domesticus Var. dromedaris persica

The vomeronasal organ (VNO) is a tubular structure in the roof of nasal cavity. The important role of this organ is olfaction of sexual odour. In this study, position, anatomical structure and histology of VNO in Iranian camels (camelus domesticus var. dromedaris persica) were determined. Fourteen healthy male camel heads were collected from an industrial slaughterhouse in Tehran, Iran, for anatomical and histological studies (seven each). The length of VNO and width of dental pad and the number and width of palatine crests were measured. For anatomical studies, the mandible was removed, and maxilla and nasal cavity was cut longitudinally and transversely. For histological studies, the mandible was removed, and first 0.5 cm of initial part of VNO was cut. Then, nasal cavity was cut in some segments with 2 cm thickness. The width of VNO was 3.85 ± 0.31 cm and 1.57 ± 0.18 cm in front and distal parts, respectively. The length of VNO was 15.61 ± 0.59 cm. In histological examinations, VNO was surrounded by J‐shape hyaline cartilage. The lining epithelium of lateral wall of VNO was originated from respiratory epithelium, while it had an olfactory epithelium origin in the medial wall. Lamina propria and tunica submucosa were a cavernous connective tissue with seromucous gland with abundant of serous secretory units. The lumen of VNO opens into nasal cavity. The presence of olfactory epithelium found in our study indicates an important role for VNO in pheromone perception and beginning of sexual behaviour.     

Ultrastructural characteristics of black marsh turtle spermatozoa obtained by electroejaculation

The black marsh turtle (Geoemydidae: Siebenrockiella crassicollis) is a freshwater turtle that occurs in equatorial tropical climates in South East Asia. The semen of S. crassicollis was investigated by electroejaculation. The spermatozoa of S. crassicollis are filiform in shape with curved heads. The entire length, midpiece to tail length, tail width and tail length of the spermatozoa were 71.33 ± 1.55 μm, 49.92 ± 1.13 μm, 0.43 ± 0.02 μm and 48.53 ± 0.25 μm, respectively. The head length, head width across the middle and head width across the base were 14.00 ± 0.38 μm, 0.79 ± 0.03 μm and 0.91 ±0.0.03 μm, respectively. The acrosomal region of the S. crassicollis spermatozoa was narrower than the head, with an acrosomal length and width at the annulus of 2.90 ± 0.13 μm and 0.43 ± 0.01 μm, respectively. The midpiece of the S. crassicollis spermatozoa was narrower than the head and contained 30–40 mitochondrial balls, each with a ball diameter of 0.16 ± 0.002 μm. The midpiece length, midpiece width and tail length were 4.92 ± 0.16, 0.78 ± 0.03 and 48.53 ± 0.25 μm, respectively. This study presents the characteristic appearance of a freshwater turtle spermatozoa in Southeast Asia, as observed under an electron microscope. The spermatozoa of Siebenrockiella crassicollis are morphologically different from those of other freshwater turtles from other regions described in previous studies.     

Harderian Gland in Canadian Ostrich (Struthio camelus): A Morphological and Histochemical Study

Heads of ten healthy adult ostrich obtained from slaughter house were the constituted materials of the study. The Harderian gland (HG) was dissected out, and all of the gross morphometrical parameters including length, width and thickness as well as weight of left and right glands were recorded. Tissue sections were stained, using haematoxylin and eosin, Masson trichrome, periodic acid‐Schiff and Alcian blue (pH 2.5) techniques. In ostrich, HG was an orbital organ located ventromedially around the posterior part of the eyeball. It was an oval flatted shape, light pink colour with irregular outline and was pointed in the dorsal end. Its mean length was 35.30 ± 2.84 mm and 35.55 ± 3.58 mm in left and right sides, respectively, and mean width 15.30 ± 1.20 mm and 15.65 ± 1.18 mm in left and right sides, respectively. There was no significant difference between length, thickness, weight and width of left and right glands. Histological results showed that the glandular epithelium was multilobular and compound tubuloalveolar. The gland was surrounded by a connective tissue capsule, and the epithelium was lined by simple columnar epithelial cells of varying height. The secretion of HG was mucous and the secretion type was apocrine. Mucosubstance analysis revealed that secretory units contained acidic and neutral glycoproteins. The granules within the epithelial cells lining the intralobular and inter‐lobular excretory ducts of the gland were positive for periodic acid‐Schiff and Alcian blue (pH 2.5).     

Sperm cryopreservation in the Far‐Eastern wildcat (Prionailurus bengalensis euptilurus)

The Far‐Eastern wildcat (Prionailurus bengalensis euptilurus) is a rare and poorly investigated nondomestic felid species. An attempt of freezing and cryopreserving Far‐Eastern wildcat spermatozoa in CaniPlus Freeze (CPF) medium is reported. Sperm was collected by electroejaculation from five adult Far‐Eastern wildcat captive‐born males. Epididymal spermatozoa from five adult randomly bred domestic cat males were used as a reference. The viability of frozen–thawed spermatozoa evaluated by double staining with SYBR Green I and PI followed by the subsequent confocal laser scanning microscopy (CLSM) was 38.2% ± 3.0% for the domestic cat and 38.0% ± 10.2% for the Far‐Eastern wildcat. The motility of frozen–thawed spermatozoa was 30.8% ± 9.8% for the domestic cat and 33.7% ± 15.1% for the Far‐Eastern wildcat. Sperm morphology was assessed by light microscopy. The total percentage of normal spermatozoa after freezing and thawing was 51.9 ± 5.9 for the domestic cat and 55.0% ± 6.4% for the Far‐Eastern wildcat. Defects of flagella were the most frequently observed abnormalities in both species (32.2% ± 4.8% and 30.8% ± 4.4% of all reported anomalies for the domestic cat and Far‐Eastern wildcat, respectively). Domestic cat epididymal and Far‐Eastern ejaculatory spermatozoa fertilized in vitro‐matured oocytes of the domestic cat (30.0% ± 5.5% and 35.5% ± 15.0%, respectively). Taken together, these results suggest that the freezing of Far‐Eastern wildcat spermatozoa with CPF medium is a suitable method for Felidae cryopreservation.     

Total antioxidant capacity and protein peroxidation intensity in seminal plasma of infertile and fertile dogs

The aim of this study was to evaluate the total antioxidant capacity and protein peroxidation intensity in seminal plasma of infertile and fertile dogs. The study was conducted on 10 infertile and 10 fertile dogs of various breeds. Infertility was defined as conception failure at least three matings with different bitches. Semen was collected by manual manipulation. The sperm concentration and motility parameters were evaluated using CASA Hamilton Thorne, Vers. IVOS 12.3. The morphology of spermatozoa and the percentage of live and dead sperm cells were assessed microscopically, total antioxidant capacity and the content of SH‐groups in seminal plasma were determined spectrophotometrically, the contents of protein peroxidation markers in seminal plasma, bityrosine and formylokinurenine, were determined using spectrofluorimetric methods. Sperm concentration and total sperm count were significantly (p < 0.05) lower in infertile dogs than in fertile dogs (99.92 ± 3 0.05 × 10⁶/ml vs. 282.07 ± 48.27 × 10⁶/ml; 214.19 ± 114.74 × 10⁶ vs. 747.57 ± 210.94 × 10⁶, respectively). The percentage of spermatozoa with normal morphology and the most determined motility parameters differed significantly (p < 0.05) between both groups. The mean values of total antioxidant capacity in the seminal plasma were significantly (p < 0.05) lower (19.95 ± 20.94 vs. 25.66 ± 23.18 µmol/g protein), whereas the mean contents of bityrosine and formylokinurenine in seminal plasma were significantly (p < 0.05) higher in infertile dogs than in fertile dogs (3.71 ± 4.83 µg/mg protein vs. 1.55 ± 2.00 µg/mg protein and 0.37 ± 0.45 µg/mg protein vs. 0.14 ± 0.08 µg/mg protein, respectively). In conclusion, the obtained results suggest that the poor semen quality and infertility in dogs could be associated with lowered total antioxidant capacity and increased protein peroxidation in seminal plasma as a consequence of oxidative stress.     

Seasonal variations in the morphology of the seminal vesicles of the African straw-coloured fruit bat (Eidolon helvum)

The study investigated seasonal changes in the morphology of the seminal vesicles of Eidolon helvum in a typical African tropical environment. Seminal vesicles of forty-eight bats were examined during the early rainy, late rainy and peak dry seasons, using gross anatomical, histological and ultrastructural techniques. The interlobular connective tissue septa from the external capsule demarcated the glandular parenchyma into numerous lobules of tubulo-alveolar glands. A simple cuboidal epithelium of mono- or bi-nucleated cuboidal cells lined the glandular mucosa during the rainy season. This was replaced by a bistratified cuboidal epithelium, whose upper layer of cells showed evidence of degeneration, during the dry season. The PAS-positive secretory products appeared to be numerous during the rainy season, but few and clumped together during the dry season. The mean gross weight, interlobular septal thickness, acini diameter and epithelial height of the glands varied significantly with season. Ultrastructural features of the secretory cells during the rainy season showed well-developed cytoplasmic organelles, numerous electron lucid secretory vesicles and electron-dense granules. Secretory epithelial cells of the seminal vesicles are very active during the rainy season, while regeneration of degenerated epithelium in preparation for the next reproductive cycle occurs during the dry season.     

Effects of single layer centrifugation (SLC) on bull spermatozoa prior to freezing on post‐thaw semen characteristics

Single layer centrifugation (SLC) has been shown to select the most robust spermatozoa from the ejaculate in several species. Here the effects of SLC prior to freezing on various parameters of frozen‐thawed bovine sperm quality are reported. Semen from 8 bulls was layered on top of a species‐specific colloid, Bovicoll. After centrifugation for 20 min at 300 g, the resulting sperm pellet was resuspended in OPTIXcell^® (IMV Technologies, l′Aigle, France); the SLC‐selected sperm samples and uncentrifuged controls were frozen. On thawing, all sperm samples were analysed for membrane integrity, production of reactive oxygen species, mitochondrial membrane potential (MMP) and chromatin integrity. The SLC‐treated samples had a higher percentage of live, superoxide‐positive spermatozoa than uncentrifuged samples (27.9 ± 5.1% versus 21.7 ± 6.7%; p = .03). They had a higher proportion of spermatozoa with high mitochondrial membrane potential than uncentrifuged samples (55.9 ± 8.2% versus 40.5 ± 15.1%; p = .03) and also a lower proportion of spermatozoa with low mitochondrial membrane potential than non‐treated samples (42.0 ± 8.5% versus 55.9 ± 14.4%; p = .04). No significant effects of treatment were found for membrane integrity or chromatin integrity. The effect of bull was significant on the proportions of dead, superoxide‐positive spermatozoa and live, hydrogen peroxide‐negative spermatozoa, as well as on membrane integrity, but it was not significant for mitochondrial membrane potential or chromatin integrity. These results suggest that SLC selects the most metabolically active bull spermatozoa from the rest of the population in normal ejaculates; the pattern of reactive oxygen species production may be different in SLC‐selected spermatozoa compared to unselected samples.     

Morphology and Morphometry of Seminiferous Tubules in Crioulo Horses

This study aimed to assess the biometrics of the testes and the morphology of the seminiferous tubules of Crioulo horses. We studied 10 sexually mature stallions (3–6 years of age). After orchiectomy, testes were perfused with Karnovsky's solution and then embedded in glycol methacrylate. Testis sections (4 μm) were cut and stained with toluidine blue and a solution of 1% sodium borate. The histological images were digitized, and the morphometric analysis was performed using ImageJ software. The average weight of the stallions was 377.5 kg, and the average weight of both testicles was 162.9 g. The percentage of testicular parenchyma occupied by the seminiferous tubules and the intertubular tissue was 77.97% ± 6.34% and 22.03% ± 6.34%, respectively. The average tubular diameter was 205.00 ± 36.91 μm, whereas the average height of the seminiferous epithelium was 70.56 ± 2.82 μm. Average tubular length per testicle and average tubular length per gram of testicle were 4,085.10 ± 1,170.68 m and 26.09 ± 10.63 m/g, respectively. The characteristics of the eight stages of the seminiferous epithelium cycle were similar to those reported in other horse breeds. We conclude that the morphometry of the seminiferous tubules of Crioulo horse resembles what has been reported in other horse breeds. The volumetric proportion of the seminiferous tubules and the Leydig cells of the Crioulo horse is one of the highest ever reported for stallions.     

Effects of cooling rates on the quality of Prochilodus lineatus (Valenciennes, 1836) sperm

This study aims to investigate the effect of different cooling rates on the semen cryopreservation of curimba (Prochilodus lineatus). Nineteen ejaculates were obtained from adults males and cryopreserved at 15°C/min (CR15), 30°C/min (CR30) (controlled temperature inside and outside straw, speed was stable during freezing) and direct freezing in liquid nitrogen vapour (~35.6°C/min) (CRNV). The straws were thawed and seminal parameters evaluated. DNA fragmentation through the comet assay was assessed. A fresh sperm sample was not frozen and used for analyses. Data were submitted to an analysis of variance (ANOVA), and means were compared by Scott–Knott test (p < 0.05) using the R Software. Mean motility percentage was 100%, and motility duration was 39.5 ± 5.7 s for the fresh sperm (subjective analysis); 58.9 ± 8.0% and 24.5 ± 5.7 s for CR15; 64.8 ± 4.8% and 26.5 ± 7.1 s for CR30; and 50.1 ± 16% and 25.7 ± 4.7 s for CRNV, respectively. Motility percentages were higher and equal between CR15 and CR30 compared to CRNV (p < 0.05). Some sperm motion kinetics, namely average path velocity (VAP) and straight line velocity (VAS), were higher for CR30 (p < 0.05), while curvilinear velocity (VCL) and velocity progression (PRO) were lower for CRNV (p < 0.05). Straightness (STR) and wobble (WOB) were the same among treatments (p > 0.05). Sperm morphology results indicated higher means for total morphological sperm alterations in CRNV. All cooling rates caused sperm DNA fragmentation, although CR30 provided a less harmful effect. This is the first report for cryopreserved P. lineatus sperm preserved under different controlled cooling rates. The cooling rate of 30°C/min is indicated for the cryopreservation of this fish sperm as it led to the lowest detrimental spermatozoa effects.     

Organogenesis and foetal haemodynamics during the normal gestation of healthy black‐rumped agoutis (Dasyprocta prymnolopha,Wagler, 1831) bred in captivity

The objective of this study was to define the patterns of organogenesis and foetal haemodynamics during the normal gestation of healthy agoutis (Dasyprocta prymnolopha) kept in captivity. Thirty pregnant agoutis that ranged in size from small to medium and weighed between 2.5 and 3 kg underwent B‐mode and Doppler ultrasonography for the biometric evaluation of the foetal organs. The foetal aortic blood flow proved to be predominantly systolic, and the measured flow velocity was 78.89 ± 2.95 cm/s, with a maximum pressure gradient of 2.12 ± 0.27 mmHg. The liver was characterized by its large volume, occupying the entire cranial aspect of the abdominal cavity, and it was associated cranially with the diaphragm and caudally with the stomach. The flow velocity in the portal vein was estimated to equal 12.17 ± 2.37 cm/s, with a resistivity index of 0.82 ± 0.05. The gallbladder was centrally located and protruded cranially towards the diaphragm. The spleen was visualized as an elongated structure with tapered cranial and caudal extremities, and the foetal kidneys were visualized bilaterally in the retroperitoneal region, with the right kidney positioned slightly more cranially than the left. The morphological characterization and hemodynamic analysis of the foetal organs of black‐rumped agoutis via B‐mode and Doppler ultrasonography allow determination of the vascular network and of reference values for the blood flow required for perfusing the anatomical elements essential for maintaining the viability of foetuses at different gestational ages.     

Detection of intracellular reactive oxygen species (superoxide anion and hydrogen peroxide) and lipid peroxidation during cryopreservation of alpaca spermatozoa

The objective of this study was to detect changes in intracellular reactive oxygen species (superoxide anion and hydrogen peroxide) production and lipid peroxidation during cryopreservation of alpaca spermatozoa. Twelve alpaca semen samples were conventionally cryopreserved. Intracellular superoxide anion and hydrogen peroxide were evaluated by fluorescence microscopy using dihydroethidium (DHE)/YO‐PRO‐1 and dichlorofluorescein diacetate (H2DCFDA)/propidium iodide (PI), respectively. Evaluations were performed during cooling curve at (1) 25°C, (2) 15°C, (3) 5°C/0 min, (4) 5°C/15 min, (5) 5°C/30 min and (6) after freezing/thawing. Evaluation of lipid peroxidation by measuring malondialdehyde (MDA) was performed at 25°C, 5°C/30 min and after thawing. Maximum percentages of total spermatozoa producing superoxide anion and hydrogen peroxide were found at 5°C/30 min (62.8 ± 6.3% and 30.5 ± 5.6%, respectively), and these results were higher (p < .05) than initial (25°C: 10.8 ± 3.8% and 6.8 ± 0.7%, respectively) and after thawing (29.8 ± 9.5% and 7.5 ± 1.8%, respectively) values. However, considering only viable spermatozoa, production of superoxide anion and hydrogen peroxide during overall stabilization at 5°C (>76% and >91%, respectively) and after thawing (74.9 ± 5.0% and 78.9 ± 2.2%, respectively) was higher (p < .05) than initial values at 25°C (38.7 ± 3.1% and 53.6 ± 2.0%, respectively). Lipid peroxidation at 25°C, 5°C/30 min, and post‐thawing were 346.5 ± 99.8, 401.1 ± 64.8 and 527.7 ± 142.8 ng/ml MDA, respectively. These results showed that high percentage of viable alpaca spermatozoa produces intracellular reactive species oxygen (ROS) during the cryopreservation process of alpaca semen.     

Comparison of two methods of seminal plasma removal on buffalo (Bubalus bubalis) sperm cryopreservation

Cryopreservation causes damage to spermatozoa, and methods minimizing this damage are therefore needed. Although much discussed, seminal plasma removal has become an alternative to improve sperm quality and viability after freezing and has been applied to different species in attempt to obtain good results. The objective of this study was to evaluate semen quality in buffaloes submitted to two methods for seminal plasma removal (filtration and centrifugation). Semen samples were collected from seven Murrah buffalo bulls (Bubalus bubalis) once a week for 8 weeks. Each ejaculate was divided into three groups: control (presence of seminal plasma), centrifugation and filtration. Sperm kinetics was evaluated with the computer‐assisted sperm analysis (CASA) system. Plasmalemma and acrosomal membrane integrity, mitochondrial membrane potential and reactive oxygen species (ROS) were measured by flow cytometry, and lipid peroxidation was evaluated by the thiobarbituric acid reactive substances (TBARS) assay. Seminal plasma removal did not improve sperm kinetics compared to the control group. Centrifugation increased the number of cells with damaged acrosomal membranes (0.77 ± 0.05) and filtration caused greater plasmalemma and acrosomal membrane damage (22.18 ± 1.07). No difference in the mitochondrial membrane potential was observed between groups. In contrast, ROS production was higher in the centrifugation group compared to the control and filtration groups, although no differences in TBARS formation were detected. In conclusion, seminal plasma removal did not improve the quality of thawed buffalo semen compared to control in terms of sperm kinetics, membrane integrity, mitochondrial membrane potential or lipid peroxidation.     

Lipid and protein oxidation levels in spermatozoa and seminal plasma of Asian Elephants (Elephas maximus) and their relationship with semen parameters

Peroxidation damage to spermatozoa and seminal plasma has an important role in sperm quality. Thus, the objective of this study was to determine the levels of lipid and protein oxidation in spermatozoa and seminal plasma of Asian elephants (Elephas maximus) with varying percentage of progressive motility. Lipid and protein oxidation was measured by the thiobarbituric acid‐reactive species (TBARS) assay and the 2, 4‐dinitrophenylhydrazine (DNPH) carbonyl groups assay, respectively. Fresh semen samples were collected from Asian elephants and classified according to the percentage of motile spermatozoa into good (>60%) and poor (≤20%) motility. Results revealed that seminal plasma malondialdehyde (MDA) and seminal plasma protein carbonyls (PCs) were significantly higher in poor motility than in good motility (p < .05). The MDA and PC levels in seminal plasma were negatively correlated with the percentages of progressive motility (p < .05). In addition, the negative correlation between sperm concentration and seminal plasma MDA level was investigated (p < .05). The sperm viability was also negatively correlated with sperm PC level (p < .05). This study indicated that lipid and protein oxidation has deleterious effect on semen quality of Asian elephants.     

Macroscopic Anatomy of the Lower Respiratory System in Mole rats (Spalax leucodon)

The morphologic and morphometric features of the lower respiratory system in mole rats were examined. It was seen that the low respiratory system of this species leading a special life under highly hypoxic/hypercapnic conditions underground is structurally similar to other mammals living on land in terms of the parts examined; trachea was formed by 29.5 ± 4 oval‐formed cartilaginous tracheals arranged backwards and became gradually more stenotic diameter from cranial to the caudal of the neck. The trachea was separated in two principal bronchus at the fourth thoracal intercostal spatium level. The angle between the two main principal bronchi was 60.5 ± 2.35°. The lung constituted 1.29 ± 0.03% of the body weight and the right lung was heavier than the left lung. Fissura inter‐lobaris was deep and separated the lung lobes wholly, and the right lung was separated in four lobes, whereas the left lung was not separated into the lobes. Also, the medial lobe of the left lung was the lightest lobe.