Comparative study of alpha-glucosidase activities in blood and seminal plasma of rams

1. This study was undertaken to characterize the alpha-glucosidase present in blood and seminal plasma of two strains of pure-bred rams which are known as seasonal breeders. 2. pH profiles and activity levels were investigated in blood and seminal plasma using a sensitive spectrophotometric assay with para-nitrophenyl-alpha-D-glucoside as substrate. 3. According to their pH optimum, blood plasma and seminal plasma alpha-glucosidases were typically neutral and acid enzymes and significant differences were recorded in their physico-chemical properties, establishing the tissue specificity of the enzyme. 4. Notwithstanding the tissue under study, the nature of the alpha-glucosidase activity was similar in both strains of pure-bred rams.     

Effects of increased scrotal temperature on semen quality and seminal plasma proteins in Brahman bulls

Environmental temperature has effects on sperm quality with differences in susceptibility between cattle subspecies and breeds, but very little is known about the seminal plasma protein (SPP) changes resulting from testicular heat stress. Scrotal insulation (SI) for 48 hr was applied to Brahman (Bos indicus) bulls. Semen was collected at 3‐day intervals from before, until 74 days post‐SI. The changes in sperm morphology and motility following SI were comparable to previously reported and differences were detected in measures of sperm chromatin conformation as early as 8 days post‐SI. New proteins spots, in the SPP two‐dimensional (2‐D) gels, were apparent when comparing pre‐SI with 74 days post‐SI, and SPP identified as associated with mechanisms of cellular repair and protection. Similar trends between 2‐D gel and Sequential Window Acquisition of All Theoretical Mass Spectra (SWATH‐MS) data was observed, with SWATH‐MS able to quantify individual SPP that otherwise were not resolved on 2‐D gel. The SPP assessment at peak sperm damage (21–24 days) showed a significant difference in 29 SPP (adjusted p < .05), and identified six proteins with change in abundance in the SI group. In conclusion both spermatozoa and SPP composition of bulls are susceptible to temperature change incurred by SI, and SPP markers for testicular heat insults may be detected.     

Effect of histamine and acetylcholine on hypophysial stalk plasma dopamine and peripheral plasma prolactin levels.

To further define the role of dopamine in the regulation of prolactin secretion, we studied the effect on prolactin and hypothalamic dopamine secretion of histamine and acetylcholine (ACh) injected into the lateral ventricle of urethane anesthetized diestrus-1 rats. Histamine (10 μg) caused a 592% increase in plasma prolactin levels and a 26% decrease in stalk plasma dopamine levels. ACh (50 μg) caused a 2090% increase in plasma prolactin levels but no significant change in stalk plasma dopamine concentration.To determine if the 26% fall in stalk plasma dopamine following histamine administration could account for the 6-fold increase in plasma prolactin, we measured the effect on prolactin secretion of a similar decrease in administered dopamine. During an infusion of physiologic levels of dopamine, a 25% decrease in arterial plasma dopamine concentration resulted in only a 2-fold increase in prolactin secretion.The results of these experiments suggest that the effect of histamine on prolactin secretion may be mediated in part by decreased hypothalamic secretion of dopamine but that an additional hypothalamic hormone is probably involved. The stimulatory effect of ACh on prolactin secretion is not mediated by dopamine. These data are consistent with the growing evidence for the participation of multiple hypothalamic factors in the regulation of prolactin secretion.     

Protective effect of esterified glucomannan on aflatoxin-induced changes in testicular function,sperm quality,and seminal plasma biochemistry in rams

The aim of this study was to determine the effect of aflatoxin (AF) on spermatologic, biochemical, and testis parameters in rams, and the protective efficiency of esterified glucomannan (EG) co-administered with AF. Thirty-two Merino rams (12–14 months old) were used. The experimental design consisted of four dietary treatments. The control group was fed commercial feed. The AF group was fed with commercial feed plus 250 μg/d of total AF. The EG group received commercial feed plus 2 g/d of EG. The AF + EG group was given commercial feed plus 250 μg/d of total AF and 2 g/d of EG. There were treatment, time, and treatment-by-time interaction effects on sperm motility, abnormal spermatozoa, damaged acrosome, and dead spermatozoa (P < 0.01). The percentage of motile sperm was lower and the percentages of abnormal sperm, sperm with damaged acrosomes, and dead sperm were greater in the AF group than in the control, AF+EG, and EG groups, as from week 3 until the end of week 12 (P < 0.05). As from week 3, hyaluronidase activity in the seminal plasma increased significantly in the AF group, compared with the control. The co-administration of AF+EG was found to be effective in preventing the increase in hyaluronidase activity. As week 4, malondialdehyde (MDA) levels were significantly higher in the AF group compared with the control. The combined administration of AF+EG was found to be effective in lowering the MDA levels, increased by AF, to the levels measured in the control (P < 0.05). Although glutathione (GSH) levels were determined to have significantly decreased in the AF group in comparison to the control, it was observed that, in the group co-administered with AF and EG, particularly after week 7, the GSH levels, which had decreased owing to AF, were largely ameliorated (P < 0.05). In conclusion, AF adversely affected spermatologic, biochemical, and testis parameters, and the combined administration of EG with AF reversibly eliminated these adverse effects in rams.     

Cimetidine and plasma levels of gonadotropins,prolactin and gonadal steroids in women.

The endocrine effects of cimetidine (Tagamet) during the menstrual cycle were investigated in seven healthy female volunteers. The subjects were studied for six menstrual cycles divided into the pretreatment phase, a phase of therapy with 1.2 g of orally administered cimetidine daily for two cycles, and a post-treatment phase. Cimetidine therapy induced a significant increase in the mean plasma level of follicle-stimulating hormone during the periovulatory period, followed by modest but sustained hyperprolactinemia throughout the luteal phase of each cycle. No significant changes were found in the mean plasma levels of luteinizing hormone and progesterone, and the mean plasma estradiol level was significantly decreased only in the midproliferative phase of each cycle. The mean plasma prolactin levels after a bolus injection of thyrotropin-releasing hormone in the midluteal phase during cimetidine administration did not differ from the mean control levels, which indicates that cimetidine modulates the release of prolactin at the suprapituitary locus. However, the significance of the endocrine changes remains to be established.     

Immunomodulatory cytokines in human seminal plasma correlate with immunomodulatory steroids

7alpha-Hydroxy-dehydroepiandrosterone and its 7beta-hydroxyepimer, which act as local immunomodulatory agents, dehydroepiandrosterone, cortisol, and major androgens, together with four cytokines-interleukins 2, 4, 10, and IFN-gamma, reflecting the activity of TH1 or TH2 cells present in semen, were measured in seminal plasma from 35 male donors. Cortisol, dehydroepiandrosterone, its sulfate, 7-hydroxy-dehydroepiandrosterone epimers, testosterone, and estradiol were also measured in their blood serum. Steroids and interleukins in semen as well as serum steroids and seminal interleukins were mutually correlated to find out whether a relationship between immunomodulatory steroids and cytokines influencing the immune environment does exist. A highly significant (P<0.001) positive correlation was found between seminal 7beta-hydroxy-dehydroepiandrosterone and IFN-gamma, while a negative correlation was found between cortisol and IL-10. Highly significant positive correlations were also found between serum 7alpha-hydroxy-dehydroepiandrosterone and seminal IFN-gamma and between serum 7beta-hydroxy-dehydroepiandrosterone and seminal IL-2, while a negative correlation was found between serum dehydroepiandrosterone and seminal IL-10. Different and in some instances, even contradictory findings concerning the influence of dehydroepiandrosterone and cortisol on TH1 and TH2 cytokines were observed in seminal plasma as compared to those found by others in serum. The differences can be ascribed to the different environments of mucosal and systemic immunity. Correlations between the levels of steroids and cytokines in seminal plasma did not always correspond to the correlations between given cytokines and hormones in sera. The results, however, are in agreement with our recent finding of an autonomous production of these steroids in the male reproductive tract.     

Effect of constant-release implants of melatonin on seasonal cycles in reproduction, prolactin secretion and moulting in rams

Seasonal cycles in the size of the testes, blood plasma concentration of testosterone, FSH and prolactin, intensity of the sexual skin flush, timing of rutting behaviour and moulting of the body coat were recorded in Soay rams after s.c. implantation of melatonin contained in a Silastic envelope which increased the circulating blood levels of melatonin to 200-600 pg/ml for many months. Two groups of 8 adult rams were held under alternating periods of short days (8L:16D) and long days (16L:8D) to drive the seasonal cycles and the treatments with melatonin were initiated during the long or short days, and one group of 8 ram lambs was kept out of doors and given implants during the long days of summer (4 melatonin-implanted and 4 control (empty implants) rams per group). The treatments demonstrated that melatonin implants during exposure to long days resulted in a rapid 'switch on' of reproductive redevelopment similar to that produced by exposure to short days melatonin implants prevented the rams from showing the normal responses to changes in the prevailing photoperiod rendering them nonphotoperiodic; and long-term cyclic changes in testicular activity, prolactin secretion and other characteristics occurred in the melatonin-implanted rams; the pattern was similar to that previously observed in rams exposed to prolonged periods of short days. The overall results are consistent with the view that melatonin is the physiological hormone that relays the effects of changing photoperiod on reproduction and other seasonal features, and that continuous exogenous melatonin from an implant interferes with the normal 'signal' and produces an over-riding short-day response.     

Carnitine, acetylcarnitine and the activity of carnitine acyltransferases in seminal plasma and spermatozoa of men, rams and rats.

The concentration of total carnitine (i.e. carnitine plus acetylcarnitine) was measured in seminal plasma and spermatozoa of men and rams. In ram semen, there was a close correlation between the concentration of spermatozoa and that of total carnitine in the seminal plasma, indicating that the epididymal secretion was the sole source of seminal carnitine. The percentage of total carnitine present as acetylcarnitine was 40% in seminal plasma and 70-80% in spermatozoa. The acetylation state of carnitine in seminal plasma was apparently not influenced by the metabolic activity of spermatozoa in ejaculated ram semen as no change was found in the plasma concentration of carnitine or acetylcarnitine up to 45 min after ejaculation. In spermatozoa, the activity of carnitine acetyltransferase (EC 2.3.1.7) was approximately equivalent to that of carnitine palmitoyltransferase (EC 2.3.1.21); and the activity of these enzymes was similar in ram and human spermatozoa but greater in rat spermatozoa. It is concluded that there is no correlation between the content of either total carnitine or the carnitine acyltransferases and the respiratory capacity of spermatozoa.     

Response of pre-pubertally castrated rams and ewes to artificial photoperiod--changes in plasma LH and prolactin

Castrate rams and ovariectomized ewes were maintained in the presence of entire rams and ewes and subjected to successive periods of alternating 6 h light:18 h darkness ('short' days) and 18 h light:6 h darkness ('long' days) preceded by a period of 12 h light:12 h darkness ('constant' light days). Plasma concentrations of LH and prolactin were measured in the castrate animals in order to determine how LH and prolactin secretion responded to the artificial light regime and corresponding periods of elevated or depressed testicular and ovarian activity in the entire rams and ewes. There was no variation in mean plasma LH concentrations or LH pulse frequency with either the changes in photoperiod or the phases of gonadal activity in the entire animals. However, there was a highly significant (P less than 0.001) relationship between prolactin secretion and the artificial photoperiod in both castrate groups with high and low levels coinciding with long and short days respectively. In addition, there was a marginally significant (P less than 0.1) relationship between prolactin secretion in the castrate ram and the stage of testicular activity in the entire rams with elevated levels associated with regressed activity. Prolactin secretion in the ovariectomized ewes was significantly (P less than 0.05) related to the phase of ovarian development with high levels associated with acyclic activity. It is concluded that LH secretion and pituitary responsiveness to exogenous GnRH were not modified by the artificial light regime. However, the changing light pattern was physiologically 'perceived' by the castrate animals as indicted by a concomitant variation in plasma prolactin concentrations.     

Effect of dietary energy on seminal plasma insulin-like growth factor-I (IGF-I), serum IGF-I and testosterone levels, semen quality and fertility in adult rams

The objective of the present study was to modulate seminal plasma insulin-like growth factor-I (IGF-I) by dietary energy and assess the relationship among testosterone and IGF-I levels, semen quality and fertility in adult rams. Twenty-four 1-yr old adult Nellore rams were equally divided into three groups (n = 8) and fed with three different concentrate mixtures formulated using conventional ingredients and finger millet (Eleucine corocana) straw to ensure rams received with similar amount of crude protein with three levels of energy. Rams in low-energy group were offered diets with 20% less energy than the control energy group (optimum energy, 100%, recommended energy level), whereas rams in high energy group were offered diets with 20% more energy than the optimum energy group. Semen was collected from rams 60 days after start of the experimental feeding. The percentages of progressive forward motility, functional membrane integrity and mitochondrial membrane potential of the spermatozoa were significantly (P < 0.05) higher in control and high energy groups as compared to low-energy group. Feeding of low-energy diet significantly (P < 0.05) decreased spermatozoa VSL, VCL and VAP when compared to control and high energy fed groups. The number of spermatozoa binding/oocyte was significantly (P < 0.05) higher in control (11.23 ± 0.20) and high energy (10.57 ± 0.19) groups as compared to the low energy (6.14 ± 0.01) group. The serum and seminal plasma IGF-I levels were significantly (P < 0.05) higher in control and high energy fed groups as compared to the low-energy group. The serum testosterone and cholesterol levels were significantly (P < 0.05) higher in the control group as compared to the low-energy group. The seminal plasma fructose levels in optimum energy fed animals were significantly (P < 0.05) higher as compared to other two groups. The seminal plasma IGF-I level had positive correlation with progressive forward motility (r = 0.7) and other velocity (linearity, r = 0.7; straightness, r = 0.7) parameters. The study suggested that the modulation of seminal plasma IGF-I levels by dietary energy is possible and the optimum level of seminal plasma IGF-I is necessary and sufficient to influence semen quality.     

Effects of heating the testes and epididymides of rams by scrotal insulation on fertility and embryonic mortality in ewes inseminated with frozen semen.

Fertilization rate and embryonic mortality were assessed in 636 ewes inseminated in each uterine horn with 50 x 10(6) frozen spermatozoa from four control rams and from four rams submitted to a moderate (1.4-2.2 degrees C), but repeated, intermittent (16 h/day for 21 consecutive days) increase in their subcutaneous scrotal temperature by means of scrotal insulation. Pregnancy was assessed twice in each ewe from concentration of progesterone in blood plasma at 17 days and by ultrasound at 65 days after insemination. No differences were observed in the pregnancy rate at 17 days between ewes inseminated with semen collected from control rams (56.0, 65.2, 66.7 and 60.3%) and from heated rams (60.6, 71.8, 63.6 and 48.2%) before or after 4, 15 and 21 days of heating, respectively. In contrast, the rate of embryonic mortality between 17 and 65 days after insemination was significantly higher at days 4, 15 and 21 in the heated rams (78.7, 78.6 and 93%) than in the control rams (55, 59 and 65.7%). These results indicate that an intermittent slight, but repeated, increase in the subcutaneous scrotal temperature could induce a significant increase in the embryonic mortality rate. As these changes were apparent on day 4 of heating, an effect must have occurred on sperm stored in the epididymis.     

Seasonality of semen production and plasma luteinizing hormone,testosterone and prolactin levels in Romney,Merino and polled dorset rams

An attempt was made to define the seasonality of reproduction in rams in the southern hemisphere by repeated measurement of semen characteristics and of plasma luteinizing hormone (LH), testosterone and prolactin concentrations. These parameters were studied for 16 months in Romney rams on pasture, with Merino and Polled Dorset rams included for comparison.Semen from all three breeds showed regular seasonal changes in ejaculate volumes, with peak values being recorded during March. A similar autumnal peak of seminal fructose levels was noted for ejaculates from Romney and Merino rams, but not for those from Polled Dorsets. Most of the other semen parameters measured showed little tendency for seasonal variations. However, a change in semen collection technique, from predominantly artificial vagina to entirely electroejaculation, may have masked some seasonal changes.Plasma hormone levels also varied in a regular manner, with peak levels occurring in summer and autumn: highest levels for prolactin were recorded in November–March, for LH in December–February and for testosterone in January–March. An exception to this pattern was recorded from the Merino rams, for which there was no definite peak of LH secretion.It is suggested that these seasonal changes resulted primarily from changes in daily photoperiod.     

Semen characteristics, scrotal circumference and bacterial isolates of fine wool range rams

During 1983, 887 fine wool rams were subjected to breeding soundness evaluation to determine effects of age and scrotal circumference on semen characteristics. Of rams evaluated, 94.4, 84.8, and 81.9% of young (6 yr), respectively, were rated as satis-factory. Old rams had fewer (P < 0.05) motile cells and more (P < 0.05) abnormal cells than young rams. Among older rams there was a higher percentage (P < 0.05) with testicular lesions than among young or mature rams. Scrotal circumference was positively correlated (P < 0.01) with semen volume and percentage of motile normal cells. Motility was negatively correlated (P < 0.01) with percentage of abnormal cells. Mature and old rams with large (>/=36.7 cm) testis had more (P < 0.10) abnormal cells than those in the same age groups with smaller testis. Scrotal circumference was positively correlated (P < 0.10) to volume and motility in mature and old rams, while motility was negatively correlated (P < 0.01) to percentage of abnormal cells in all age groups. Previous semen testing reduced (P < 0.05) the percentage of mature rams with leucocytes. Vaccination against epididymitis reduced (P < 0.05) the incidence of mature rams with leucocytes and testicular lesions. Brucella ovis was recovered from 54 (67.5%) of 80 ejaculates cultured. Among rams infected with B . ovis , only three (5.6%) were vaccinated against B . ovis and had their semen tested previously.     

Effect of thermal exposure on physiological adaptability and seminal attributes of rams under semi-arid environment

Thermal stress in hot semi-arid environment is a major limitation of sheep production in tropical and sub-tropical climatic condition. The animals tend to maintain homeostasis through physiological adjustments in a hot environment (maximum temperature reaches up to 47.5 °C). Therefore, the present study was carried out to assess the effect of thermal exposure on physiological adaptability and seminal attributes of rams under semi-arid environment. The experiment was conducted for eight weeks involving sixteen Malpura crossbred rams (GMM: Garole X Malpura X Malpura). The rams were divided equally into two groups, designated as G1 and G2, respectively. The rams in G1 (Control) group were kept in a sheep shed under naturally prevailing environment without artificial manipulation of ambient temperature (Temperature 30.48±0.38 °C; Relative Humidity 28.59±1.15%). The rams of G2 group were exposed to different temperature at different hours of the day (38 °C at 1000–1100 h; 40 °C at 1100–1200 h; 42 °C at 12:00–1300 h; 43 °C at 1300–1400 h; 44 °C at 1400–1500 h and 42 °C at 1500–1600 h) in a climatic chamber for thermal exposure. Physiological responses, blood biochemical profile, blood endocrine profile, sexual behavior and seminal attributes were measured for both the groups. Thermal exposure significantly (P<0.05) increased the water intake; respiration rate, rectal temperature and skin temperature at afternoon in rams. Exposure of rams to thermal stress (G2) significantly (P<0.05) increased cortisol level and decreased tri-ido-thyronine level. The latency period after the first ejaculation, decreased significantly (P<0.05) in G2. The percentage of rapid motile sperm, linearity and average path velocity of sperm were also altered significantly (P<0.05) in thermal exposed rams as compared to control. However, comparable feed intake, body weight, and major blood biochemical parameters, as well as acceptable semen quality attributes of all the rams indicated that the Fec B gene introgressed Malpura cross rams adapted to the thermal exposure under semi-arid tropical climate.     

Effects of scrotal insulation in Nellore bulls (Bos taurus indicus) on seminal quality and its relationship with in vitro fertilizing ability

The objectives of this study were to assess the effects of induced testicular degeneration in Bos taurus indicus (Nellore) bulls on changes in seminal characteristics and fertilizing ability of sperm. Four Nellore bulls (30–36-month-old, 500–550 kg) with good seminal quality (>80% motile and morphologically normal sperm) had scrotal insulation applied for 5 d. Semen was collected by electroejaculation and cryopreserved at the pre-insulation moment, and 7, 14, and 21 d after insulation was removed. Gross motility, vigor of sperm movement (1–5), acrosome integrity, sperm morphology (phase-contrast microscopy), nuclear vacuoles and abnormal chromatin (Feulgen-stain) were determined after sperm preparations for in vitro fertilization (IVF). Prior to IVF, sperm were separated using a Percoll gradient (45% and 90%). Normal sperm decreased (P < 0.05) 14 and 21 d after insulation was removed. On 14 and 21 d, the incidence of head defects (9.7 ± 0.6 and 17.0 ± 0.8, respectively; mean ± S.E.M.) was higher (P < 0.05) in agreement with the incidence of nuclear vacuoles (14.0 ± 5.0 and 12.3 ± 2.3) and abnormal chromatin (24.4 ± 7.2 and 30.8 ± 2.8). Although the frequency of cleaved oocytes decreased only on 21 d (P < 0.05), blastocyst rates were lower (P < 0.05) than pre-insulation on 14 and 21 d. In regression analyses, only nuclear vacuoles, head defects and intact acrosome accounted for differences in cleavage (R² = 0.38, 0.48, and 0.30, respectively) and blastocyst rates (R² = 0.35, 0.37, and 0.44). Abnormal chromatin was associated only with blastocyst rates (R² = 0.35). In conclusion, blastocyst rate was more sensitive than cleavage rate and the assessment of nuclear integrity is recommended to predict the fertilizing ability of bull sperm.     

Effects of scrotal insulation in Nellore bulls (Bos taurus indicus) on seminal quality and its relationship with in vitro fertilizing ability

The objectives of this study were to assess the effects of induced testicular degeneration in Bos taurus indicus (Nellore) bulls on changes in seminal characteristics and fertilizing ability of sperm. Four Nellore bulls (30–36-month-old, 500–550 kg) with good seminal quality (>80% motile and morphologically normal sperm) had scrotal insulation applied for 5 d. Semen was collected by electroejaculation and cryopreserved at the pre-insulation moment, and 7, 14, and 21 d after insulation was removed. Gross motility, vigor of sperm movement (1–5), acrosome integrity, sperm morphology (phase-contrast microscopy), nuclear vacuoles and abnormal chromatin (Feulgen-stain) were determined after sperm preparations for in vitro fertilization (IVF). Prior to IVF, sperm were separated using a Percoll gradient (45% and 90%). Normal sperm decreased (P < 0.05) 14 and 21 d after insulation was removed. On 14 and 21 d, the incidence of head defects (9.7 ± 0.6 and 17.0 ± 0.8, respectively; mean ± S.E.M.) was higher (P < 0.05) in agreement with the incidence of nuclear vacuoles (14.0 ± 5.0 and 12.3 ± 2.3) and abnormal chromatin (24.4 ± 7.2 and 30.8 ± 2.8). Although the frequency of cleaved oocytes decreased only on 21 d (P < 0.05), blastocyst rates were lower (P < 0.05) than pre-insulation on 14 and 21 d. In regression analyses, only nuclear vacuoles, head defects and intact acrosome accounted for differences in cleavage (R² = 0.38, 0.48, and 0.30, respectively) and blastocyst rates (R² = 0.35, 0.37, and 0.44). Abnormal chromatin was associated only with blastocyst rates (R² = 0.35). In conclusion, blastocyst rate was more sensitive than cleavage rate and the assessment of nuclear integrity is recommended to predict the fertilizing ability of bull sperm.