Effect of small doses of naloxone on sexual exhaustion in White New Zealand male rabbits

The objective of the present study was to determine the effect of small doses of naloxone on sexual exhaustion in White New Zealand male rabbits. Twelve young and 12 adult male rabbits 6–12 months old and 14–20 months of age, respectively, were selected from a commercial farm. Each male rabbit was housed individually in galvanized cages (90 cm × 60 cm × 40 cm). The rabbits were housed in an open shed exposed to natural photoperiod (12 L 12 D, 19°N). Daily temperature fluctuated through the year from 28 to 16 °C. Humidity was 45 ± 5%. Water and food (rabbit chow PMI) was supplied ad libitum. After sexual behaviour for each studied group was established, the males were given a 6-day rest, and 3 days before next trial, six males of each group (treated) received a subcutaneous implant of 8 mg of naloxone in a crystalline nitrocellulose pellet formulated to be completely absorbed in 15 days. The remaining six males were sham-treated (control). At the end of the resting period as previously described, the sexual behavior of each group was studied and compared using a Mann–Whitney statistical U-test. The effect of naloxone on sexual behavior was analyzed with a Wilcoxon test for correlated samples. With regard to sexual activity between young and adult rabbits, it was observed that there was a significant difference between groups (P = 0.00275, Z = 2.8823, adjusted Z = 2.99.43) showing that younger rabbits mounted/ejaculated from 9 to 10 females compared with 6 to 8 mounted/ejaculated by older rabbits. When naloxone was administered to both groups, there was a significant difference when comparing sexual behavior before and after administration of naloxone (table first and second trial). Young rabbits treated with naloxone mounted/ejaculated 11–12 females while older rabbits mounted nine females before reaching sexual exhaustion. A significant difference was observed when comparing the number of estrous females that were mounted/ejaculated between groups. Environmental photoperiod and temperature changes were not considered. It was concluded that endogenous opioids are important modulators of behavioral and hormonal interactions related to sexual behavior.     

The effect of a combination of permanent breeding cage and low housing density on the reproductive success of farmed blue foxes

Social factors are known to affect the reproduction of many canids both in the wild and in farms. For example, reproduction in farmed silver foxes is regulated by social stress; foxes seem to benefit from noncramped housing conditions and permanent breeding cages. However, no comparable studies have been carried out in farmed blue foxes.

The aim of our experiment was to create an alternative, improved, economically viable and practical housing solution for blue foxes. Therefore, we compared reproductive performance of blue foxes in permanent breeding cages with low animal densities (L group, N = 79) and traditional housing with its changing social environment with high animal density (H group, N = 74). The reproductive data from the L and H groups were compared separately for primiparous and multiparous vixens because the reproductive performance in primiparous vixens was substantially lower (P < 0.001) than in multiparous vixens.

Altogether, 41 and 39% of the primiparous vixens in the H and L group whelped (P > 0.05), but only 28 and 34%, respectively, weaned at least one cub (P > 0.05), i.e., 72 and 66% of the primiparous vixens did not reproduce in the H and L group, respectively (P > 0.05). The total reproductive performance, expressed as cubs at weaning per breeding female, was 1.7 ± 3.5 for the H and 1.6 ± 2.9 for the L group (P > 0.05). In the primiparous vixens, the only statistically significant difference observed between the two housing systems was that the onset of oestrus occurred five days earlier in the H than in the L group (P < 0.05).

All multiparous vixens in the L group exhibited oestrus compared to 94% in the H group (P > 0.05). Furthermore, there was a nonsignificant (ns) trend for fewer barren females (9% versus 17%), more successfully reproducing vixens (83% versus 74%) and a higher number of live-born cubs (10.9 ± 4.7 versus 9.4 ± 3.9) in the L than in H group in the multiparous vixens (for all P > 0.05). This resulted in 1.7 and 1.4 cubs more per breeding and per mated vixen, respectively, at weaning in the L group (7.3 ± 5.0) compared to the H group (5.6 ± 4.2), but also this difference was nonsignificant.

Although our present results lack statistical significance, they are promising enough to encourage field experiments with sufficiently large number of animals to prove or disprove these preliminary findings that lower housing density and permanent breeding cage, together or separately, may enhance reproduction particularly in multiparous blue fox vixens.     

Body condition and protein supplementation positively affect periovulatory ovarian activity by non LH-mediated pathways in goats

Effects of rumen undegradable intake protein (UIP) supplementation on ovarian activity and serum insulin, GH, and LH were evaluated in goats having low or high body condition (BC). Goats with either low BC (n = 16, 28.7 ± 0.8 kg BW, BC = 2.1 ± 0.3) or high BC (n = 16, 38.4 ± 0.8 kg, BC = 3.2 ± 0.3) received, during 40-days, one of the two protein supplementation levels: without UIP or with UIP (120 g goat⁻¹ d⁻¹). Oestrus was synchronized with two i.m. doses of PGF₂, and jugular blood samples were collected from 36 to 42 h after the second prostaglandin injection at 15 min intervals. Serum concentrations of insulin, LH, and GH were measured The number of preovulatory follicles and the number of corpora lutea (CL) were evaluated by transrectal ultrasonography at 1 and 4 days after the second prostaglandin dose, respectively. Does with higher BC had more CL than those in the lower condition group (2.8 ± 0.2 versus 1.8 ± 0.2, P < 0.05). Similarly, goats receiving UIP supplementation had more follicles (2.6 ± 0.2 versus 1.9 ± 0.2, P < 0.05) and tended to have more CL (2.6 ± 0.2 versus 2.0 ± 0.2, P = 0.05) than does not receiving UIP. Neither BCS nor UIP supplementation affected serum GH or LH concentrations, pulsatility, or area under the curve. High BC does produced more insulin (1.92 ± 0.17 versus 0.81 ± 0.17 ng/mL, P < 0.01 ng/mL) than lower BC goats; the same for UIP-supplemented (1.69 ± 0.18 versus 1.04 ± 0.18, P < 0.05). Results suggest that the increased ovarian activity observed in both UIP-supplemented and higher BC goats was not the result of changes in LH or GH, suggesting effects at a local level, through changes in insulin in a non-GnRH-gonadotrophin dependent manner.     

Relationship between male-male and male-female sexual behavior in 5-6-month-old male lambs

To characterize male–male sexual behavior during lamb development, to relate it with lamb body and testicular growth, and with sexual behavior toward estrual ewes, 40 Milchschaf male lambs, weaned at 45 days of age, were kept with ewes that were nursing younger lambs. Experimental lambs were weighed and scrotal circumference was measured every 2 weeks. Male–male sexual behavior was observed during 1–2 h every 2 weeks after birth until 7 months of age. Observations were recorded more intensively (3–4 h on five different days) for 2 weeks (5–6 months of age) as male–male sexual behavior increased during that period. Both mounting and mounted lambs were identified. An individual mounting index (MI) was calculated. To study male–female sexual behavior, lambs were individually located with two estrual ewes, and during 5 min the number of ano-genital sniffing, lateral approaches, mounts, and mounts with ejaculation were recorded. From those data, a libido index was also calculated. Male–male mounts (n = 308) were observed. Courtship behavior was displayed in 25% of interactions; mounts were accepted in 72.1% of attempted mounts. Mounts without previous courtship were accepted more frequently than mounts with previous courtship (P = 0.002). Lamb weight and scrotal circumference were not different according to MI groups. Lambs that mounted more times estrual ewes (first tertile) had greater (P = 0.04) MI (0.61 ± 0.10) than lambs with medium (0.27 ± 0.09) and less (0.30 ± 0.10) MI. The regression between MI and heterosexual libido index was r = 0.33 (P < 0.05). In summary, intensive male–male sexual activity during a short period of male lamb development was observed. There was a positive relationship between sexual behavior of male lambs towards other male lambs and towards estrual ewes.     

Prolactin regulation of testicular development and sexual behavior in yearling Suffolk rams

The aim of this study was to determine how the yearly prolactin rhythm might affect the sexual development of Suffolk rams (latitude 50°N). Five rams were injected daily with bromocriptine (35–45 μg kg⁻¹ body weight) for 1 year, beginning in January (early winter) when rams were 11 months of age. Five control rams each received daily injections of the vehicle. In the controls, blood prolactin was <7.5 ng ml⁻¹ in winter, increased (P < 0.01) to a peak of 172.6 ± 11.9 ng ml⁻¹ after the spring equinox, and remained high during summer before declining (P < 0.01) to 29.6 ± 6.6 ng ml⁻¹ at the autumn equinox. Suppression of the seasonal rise in prolactin secretion with bromocriptine slowed testicular growth (50%; P < 0.05) in April and May (spring), thus delaying the time of peak testis size and sperm production by 1 month. Serum testosterone level was lower (50%; P < 0.01) in the treated rams than the controls in June and July (early summer), due mainly to reduced stimulation of the testes by smaller (P < 0.01) LH pulse releases or to smaller (P < 0.01) testosterone responses to LH releases, respectively. Suppression of prolactin also seemed to disrupt the central activation of gonadotropin secretion in that seasonal increases in serum FSH level and LH pulse amplitude and frequency were unusually slow (P < 0.05). These anomalies did not affect testis growth, which was normal from June until development was complete. Rams were sexually inexperienced when libido was first tested in July (non-breeding season). Both groups were equally capable of learning and expressing sexual behavior (i.e. normal mounting and ejaculation frequencies), which was more intense in September (breeding season; P < 0.05). Results support the hypothesis (based on the location of prolactin receptors) that the spring increase in prolactin secretion could target both the testes and the hypothalamic–pituitary system and be involved in the seasonal regulation of sexual function in the young adult Suffolk ram.     

Examination of cyclic changes in bovine luteal echotexture using computer-assisted statistical pattern recognition techniques

B-mode sonography is a well-established diagnostic tool for determination of cycle stage in gynaecology. The aim of this study was to determine whether computer-assisted texture analysis of B- mode sonographic images of bovine luteal glands provides further information about the animal's plasma progesterone concentration and cycle stage. Four Simmenthal cows were examined during two consecutive estrous cycles with an ultrasound device equipped with a 7.5 MHz microconvex probe. During each examination three B-mode images of the corpus luteum (CL) were digitized and analyzed off-line using a computer-assisted texture analysis program. Size, echogeneity, and echotexture of the CL were characterized by the following texture parameters: area of cross-sectional planes of the CL (A), mean gray level (MGL), correlation (CORR), run percentage (RPERC), and long-run emphasis (LREM). Plasma progesterone levels (P4) were also determined. All parameters showed characteristic changes during the estrous cycle (P < 0.05). Variance component estimates for the effect of Day of estrous cycle on A, MGL, CORR, RPERC, and LREM were 56.6%, 64.6%, 77.6%, 89.9%, and 86.0%, respectively, and 20.6%, 24.5%, 7.2%, 0.0%, and 14.0% for the influence of the individual cow. The factor estrous cycle within cows was responsible for 22.8%, 10.9%, 15.2%, 10.1%, and 0.0% of the variability of A, MGL, CORR, RPERC and LREM values, respectively. Cyclic changes were similar in A and P4. In contrast to P4, which decreased already between Days –5 and –3 (Day 0 = ovulation), A stayed at constant high values until Day –3. Mean MGL values were higher (P < 0.05) on Days 7, 9, and 13 compared to Days 3 and –3. Mean CORR values were constantly high (P > 0.05) during the first days after ovulation and decreased continuously (P < 0.05) between Days 5 and 13. Thereafter, mean CORR values remained low (P < 0.05) until the next ovulation, except on Day –3 (P < 0.05). Mean RPERC rose between Days 1 and 9 from low to high values (P < 0.0001) remained at these high values (P > 0.05) between Days 9 and 15, and decreased (P < 0.05) afterwards to baseline values on Day –1. Mean LREM inclined steeply (P < 0.0001) from minimum to maximum between Days 1 and 5. From Days 7 to –3, mean LREM remained (P > 0.05) at a constant level close below the maximum value, and decreased to baseline values on Day –1. The results of this study show that statistical pattern recognition techniques provide new information about the luteal glands, thus facilitating a more accurate differentiation between different cycle stages in cows.     

Control of postpartum anestrous with an intra-vaginal progesterone device plus eCG or calf removal for 120 h in suckled crossbred cows managed in a pasture-based system

To control postpartum anestrus and reduce calving to conception interval, 167 crossbred non-pregnant cows that were 90–130 days postpartum were allotted randomly to one of the following treatments: PH (n = 59), intra-vaginal sponge with 250 mg of medroxyprogesterone acetate (MAP) for 7 days plus 50 mg of MAP and 5 mg 17-β estradiol (17β-E) in the first day of treatment (day −8), 500 UI eCG (day −3) and 1.5 mg 17β-E in 24 h after sponge removal (day 0); CR (n = 57), temporary calf removal for 120 h; CG (n = 51), control group without treatment. Estrus rate differed among treatments (P < 0.01) being greater in PH (78.2%), followed by CR (52.0%) and CG (22.9%). A greater proportion of cows in the PH (80.0%) and CR (54%) groups had ovulations when compared to CG (35.4%). Intervals to first estrus were 13.5 ± 6.3 days, 26.1 ± 6.4 days and 52.5 ± 7.5 days for the PH, CR and CG groups, respectively. First insemination conception was similar in the three groups. Postpartum intervals to first breeding (PFS) and to conception (PCI) were longer in CG than PH and CR groups (P < 0.05; P < 0.01). The PH and CR groups had a similar PFS but PCI was different (P < 0.02). Accumulated pregnancy rate at 30 and 60 but not at 90 days were different (30 days: P < 0.09; P < 0.01; P < 0.09; 60 days: P < 0.06; P < 0.01; P < 0.03) among treatments. After 90 days post-treatment, 9%, 18% and 33% of cows from the PH, CR and CG groups had not conceived. Similarly, 5.4%, 6.0% and 12.5% of cows from the PH, CR and CG groups, respectively, were culled from the herd because of lack of pregnancy after 180 days post treatment. In the group of cows evaluated by ultrasonography, only those cows having larger ovaries and dominant follicles had ovulations. It was concluded that the hormonal treatment was more efficient in inducing a fertile estrus and reducing calving to conception interval followed by the calf removal for 120 h. Each method can be considered as an important tool to reduce the postpartum anestrous period in dual purpose herds when AI is conduct in the tropics.     

Different effects of ATP on the contractile activity of mice diaphragmatic and skeletal muscles

Apart from acetyl-choline (Ach), adenosine-5′-trisphosphate (ATP) is thought to play a role in neuromuscular function, however little information is available on its cellular physiology. As such, effects of ATP and adenosine on contractility of mice diaphragmatic and skeletal muscles (m. extensor digitorum longa—MEDL) have been investigated in in vitro experiments. Application of carbacholine (CCh) in vitro in different concentrations led to pronounced muscle contractions, varying from 9.15 ± 4.76 to 513.13 ± 15.4 mg and from 44.65 ± 5.01 to 101.46 ± 9.11 mg for diaphragm and MEDL, respectively. Two hundred micromolars of CCh in both muscles caused the contraction with the 65% (diaphragm) to 75% (MEDL) of maximal contraction force—this concentration was thus used in further experiments. It was found that application of ATP (100 μM) increased the force of diaphragmatic contraction caused by CCh (200 μM) from 335.2 ± 51.4 mg (n = 21) in controls to 426.5 ± 47.8 mg (n = 10; P < 0.05), but decreased the contractions of MEDL of CCh from 76.6 ± 6.5 mg (n = 26) in control to 40.2 ± 9.0 mg (n = 8; P < 0.05). Application of adenosine (100 μM) had no effect on CCh-induced contractions of these muscles.

Resting membrane potential (MP) measurements using sharp electrodes were done at 10, 20 and 30 min after the application of ATP and adenosine. Diaphragm showed depolarization from 75 ± 0.6 down to 63.2 ± 1.05, 57.2 ± 0.96 and 53.6 ± 1.1 mV after 10, 20 and 30 min of exposition, respectively (20 fibers from 4 muscles each, P < 0.05 in all three cases). Adenosine showed no effect on diaphragmatic MP. Both agents were ineffective in case of MEDL.

The effects of ATP in both tissues were abolished by suramin (100 μM), a P2-receptor antagonist, and chelerythrin (50 μM), a specific protein-kinase C (PKC) inhibitor, but were not affected by 1H-[1,2,4]-oxadiazolo-[4,3-]-quinoxalin-1-one (ODQ, 1 μM), a guanylyl-cyclase inhibitor, or by adenosine-3,5-monophosphothioate (Rp-cAMP, 1 μM), a protein-kinase A (PKA) inhibitor.

Besides the action on contractile activity, ATP (100 μM) led to a significant (P < 0.001) depolarization of diaphragm muscle fibers from 74.5 ± 2.3 down to 64 ± 2.1, 58.2 ± 2.2 and 54.3 ± 2.4 mV after 10, 20 and 30 min of incubation, respectively. Incubation of MEDL with the same ATP concentration showed no significant change of MP.

Denervation of the muscles for 28 days led to a decrease of CCh-induced contractions of diaphragm down to 171.1 ± 34.5 mg (n = 11, P < 0.05), but increased the contractile force of MEDL up to 723.9 ± 82.3 mg (n = 9, P < 0.01). Application of ATP elevated the contractility of denervated diaphragm caused by CCh up to normal values (311.1 ± 79.7 mg, n = 6, P > 0.05 versus control), but did not significantly affect of contractility of MEDL, which became 848.1 ± 62.7 mg (n = 6).

These results show that the effects of ATP on both diaphragmatic and skeletal muscles are mediated through P2Y receptors coupled to chelerytrin-sensitive protein-kinase C.     

The relationship between heart rate and energy expenditure in Alpine, Angora, Boer and Spanish goat wethers consuming different quality diets at level of intake near maintenance or fasting

Six Alpine (AL; 38.4 ± 3.0 kg), Angora (AN; 23.1 ± 2.7 kg), Boer (BO; 40.8 ± 4.5 kg) and Spanish (SP; 33.6 ± 2.2 kg) wethers (1.5 yr of age) were used to determine the effects of time of the day and potential interactions between time, genotype and diet quality on energy expenditure (EE), heart rate (HR) and EE:HR when fed near maintenance and fasting. The experiment consisted of four simultaneous crossovers, with 21 d for adaptation before measures. Diets were 60% concentrate (CON: 15% CP) and ground alfalfa hay (FOR: 23% CP), offered in two meals at 8:00 and 16:00 h. Energy expenditure was determined from O₂ consumption and production of CO₂ and CH₄ over 2-day periods in fed and fasting states (total 4-day fasting period). Fasting EE was higher during the day than night, with values generally highest at 16:00–17:00 h. Animal within breed affected EE, HR and EE:HR (P < 0.05). The diurnal pattern in EE varied with diet (P < 0.05), although total daily EE was not different between diets. Before the morning meal, there were a number of hours during which EE was greater for CON than for FOR. However, at both meals the rise in EE was considerably greater for FOR versus CON, lasting for 3–4 h. The same general pattern in HR was observed, although the period of time when there was a dietary difference after the afternoon meal was shorter. For both fed and fasted goats, EE:HR differed among hours of the day (P < 0.05). EE:HR tended (P < 0.09) to differ between diets (5.99 and 6.21 for CON and FOR, respectively) and to be affected (P < 0.09) by an interaction between breed and diet (AL: 5.84 and 6.38; AN: 5.91 and 5.73; BO: 6.05 and 6.58; and SP: 6.17 and 6.15 kJ/(kg BW^0.75 × day):heart beats/min) for CON and FOR, respectively. In conclusion, for use of HR to predict EE by goats, it appears desirable to determine the ratio of EE:HR with a diet similar to that consumed during prediction and over an extended period of time.     

Interaction of calf suckling, use of oxytocin and milk yield with reproductive performance of dairy buffaloes

Calf suckling and oxytocin injections are commonly used for pre-milking stimulus in dairy buffaloes under field conditions. A study was conducted to investigate effect of these treatments on reproductive performance. Fifty one Nili-Ravi buffaloes were monitored from parturition up to 150 days postpartum through rectal examination. Data on milk yield, body condition score (BCS) and reproductive parameters were recorded weekly. Postpartum ovulation interval (POI) was determined by presence of an ovulation depression or a very soft corpus luteum haemorrhagicum and was confirmed through milk progesterone levels (MPL). Suckling was used to stimulate milk let down, and where the calf had died, injection of oxytocin was resorted to. Milk samples were analyzed for MPL using radioimmunoassay (RIA) and fat; and milk yield was converted to 4% fat corrected milk (FCM). The mean postpartum uterine involution length (PUI) was 34.30 ± 1.33 days. Mean POI was 59.37 ± 4.76 days and mean postpartum estrus interval (PEI) was 69.03 ± 6.03 days. Suckling period averaged 26.40 ± 5.57 days and correlated with POI (r = 0.19, P < 0.01) and PEI (r = 0.23, P < 0.01). POI was shortest in buffaloes suckled for one month (P < 0.05). Oxytocin was used with a mean dosage of 7.50 IU, delaying placental expulsion time (PET) and POI but shortening PEI. BCS shortened PET, POI and PEI (P < 0.01). Mean FCM was 14.50 ± 0.20, ranging from 2 to 35 kg/d; and was higher in estrus group; correlating positively with POI (r = 0.31, P < 0.01). MPL were 1.37 ± 0.17 ng/ml and increased after ovulation, remaining greater than 1.5 ng/ml from Day 4 to 14 of the estrus cycle, followed by a rapid decline up to next estrus. BCS in buffaloes resuming oestrus was constantly higher than those failing to resume ovarian cyclicity. Live weight, prepartum was 510.0 ± 5.9 kg with a loss of 3.7 ± 2.12 kg, 30 days postpartum. The present study suggests a lower reproductive efficiency of dairy buffaloes under the peri-urban farming system reflected by ovarian cyclicity in 68.63% buffaloes within 150 days postpartum and silent estrus in 51.5% of the cases. Increasing suckling duration and use of oxytocin delayed POI, however, POI was shortest in buffaloes suckled for one month. The high yielding buffaloes also manifested better reproductive cyclicity; while moderate yielder showed shorter ovulation intervals and higher conception rate.     

Chromosomal aberrations in tire plant workers and interaction with polymorphisms of biotransformation and DNA repair genes

We evaluated chromosomal aberrations in lymphocytes of 177 workers exposed to xenobiotics in a tire plant and in 172 controls, in relation to their genetic background. Nine polymorphisms in genes encoding biotransformation enzymes and nine polymorphisms in genes involved in main DNA repair pathways were investigated for possible modulation of chromosomal damage. Chromosomal aberration frequencies were the highest among exposed smokers and the lowest in non-smoking unexposed individuals (2.5 ± 1.8% vs. 1.7 ± 1.2%, respectively). The differences between groups (ANOVA) were borderline significant (F = 2.6, P = 0.055). Chromosomal aberrations were higher in subjects with GSTT1-null (2.4 ± 1.7%) than in those with GSTT1-plus genotype (1.8 ± 1.4%; F = 7.2, P = 0.008). Considering individual groups, this association was significant in smoking exposed workers (F = 4.4, P = 0.040). Individuals with low activity EPHX1 genotype exhibited significantly higher chromosomal aberrations (2.3 ± 1.6%) in comparison with those bearing medium (1.7 ± 1.2%) and high activity genotype (1.5 ± 1.2%; F = 4.7, P = 0.010). Both chromatid- and chromosome-type aberration frequencies were mainly affected by exposure and smoking status. Binary logistic regression analysis revealed that frequencies of chromatid-type aberrations were modulated by NBS1 Glu185Gln (OR 4.26, 95%CI 1.38–13.14, P = 0.012), and to a moderate extent, by XPD Lys751Gln (OR 0.16, 95%CI 0.02–1.25, P = 0.081) polymorphisms. Chromosome-type aberrations were lowest in individuals bearing the EPHX1 genotype conferring the high activity (OR 0.38, 95%CI 0.15–0.98, P = 0.045). Present results show that exposed individuals in the tire production, who smoke, exhibit higher chromosomal aberrations frequencies, and the extent of chromosomal damage may additionally be modified by relevant polymorphisms.     

PAH-DNA adducts in environmentally exposed population in relation to metabolic and DNA repair gene polymorphisms

Epidemiologic studies indicate that prolonged exposure to particulate air pollution may be associated with increased risk of cardiovascular diseases and cancer in general population. These effects may be attributable to polycyclic aromatic hydrocarbons (PAHs) adsorbed to respirable air particles. It is expected that metabolic and DNA repair gene polymorphisms may modulate individual susceptibility to PAH exposure. This study investigates relationships between exposure to PAHs, polymorphisms of these genes and DNA adducts in group of occupationally exposed policemen (EXP, N = 53, males, aged 22–50 years) working outdoors in the downtown area of Prague and in matched “unexposed” controls (CON, N = 52). Personal exposure to eight carcinogenic PAHs (c-PAHs) was evaluated by personal samplers during working shift prior to collection of biological samples. Bulky-aromatic DNA adducts were analyzed in lymphocytes by ³²P-postlabeling assay. Polymorphisms of metabolizing (GSTM1, GSTP1, GSTT1, EPHX1, CYP1A1-MspI) and DNA repair (XRCC1, XPD) genes were determined by PCR-based RFLP assays. As potential modifiers and/or cofounders, urinary cotinine levels were analyzed by radioimmunoassay, plasma levels of vitamins A, C, E and folates by HPLC, cholesterol and triglycerides using commercial kits. During the sampling period ambient particulate air pollution was as follows: PM10 32–55 μg/m³, PM2.5 27–38 μg/m³, c-PAHs 18–22 ng/m³; personal exposure to c-PAHs: 9.7 ng/m³ versus 5.8 ng/m³ (P < 0.01) for EXP and CON groups, respectively. The total DNA adduct levels did not significantly differ between EXP and CON groups (0.92 ± 0.28 adducts/10⁸ nucleotides versus 0.82 ± 0.23 adducts/10⁸ nucleotides, P = 0.065), whereas the level of the B[a]P-“like” adduct was significantly higher in exposed group (0.122 ± 0.036 adducts/10⁸ nucleotides versus 0.099 ± 0.035 adducts/10⁸ nucleotides, P = 0.003). A significant difference in both the total (P < 0.05) and the B[a]P-“like” DNA adducts (P < 0.01) between smokers and nonsmokers within both groups was observed. A significant positive association between DNA adduct and cotinine levels (r = 0.368, P < 0.001) and negative association between DNA adduct and vitamin C levels (r = −0.290, P = 0.004) was found. The results of multivariate regression analysis showed smoking, vitamin C, polymorphisms of XPD repair gene in exon 23 and GSTM1 gene as significant predictors for total DNA adduct levels. Exposure to ambient air pollution, smoking, and polymorphisms of XPD repair gene in exon 6 were significant predictors for B[a]P-“like” DNA adduct. To sum up, this study suggests that polymorphisms of DNA repair genes involved in nucleotide excision repair may modify aromatic DNA adduct levels and may be useful biomarkers to identify individuals susceptible to DNA damage resulting from c-PAHs exposure.     

Effects of anti-oxidant additives on microscopic and oxidative parameters of Angora goat semen following the freeze–thawing process

The anti-oxidant system of reduced glutathione (GSH), glutathione peroxidase (GSH-PX), catalase (CAT), and superoxide dismutase (SOD) has been described as a defense functioning mechanism against lipid peroxidation (LPO) in semen, and is important in maintaining sperm motility and viability. This anti-oxidant capacity of sperm cells may be insufficient in preventing LPO during the freeze–thawing process. The aim of this study was thus to determine the influence of varying doses of anti-oxidant additives on standard semen parameters, lipid peroxidation and anti-oxidant activities after the freeze–thawing of goat semen. Ejaculate samples (artificial vagina) obtained from 4 mature Angora goats were evaluated and pooled at 37 °C. The semen samples diluted with a Tris-based extender, containing taurine (25, 50, 75 mM), trehalose (25, 50, 75 mM), and cysteine (5, 10, 15 mM), and an extender containing no anti-oxidant additives (control) were again evaluated. Diluted semen was cooled down to 5 °C and frozen in 0.25 ml French straws, prior to being stored in liquid nitrogen. Frozen straws were thawed in a water bath (37 °C) for 30 s for microscopic sperm evaluation. Upon evaluation of parameters for semen quality, the use of a Tris-based extender supplemented with anti-oxidant additives was found to cause no significant improvement in sperm mortality, when compared to the controls. Increasing doses of taurine and trehalose decreased (P < 0.05) the sperm motility following the freeze–thawing of the goat semen. In biochemical assays, the application of taurine (75 mM) produced the lowest level of malondialdehyde (MDA) (4.46 ± 0.31 nmol/ml), compared to the controls (P < 0.001). Lower GSH levels were higher in the groups in which cysteine was included at 10 and 15 mM (3.27 ± 0.11 and 3.45 ± 0.28 nmol/ml) – compared to the group which received 5 mM cysteine, as well as the controls (2.27 ± 0.08 and 2.50 ± 0.08 nmol/ml respectively, P < 0.001). Compared to the controls, taurine at a concentration of 25 and 75 mM, and increasing doses (50 and 75 mM) of trehalose, significantly increased the GSH-PX activity (P < 0.01). The maintenance of CAT activity was demonstrated to be higher with the addition of 10 and 15 mM cysteine, compared to the other groups (P < 0.001). Vitamin A (VitA) levels were significantly higher, compared to the controls (267.34 ± 9.68 mg/dl and 267.34 ± 9.68 mg/dl, respectively), when 25 mM taurine (329.61 ± 6.35 mg/dl) and 10 mM (318.64 ± 6.34 mg/dl) cysteine was added to the extender (P < 0.001). The results of this study provide a new approach to the cryopreservation of Angora goat semen and could contribute to the improvement of this technology in the goat industry.     

Development to the blastocyst stage of immature pig oocytes arrested before the metaphase-II stage and fertilized in vitro

In vitro fertilization (IVF) and embryonic development of mature and meiotically arrested porcine oocytes were compared in the present study. After in vitro maturation (IVM) of cumulus-oocyte complexes for 48 h, 75.4% of them extruded a visible polar body (PB). Most of the oocytes with a first polar body (PB+ group) were at the metaphase-II (M-II) stage (91.4%). Most of the oocytes without a visible polar body (PB− group) appeared to be arrested at the germinal vesicle (GV) (41.6%) and metaphase-I (M-I) (34.0%) stages. After IVF of oocytes (day of IVF = Day 0), there was no difference between PB⁺ and PB⁻ groups in rates of sperm penetration, mono-spermy, however oocyte activation rate after penetration was greater in the PB+ than in the PB− group (P < 0.05). On Day 2, there was no difference between rates of embryos cleaved at the 2–4 cell stages in PB+ and PB− groups (42.1 ± 48.8% and 33.6 ± 2.1%, respectively). On Day 4, the rate of PB+ embryos developing beyond the 4-cell stage was greater than that of PB− embryos (P < 0.05, 31.7 ± 3.9% and 14.1 ± 1.5%, respectively), and PB+ embryos had more cells than the PB− embryos (P < 0.05, 8.3 ± 0.4 and 6.0 ± 0.8 cells, respectively). On Day 6, a greater proportion of PB+ embryos developed to the blastocyst stage than did PB− embryos (P < 0.05, 34.6 ± 2.4% and 20.7 ± 2.8%, respectively). However, when the GV oocytes of the PB− group were not included in recalculations, there was no difference in blastocyst rates between M-I arrested and M-II oocytes (35.3 and 34.6%, respectively). The number of blastomere nuclei in embryos obtained from the PB+ group (52.0 ± 2.5) was greater than that from the PB− group (P < 0.05, 29.1 ± 2.8). The proportion of degenerated parts in the blastocysts, as determined by morphological appearance, was the same in the PB+ and PB− groups. Although the quality of PB+ embryos was enhanced as compared with that of the PB− group, the proportion of inner cell mass and trophectoderm cells in PB+ and PB− blastocysts did not differ (1:1.9 and 1:2.2, respectively). Chromosome analysis revealed that PB+ blastocysts had more diploidy (P < 0.05, 69.7%) than did PB− blastocysts (44.0%), whereas PB− blastocysts had more triploid cells (P < 0.05, 34.0%) than did PB+ oocytes (8.4%). These results indicate that pig oocytes arrested before the M-II stage (M-I oocytes) undergo cytoplasmic maturation during maturation culture and have the same ability to develop to blastocysts after IVF as M-II oocytes, but some of them resulted in degeneration or delayed development with poor embryo quality.     

Effects of ensiled forage legumes on performance of store finishing lambs

Male Beulah speckled face lambs (initial live weight (LW) 28.8 ± 0.31 kg) were allocated to three dietary treatments to evaluate the performance of store lambs of a hill breed when offered ensiled lucerne (Medicago sativa), red clover (Trifolium pratense) or ryegrass. Second-cut silage bales (wilted and inoculated) were prepared from 3-year old lucerne and red clover stands and a 1-year old ryegrass sward. All the lambs were group-housed and offered ad libitum ryegrass silage during a 3-week co-variate period. This was followed by a week of dietary changeover period, after which the lambs were housed individually and offered their treatment diet ad libitum. All the lambs received a flat rate supplement of pelleted molassed sugarbeet (250 g fresh weight/day). Individual intakes were determined daily, and weekly measurements of LW and body condition score (CS) were made. Additional measurements were taken by scanning the lambs for depth of Longissimus dorsi (LD) muscle and subcutaneous fat. Over an experimental period of 7 weeks, the lambs offered red clover silage had a higher voluntary silage dry matter (DM) intake, total DM intake and metabolisable energy (ME) intake (P < 0.001) than lambs offered either lucerne or ryegrass silage. This resulted in a faster (P < 0.001) growth rate and increase (P < 0.001) in CS, with no difference between lucerne and ryegrass silages. The feed conversion efficiency (FCE) was 8.0 ± 0.61 kg feed/kg gain for lambs fed red clover silage, compared with 16.6 ± 2.82 and 10.6 ± 1.94 kg feed/kg gain for lucerne and ryegrass silage, respectively. The CP intake was higher (P < 0.001) for lambs fed the lucerne and red clover silages than for the ryegrass silage treatment. The concentration of plasma total protein (TP) was higher (P < 0.05) for lambs offered ryegrass silage versus lucerne and red clover silage. Urea concentrations were highest for lambs fed lucerne silage and lowest for those fed ryegrass silage (P < 0.05). The glucose concentration was higher (P < 0.05) for lambs offered red clover silage, whereas non-esterified fatty acids (NEFA) concentration was higher (P < 0.05) for lambs offered lucerne silage. Substituting ryegrass silage with red clover silage has the potential to improve the performance of finishing store lambs.     

Regrouping in lactating goats increases aggression and decreases milk production

With the aim of testing the hypothesis that regrouping decreases milk production in French Alpine goats that were lactating, a study was done using two groups (n = 8, 7). During their third month after parturition, four goats from each group were exchanged (first regrouping) between pens and left for 2 weeks, then the same two subgroups of four goats were taken back to their original pen for another 2 weeks (second regrouping). In the third regrouping, the two groups were all placed in the one pen. Milk production and social behaviour were measured daily before and after each regrouping. All regroupings led to an increase in aggressive behaviours that last by 1–2 days. Mean daily milk production decreased after first (2.82 ± 0.2 kg versus 2.53 ± 0.2 kg; P < 0.05) but not after second and third regrouping. It is concluded that aggressive behaviour increases after all regroupings, whereas milk production decreases only after the first regrouping, suggesting an important capacity of adaptation to a novel and stressful managements in the French Alpine goat. The study highlights the importance of considering effects of common practices in herd managements on social behaviour and production.     

Chromosomal aberrations by fluorescence in situ hybridization (FISH)--Biomarker of exposure to carcinogenic PAHs

The fluorescence in situ hybridization (FISH) technique with whole chromosome painting for chromosomes #1 and #4 was used to study the impact of air pollution containing higher concentrations of carcinogenic polycyclic aromatic hydrocarbons (c-PAHs) in three European cities, Prague (Czech Republic), Kosice (Slovakia) and Sofia (Bulgaria). In each site were followed an exposed group, who were police officers or bus drivers who work usually through busy streets for at least 8 h, and a reference group, who spent more than 90% of their daily time indoors.

In Prague, a significant increase was observed in percentage of aberrant cells (% AB.C.) in the police officers compared to the reference group (0.33 ± 0.25 versus 0.24 ± 0.18, p < 0.05). In Kosice, the exposed group differed from reference in the endpoints F_G/100 1.52 ± 1.18 versus 1.12 ± 1.30, p < 0.05; % AB.C. 0.30 ± 0.19 versus 0.21 ± 0.20, p < 0.05; t/1000 3.91 ± 3.18 versus 2.84 ± 3.10, p < 0.05. In Sofia were followed two exposed groups: police officers and bus drivers. All FISH endpoints were significantly higher in police officers compared to reference group (F_G/100 1.60 ± 0.99 versus 0.82 ± 0.79, p < 0.01; % AB.C. 0.25 ± 0.14 versus 0.13 ± 0.13, p < 0.01; t/1000 4.19 ± 2.65 versus 2.13 ± 2.05, p < 0.05; rcp 1.46 ± 1.07 versus 0.70 ± 0.76, p < 0.05). In bus drivers compared to reference there was an increase in % AB.C. (0.25 ± 0.18 versus 0.13 ± 0.13, p < 0.05).

This is the first study when FISH method was used to analyze the impact of environmental air pollution. According to the original hypothesis it is expected that the most important group of chemicals responsible for the biological activity of air pollution represent c-PAHs.     

Influences of fallow age and season on the foraging behavior and diet selection pattern of goats (Capra hircus L.)

A study was undertaken to determine the foraging behavior and diet selection pattern of goats maintained on communal exclosures in relation to the influence of fallow age (years since effective protection initiated; <8 years vs. >12 years) and season (long rainy, dry, and short rainy seasons). The frequency of browsing was higher (P < 0.05) in the exclosures fallowed for >12 years compared to those fallowed for <8 years and this frequency was also highest during the short rainy season. More (P < 0.05) herbaceous bites were recorded in the <8 years fallow exclosures, compared to the >12 years fallow exclosures. Herbaceous species bites were maximized during the long rainy season whereas woody species bites were maximized during the dry season. The goats foraged upon 48 plant species of which 69% were woody species, 19% grasses, and the rest comprised of various forb species. The top species that significantly (P < 0.05) contributed to the diet of goats were Acacia etbaica, Balanites aegyptica, Cadaba farinosa, Capparis tomentosa, Dichrostachys cinerea, and Ocimum forskolei. The goats targeted taller woody species than the average of the exclosures. Maurea angolensis, Jasminium abyssinicum, C. farinosa, and Ormocarpum pubescence were among the preferred species that appeared to be on the verge of disappearance in most of the exclosures investigated in this study and require immediate attention. Using goats to reduce the threat of potential bush infestation appears to be a viable option for managing and sustaining production from exclosures since the goats showed preference for D. cinerea, the species considered a prolific invader plant. There is, however, a need to conduct further research on the impacts of direct browsing by the goats and this should also entail the determination of optimum number of goats/unit area.     

Behaviour during the first stage of labour in cattle: Influence of parity and dystocia

The aim of the study was to examine typical behavioural elements in suckling cows and heifers at the first stage of labour and how these are affected by parity and dystocia. For this purpose, the parturitions of 87 cattle (10 Simmental heifers; 77 multiparous cows, 55 Simmental, 21 Simmental × Limousin) were observed from the appearance of the amniotic sacs in the cervix up to the emergence of the foetal phalanges in the rima vulvae. The animals were divided into three groups: group 1—cows with eutocia, n = 68; group 2—heifers with eutocia, n = 10; group 3—cows with dystocia, n = 9.

Compared to cows, there was a lower proportion of heifers with calm behaviour, whereas a higher proportion of heifers showed pawing with the forefeet (P < 0.05).

There was a higher proportion of cows with dystocia which showed rubbing against the wall (P < 0.05), discharge of urine (P < 0.05) and scraping on the floor (P < 0.05) compared to cows with eutocia.

The differences in behaviour of cows and heifers should be considered in a system for monitoring parturition to avoid misconstruing the normal calving situation and to obviate unnecessary obstetric intervention in heifers. Behavioural elements which were more frequently detectable in cases of dystocia should draw attention to the possibility that there may be a problem in the first stage of labour.     

Biopsied and vitrified bovine embryos viability is improved by trans10, cis12 conjugated linoleic acid supplementation during in vitro embryo culture

Bovine embryos cultured in serum-containing media abnormally accumulate lipids in the cytoplasm. This is well known to contribute to their higher susceptibility to cryopreservation and biopsied embryos are even further susceptible. We aimed to improve in vitro produced (IVP) embryos resistance to micromanipulation and cryopreservation by supplementing serum-containing media with trans-10, cis-12 conjugated linoleic acid (t10, c12 CLA). The effect of t10, c12 CLA on lipid deposition and embryonic development was also tested. After in vitro maturation and fertilization (IVF day = D0), zygotes were cultured on granulosa cells + M199 + 10% serum + 100 μM GSH supplemented with 100 μM of t10, c12 CLA (CLA group, n = 1394) or without supplementation (control group, n = 1431). Samples of D7/D8 embryos were observed under Nomarsky microscopy for lipid droplets evaluation while others were biopsied and vitrified (group B-Control, n = 24; group B-CLA, n = 23). Non-biopsied embryos were also frozen (group NB-Control, n = 49; group NB-CLA, n = 45). Biopsied cells were used for embryo sex determination. Postwarming embryo survival and viability were determined at 0 and 24 h of culture, respectively. Supplementation of t10, c12 CLA did not influence cleavage, embryo sex ratio, D7/D8 embryo rate or morphological quality. CLA embryos had higher number of small lipid droplets (P ≤ 0.003) and a smaller (P < 0.001) fat embryo index being leaner (P = 0.008) than control embryos. Embryo postwarming survival was higher in B-CLA than in B-control group (95.0 ± 7.0% versus 62.5 ± 7.9%; P < 0.001). After 24 h of culture, the viability (expansion rate) of biopsied embryos and nonbiopsied embryos, cultured with t10, c12 CLA was higher than control embryos (B-CLA = 64.6 ± 4.4% and B-control = 27.5 ± 2.5%, P = 0.01; NB-CLA = 86.0 ± 3.5% and NB-Control = 68.6 ± 7.0%, P = 0.05). Results showed that supplying t10, c12 CLA to serum-containing media decreases embryo cytoplasmic lipid deposition during in vitro culture and significantly improves resistance of IVP embryos to micromanipulation and cryopreservation.