A carcinogenic western diet does not induce somatic mutations in various target tissues of transgenic C56BL/6 mice

Although the importance of diet in human cancer is clear, most dietary studies of carcinogenesis in laboratory rodents have involved the use of large doses of a carcinogen, which is not comparable to the human situation. The use of carcinogens has been necessary because laboratory rodents have extremely low spontaneous rates of colon cancer. Newmark et al. (2001) showed, however, that a radical dietary manipulation sufficed to induce high rates of colon cancer in C57BL/6 mice. Here we report an investigation into whether or not this dietary manipulation acts by altering somatic mutation rates. We used the transgenic lambda cII locus of F₁ pups (C57BL/6 × Big Blue^®) with the same C57BL/6 genetic background. The same diet (ND), high in fat, and low in calcium, vitamin D, folic acid, choline, and fibre, that was used by Newmark et al. (2001) was fed ad libitum to dams during pregnancy and lactation in order to examine its effect on mutagenesis in development and growth. There was no significant difference in mutant frequency in the small intestine (P = 0.82), or bone marrow (P = 0.95) of pups fed a ND versus the control diet. To investigate the effect of a ND during adulthood, 6-week-old F₁ pups were fed a ND ad libitum for 6, 12 and 19 weeks. There was no significant difference in mutant frequency in the small intestine (P = 0.66) or colon (P = 0.49) at the cII locus with no significant difference in body weight. These results indicate that Western diet-induced carcinogenesis is not mediated by alterations in mutation rate and thus may act at the promotion rather than at the initiation stage of carcinogenesis.     

Growth response of major USA cowpea cultivars: II. Effect of salinity on leaf gas exchange

In an effort to elucidate the physiological processes involved in cowpea differential growth response of four major USA cowpea cultivars (CB5, CB27, 8517 and 7964) to increasing salinity, we investigated the effect of salinity on leaf gas exchange of net photosynthetic rate per unit leaf mass (Pnm) and per unit leaf area (Pna), and stomatal conductance (gs) of the four cowpea cultivars. The experiment was set up as a standard split-plot design in which cowpea plants were grown in greenhouse sand tanks irrigated with nutrient solutions. Seven salinities ranging from 2.6 to 20.5 dS m⁻¹ were constructed based on Colorado River water salt composition with NaCl, CaCl₂ and MgSO₄ as the salinization salts. Light-saturated Pnm, Pna and gs of fully expanded trifoliage were examined at the vegetative growth and flowering stages, and the data were analyzed using a split-plot analysis of variance (ANOVA) model. We found a highly significant (P ≤ 0.0001) reduction of Pnm, Pna and gs due to salinity. The responses of Pnm, Pna and gs to salinity could be further described by a general model of log(y) = a₁ + a₂x + a₃x², where y represents either Pnm, Pna, or gs; a₁, a₂ and a₃, empirical constants; x, salinity. We found that Pnm was more sensitive to salinity than Pna. Additionally, we found that increasing stomatal closure with increasing salinity might limit Pnm or Pna. While we did not find any significant difference (P > 0.05) of Pnm and Pna among the four cultivars, we did find a significant difference (P ≤ 0.05) in gs. No significant salt × cultivar interaction effect (P > 0.05) was found with Pnm, Pna and gs indicating that the four cowpea cultivars have the same response pattern of their leaf gas exchange to salinity.     

CVT-4325: a potent fatty acid oxidation inhibitor with favorable oral bioavailability

New inhibitors of palmitoyl-CoA oxidation are based on the introduction of nitrogen heterocycles in the ‘Western Portion’ of the molecule. SAR studies led to the discovery of CVT-4325 (shown), a potent FOXi (IC₅₀ = 380 nM rat mitochondria) with favorable PK properties (F = 93%, t_1/2 = 13.6 h, dog).     

Group differences in captive gorillas’ reaction to large crowds

The impact of visitor crowd size on captive animal behaviour can be difficult to interpret in many institutions due to the lack of variation in crowd size over short periods of time. In attempts to compare greater variation in crowd size, animal behaviour is often compared over conditions that create additional confounds, such as day of week or season. Fluctuations in attendance over the holiday season at Disney's Animal Kingdom^® Theme Park provided an opportunity to examine the impact of variation in crowd size on gorilla behaviour without the confounds found in many other studies. Ten western lowland gorillas (Gorilla g. gorilla) in a bachelor group (n = 4) and a family group (n = 6) were observed over a period of 8 weeks in late 2005. Observations were classified into Large Crowd and Small Crowd days and a repeated-measures ANOVA procedure was conducted to determine the effects of crowd size and social group on gorilla behaviour patterns. Few overall differences were found in behaviour during the Large Crowd and Small Crowd conditions. However, similar to previous research, gorillas were less visible during the Large Crowd condition (F_1,8 = 14.15, P = 0.01). There was also an interaction of crowd size and group (F_1,8 = 7.58, P = 0.01), indicating the bachelor group of gorillas was more aggressive during the Large Crowd condition, whereas the family group showed no increase in aggression with large crowds. These results indicate the importance of providing complex enclosures with visual barriers to allow animals to move away from large crowds if they choose. Future research should focus on the individual characteristics of animals and enclosures that may contribute to behavioural differences in visitor–animal interaction research, as well as the proximate cues associated with behaviour change when exposed to large crowds.     

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.     

A thermodynamic and crystallographic study of complexes of the highly preorganized ligand 8-hydroxyquinoline-2-carboxylic acid

The metal ion complexing properties of the ligand HQC (8-hydroxyquinoline-2-carboxylic acid) are reported. The structures of [Zn(HQCH)₂] · 3H₂O (1) and [Cd(HQCH)₂] · 3H₂O (2) were determined (HQCH = HQC with phenol protonated). Both 1 and 2 are triclinic, space group , with Z = 2. For 1 a = 7.152(3), b = 9.227(4), c = 15.629(7) Å,  = 103.978(7)°, β = 94.896(7)°, γ = 108.033(8)°, R = 0.0499. For 2 a = 7.0897(5), b = 9.1674(7), c = 16.0672(11) Å,  = 105.0240(10)°, β = 93.9910(10)°, γ = 107.1270(10)°, R = 0.0330. In 1 the Zn has a distorted octahedral coordination geometry, with Zn–N of 2.00 and 2.15 Å, and Zn–O to the protonated phenolic oxygens of 2.431 and 2.220 Å. The structure of 2 is similar, with Cd–N bonds of 2.220 and 2.228 Å, with Cd–O bonds to the protonated phenolate oxygens of 2.334 and 2.463 Å. The structures of 1 and 2, and isomorphous Ni(II) and Co(II) HQC complexes reported in the literature, show very interesting short (<2.5 Å) O–O distances in H-bonds involving the protons on the coordinated phenolates and lattice water molecules. These are discussed in relation to the possible role of short low-energy H-bonds in alcohol dehydrogenase in mediating the transfer of the hydroxyl proton of the alcohol to an adjacent serine oxygen.

The formation constants for HQC are determined by UV–Visible spectroscopy at 25 °C in 0.1 M NaClO₄ with Mg(II), Ca(II), Sr(II), Ba(II), La(III), Gd(III), Zn(II), Cd(II), Ni(II), Cu(II), and Pb(II). These show greatest stabilization with metal ions with an ionic radius above 1.0 Å. This is as would be expected from the fact that HQC forms two five-membered chelate rings on complex-formation, which favors larger metal ions. The ligand design concept of using rigid aromatic backbones in ligands to achieve high levels of preorganization, and hence the high log K values (for a tridentate ligand) and strong metal ion selectivities observed for HQC, is discussed.     

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.     

Spin-crossover iron(II) complexes [Fe(Medpq)(py)2(NCS)2] and [Fe(Medpq)(py)2(NCSe)2]: syntheses, characterization and magnetic properties

Two new spin-crossover complexes, [Fe(Medpq)(py)₂(NCS)₂] · py · 0.5H₂O (1) and [Fe(Medpq)(py)₂(NCSe)₂] · py (2) (Medpq = 2-methyldipyrido[3,2-f:2′,3′-h]-quinoxaline, py = pyridine), have been synthesized. The crystal structures were determined at both room temperature (298 K) and low temperature (110 K). Complexes 1 and 2 crystallize in the orthorhombic space group Pbca and monoclinic space group P2₁/n, respectively. In both complexes, the distorted [FeN₆] octahedron is formed by six nitrogen atoms from Medpq, the trans pyridine molecules and the cis NCX⁻ groups. The thermal spin transition is accompanied by the shortening of the mean Fe–N distances by 0.194 Å for 2. The mononuclear [Fe(Medpq)(py)₂(NCS)₂] and [Fe(Medpq)(py)₂(NCSe)₂] neutral species interact each other via π-stacking, resulting in a one-dimensional extended structure for both 1 and 2. There exist C–HX (X = S, Se) hydrogen bonds for both complexes. Variable-temperature magnetic susceptibility measurements and Mössbauer spectroscopy reveal the occurrence of a gradual spin transition. The transitions are centered at T_1/2 = 120 K for 1 and T_1/2 = 180 K for 2, respectively.     

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.     

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.     

Treatment with high-dose estrogen (diethylstilbestrol) significantly decreases plasma estrogen and androgen levels but does not influence in vivo aromatization in postmenopausal breast cancer patients

While growth factors and hormones are known to influence aromatase expression in experimental systems, little is know about potential factors influencing peripheral aromatization in postmenopausal women. The fact that peripheral aromatase activity is higher in old compared to young women and the finding of relatively high tissue estradiol (E₂) concentrations after the menopause suggests peripheral aromatization could be influenced by estrogen concentration. To test this hypothesis, we determined plasma hormone levels (n = 9) and in vivo aromatization (n = 3) in postmenopausal women suffering from advanced breast cancer before and during treatment (4 weeks) with diethylstilbestrol (DES) 5 mg three times daily. Plasma levels of cortisol (C), corticosteroid-binding globulin (CBG), and sex hormone binding globulin (SHBG) were significantly increased in all patients (P < 0.05 for all). While we found no change in total body aromatization and plasma estrone (E₁) levels, estradiol (E₂) and estrone sulfate (E₁S) were suppressed by a mean of 48.8 and 68.2%, respectively (P = 0.043 and 0.008). Surprisingly, plasma levels of androstenedione (A), testosterone (T), dehydroepiandrosterone (DHEA) and its sulfate (DHEAS) were also suppressed by a mean in the range of 32.1 to 52.6% (P < 0.05 for all androgens). In contrast, no change in plasma progesterone or 17-hydroxyprogesterone was found. Thus, one possible explanation to our findings could be that DES administered in high doses reduces 17,20-lyase activity in the adrenal gland.     

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.     

Social reinstatement responses of meat-type chickens to familiar and unfamiliar conspecifics after exposure to an acute stressor

Runway tests are considered indicative of underlying sociality in birds and their ability to make social discriminations. We evaluated whether experience of a prior stressor alters the subsequent affiliation responses of 9 or 10-day-old chicks simultaneously exposed to familiar (cagemates) and unfamiliar conspecifics placed in goal boxes at opposite ends of a runway. Birds were housed in groups of eight in home cages. Half of the birds in each home cage were used as either familiar or unfamiliar social stimuli in the goal boxes. The other half of the birds were randomly assigned either to a control (CON; n = 51) group that remained undisturbed until testing or to a stress-treatment (STR; n = 52) group that was exposed to a 5-min restraint stressor, returned to its home cage and then tested 1 h later. Birds were individually tested in the runway for 5 min and the behaviours video-recorded. During revision of tapes, the projected floor image of the runway was divided into squares and zones. The stressor decreased (P < 0.01) the time spent in close proximity (close zone; CZ) to conspecifics regardless of the familiarity of the stimulus birds. Regardless of treatment, test chicks showed shorter latencies to enter (P < 0.05) and spent longer time (P < 0.02) in the familiar than in the unfamiliar CZ suggesting that young chicks can discriminate between familiar and unfamiliar conspecifics encountered in novel surroundings. While in close proximity to familiar conspecifics, STR birds showed a reduced (P < 0.05) number of squares entered compared to CONs. This reduced locomotor activity was not accompanied by an increased activity in other zones of the runway. At the end of the trial, both CON and STR birds showed a reduced (P < 0.05) locomotor activity in the unfamiliar CZ and an increased (P < 0.05) activity in the central zone of the runway. Interestingly, no differences were detected between CON and STR birds in the total number of squares entered during the trial. These results suggest that prior stressor exposure did not affect the overall amount of locomotion but altered the spatial distribution of it. Collectively, our findings suggest that exposure to an acute stressor event subsequently affects chicks’ affiliation responses in runway tests. The way a bird will react depends on the identity (familiar or unfamiliar) of the conspecifics in its close environment.     

Pleistocene population expansions of Antarctic seals

We sequenced a portion ( c . 475 bp) of the mitochondrial control region of three species of Antarctic phocid carnivores (Weddell seal, Leptonychotes weddellii , N  = 181; crabeater seal, Lobodon carcinophaga , N  = 143; and Ross seal, Ommatophoca rossii , N  = 41) that live seasonally or permanently in the fast ice and seasonal pack ice of the western Amundsen and Ross seas of western Antarctica. We resolved 251 haplotypes with a haplotype diversity of 0.98 to 0.99. Bayesian estimates of Θ from the program LAMARC ranged from 0.075 for Weddell seals to 0.576 for crabeater seals. We used the values of theta to estimate female effective population sizes ( N_EF ), which were 40 700 to 63 000 for Weddell seals, 44 400 to 97 800 for Ross seals, and 358 500 to 531 900 for crabeater seals. We used mismatch distributions to test for historical population size expansions. Weddell seals and crabeater seals had significant, unimodal mean pairwise difference distributions ( P  = 0.56 and 0.36, respectively), suggesting that their populations expanded suddenly around 731 000 years ago (Weddell seals) and around 1.6 million years ago (crabeater seals). Both of these expansions occurred during times of intensified glaciations and may have been fostered by expanding pack ice habitat.     

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.     

Enhanced efficiency of a capillary-based biosensor over an optical fiber biosensor for detecting calpastatin

A capillary-based optical biosensor has been developed to detect calpastatin, an indicator of meat tenderness. Longissimus muscle samples (n = 11) were extracted from beef carcasses at 0 and 48 h post-mortem. These samples were assayed for calpastatin by traditional laboratory methods and with a newly developed capillary tube biosensor as well as for Warner–Bratzler shear force (WBSF) and crude protein and the responses were compared. Additionally, the response from the capillary-based biosensor was compared to a previously developed optical fiber biosensor. When the 0 and 48 h sampling periods were combined, the capillary tube biosensor was moderately accurate in predicting calpastatin activity (R² = 0.6058). There was less variation in the 0 h capillary tube biosensor compared to the 0 h pre-column (P = 0.006) and post-column optical fiber biosensors (P = 0.047), therefore the capillary tube biosensor is a more precise system of measurement. This research further advances the development of a calpastatin biosensor and makes online assessment one step closer to reality.     

Effects of Tasco (a seaweed extract) and heat stress on N metabolism and meat fatty acids in wether lambs fed hays containing endophyte-infected fescue

Wether lambs (n = 27, average BW = 40 kg) were used to test response to forage treated with Tasco-Forage (an extract of the brown kelp Ascophyllum nodosum) prior to conserving, or to direct feeding of the extract (Tasco-EX). Hays made from endophyte (Neoyphodium coenophialum)-infested tall fescue (Festuca arundinacea)-based pasture received 0 or 3 kg of Tasco/ha prior to harvest. Lambs, blocked by weight, were randomly allotted to three diets: (1) control hay, (2) treated hay, and (3) #1 + Tasco-EX fed at 1% of the diet. Hays were low in CP (<7%) so all lambs were fed soybean meal (12% of the diet) in addition to trace mineralized salt. Diets were fed at 1.5% BW to prevent refusals. Total collections (7 d) were made during periods without or with applied heat stress. After each period, rumen contents were obtained to determine pH, NH₃ and VFA. Lambs were sacrificed post-trial. A subset was used to evaluate sensory traits and muscle fatty acids. Lambs were in negative N balance during the study and Tasco treatments did not affect N metabolism. Fecal N tended (P < 0.10) to increase with short duration heat stress causing a concomitant decrease (P < 0.05) in apparent N digestibility (58.6 versus 56.1%; S.E. = 0.7). Urinary N loss decreased (P < 0.001) with heat stress (8.0 versus 5.9 g/d; S.E. = 0.2), resulting in increased (P < 0.001) N retention (−2.1 versus −0.3 g/d; S.E. = 0.2). Apparent OM digestibility was not affected by heat stress but was greater (P < 0.05) for lambs fed Tasco-EX treatment than those fed treated hay. Treatment diets decreased (P < 0.05) ruminal butyrate. Heat stress increased (P < 0.05) acetate and total VFA and decreased (P < 0.01) ruminal pH. A tendency (P < 0.11) of increased 14:1ω5, decreased (P < 0.05) 18:0 and total saturated fatty acids in muscle was observed with Tasco diets. Meat sensory characteristics were not affected by treatment. Tasco may alter some aspects of rumen or lipid metabolism but has no effect on N metabolism or meat sensory characteristics of sheep fed restricted, low-quality diets.     

Structure-activity studies on different modifications of nociceptin/orphanin FQ: identification of highly potent agonists and antagonists of its receptor

Nociceptin/orphanin FQ (N/OFQ) and its receptor system modulate a variety of biological functions and further understandings of physiological and pathological roles of this system require new potent agonists and antagonists of its receptor. Two series of N/OFQ related analogues were synthesized to investigate the relationship of different modifications. We combined modifications including: (a) Phe⁴→(pF)Phe⁴; (b) Ala⁷, Ala¹¹→Aib⁷, Aib¹¹; (c) Leu¹⁴, Ala¹⁵→Arg¹⁴, Lys¹⁵. Compared with the first series, N-terminus of the second series was changed from Phe¹ to Nphe¹. All the analogues were amidated at C-terminus. These compounds were tested in binding studies on rat brain membranes and mouse vas deferens assay. Results indicated that the compounds of the first series showed higher affinity and potency than N/OFQ (pK_i = 9.33; pEC₅₀ = 7.50). In particular, [(pF)Phe⁴, Aib⁷, Aib¹¹, Arg¹⁴, Lys¹⁵] N/OFQ-NH₂ was found to be a highly potent agonist with pK_i = 10.78 in binding studies and pEC₅₀ = 9.37 in mouse vas deferens assay. The second series all competitively antagonized the effects of N/OFQ in mouse vas deferens assay. [Nphe¹, (pF)Phe⁴, Aib⁷, Aib¹¹, Arg¹⁴, Lys¹⁵] N/OFQ-NH₂ was the best antagonist with pA₂ = 8.39 and showed high binding affinity with pK_i = 9.99. Thus modifications which increase the potency of agonist have synergistic effect on biological activity and a replacement of N-terminus leads to shift of analogues from agonist to antagonist.     

Structure and thermal decomposition of methylamine borane

The crystal structure of methylamine borane has been determined and contains parallel chains of dihydrogen-bonded CH₃NH₂BH₃ molecules. Thermal decomposition takes place from the melt (ΔH_fusion = 8.5 kJ mol⁻¹) and begins with the formation of an ionic borohydride. Hydrogen is liberated in two stages, at ca. 100 and 190 °C, with the observed rates during the first stage (ΔH = −25 kJ mol⁻¹, E_a = 115 kJ mol⁻¹) strongly dependent on temperature and time. cis- and trans-N-trimethylcyclotriborazane are formed during the first stage and subsequently cross-link to yield a non-volatile solid. Before this cross-linking, the system exhibited a high degree of volatility, with weight losses in excess of 80% observed in TG experiments using flowing gas.     

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.