Predominant expression of growth hormone secretagogue receptor in the caudal lobe of chicken pituitary and parallel developmental increase in expression of pituitary GH and proventriculus ghrelin mRNA

To examine the involvement of ghrelin in growth hormone (GH) synthesis in the chicken pituitary, the regional distribution of GH secretagogue receptor (GHS‐R)/ghrelin receptor was investigated. Quantitative real‐time polymerase chain reaction (Q‐PCR) analysis revealed that the expression levels of GHS‐R and GH mRNA in the caudal lobe were about fourfold and sevenfold higher in the cephalic lobe of 7 day‐old chickens, respectively. Immunohistochemical analysis showed that GHS‐R immunoreactivity was more abundant in the caudal lobe than in the cephalic lobe, as was the case for GH immunoreactivity. By Q‐PCR, parallel increases were observed in the expression levels of ghrelin mRNA in the proventriculus and GH mRNA in the pituitary from embryonic day 17 to day 7 after hatching, whereas no significant change was found in the expression levels of GHS‐R mRNA in the pituitary during this period. These results suggest that proventriculus‐derived ghrelin may participate in pituitary GH synthesis by acting on its receptor during late embryonic development and the early post‐hatching period in chickens.     

Differential Effects of Continuous Exposure to the Investigational Metastin/Kisspeptin Analog TAK-683 on Pulsatile and Surge Mode Secretion of Luteinizing Hormone in Ovariectomized Goats

The aim of the present study was to determine if the estradiol-induced luteinizing hormone (LH) surge is influenced by the constant exposure to TAK-683, an investigational metastin/kisspeptin analog, that had been established to depress the pulsatile gonadotropin-releasing hormone (GnRH) and LH secretion in goats. Ovariectomized goats subcutaneously received TAK-683 (TAK-683 group, n=6) or vehicle (control group, n=6) constantly via subcutaneous implantation of an osmotic pump. Five days after the start of the treatment, estradiol was infused intravenously in both groups to evaluate the effects on the LH surge. Blood samples were collected at 6-min intervals for 4 h prior to the initiation of either the TAK-683 treatment or the estradiol infusion, to determine the profiles of pulsatile LH secretion. They were also collected at 2-h intervals from –4 h to 32 h after the start of estradiol infusion for analysis of LH surges. The frequency and mean concentrations of LH pulses in the TAK-683 group were remarkably suppressed 5 days after the start of TAK-683 treatment compared with those of the control group (P<0.05). On the other hand, a clear LH surge was observed in all animals of both groups. There were no significant differences in the LH concentrations for surge peak and the peak time of the LH surge between the TAK-683 and control groups. These findings suggest that the effects of continuous exposure to kisspeptin or its analog on the mechanism(s) that regulates the pulsatile and surge mode secretion of GnRH/LH are different in goats.     

The effect of high ambient temperature on Ca,P and Mg balance and bone turnover in high‐yielding dairy cows

We investigated the effect of heat stress on Ca, P and Mg balance and bone turnover in lactating cows. In a 2 × 2 crossover design, four multiparous lactating Holstein cows were kept in a chamber and subjected to a constant moderate (18°C) ambient temperature (MT) or high (28°C) ambient temperature (HT). The cows were fed total mixed ration (Ca, 0.7%; P, 0.4%; Mg, 0.2%) ad libitum. The milk yield under HT (35.4 kg/day) tended to be lower (P < 0.10) than that under MT (43.2 kg/day). The concentrations of milk P (P < 0.05) and Mg (P < 0.01) were significantly lower under HT than MT. The Ca, P and Mg intake (P < 0.10); Ca (P < 0.10), P, and Mg (P < 0.05) secretion into milk; and Ca (P < 0.05), P (P < 0.01), and Mg (P < 0.05) absorption in the intestine were lower under HT than MT. The plasma osteocalcin, a marker of bone turnover, was significantly lower (P < 0.05) under HT than MT. Heat stress did not affect plasma C‐telopeptide of collagen type I, a bone resorption marker, and plasma parathyroid hormone concentration.     

Maintenance of mouse trophoblast stem cells in KSR-based medium allows conventional 3D culture

Mouse trophoblast stem cells (TSCs) can differentiate into trophoblast cells, which constitute the placenta. Under conventional culture conditions, in a medium supplemented with 20% fetal bovine serum (FBS), fibroblast growth factor 4 (FGF4), and heparin and in the presence of mouse embryonic fibroblast cells (MEFs) as feeder cells, TSCs maintain their undifferentiated, proliferative status. MEFs can be replaced by a 70% MEF-conditioned medium (MEF-CM) or by TGF-ß/activin A. To find out if KnockOut^TM Serum Replacement (KSR) can replace FBS for TSC maintenance, we cultured mouse TSCs in KSR-based, FBS-free medium and investigated their proliferation capacity, stemness, and differentiation potential. The results indicated that fibronectin, vitronectin, or laminin coating was necessary for adhesion of TSCs under KSR-based conditions but not for their survival or proliferation. While the presence of FGF4, heparin, and activin A was not sufficient to support the proliferation of TSCs, the addition of a pan-retinoic acid receptor inverse agonist and a ROCK-inhibitor yielded a proliferation rate comparable to that obtained under the conventional FBS-based conditions. TSCs cultured under the KSR-based conditions had a gene expression and DNA methylation profile characteristic of TSCs and exhibited a differentiation potential. Moreover, under KSR-based conditions, we could obtain a suspension culture of TSCs using extracellular matrix (ECM) coating-free dishes. Thus, we have established here, KSR-based culture conditions for the maintenance of TSCs, which should be useful for future studies.     

Approachability and contact behavior of commercial dairy calves to humans

Calves (n = 106) on four dairy farms were observed for their approachability to humans. All calves experienced similar rearing conditions: Beginning individual pen, after birth until weaning at about 2 months, where they were housed individually and fed milk and a milk replacement; Late individual pen, after weaning until grouping at about 3.5 months, where they were housed individually and fed hay, silage and concentrate feed; Beginning group pen, after grouping until 5 months, where they were housed in groups of 2–5 animals and fed hay, silage and concentrate feed; later group pen, from 5 to 7 months. The number of calves that contacted an experimenter who stood in front of their pens for 10 min was recorded on 6 separate days over 3 months. Latency to touch and time spent in activities during touching such as sucking, licking, biting and rubbing were also measured. There were no significant differences in the latency to touch and the ratio of touch to non‐touch calves between the rearing conditions and the farms. The time spent touching was significantly affected by the interaction between the rearing condition and the farm (P < 0.01). In detail, the time spent sucking (P < 0.001) and licking (P < 0.01) was different between the rearing condition × farm variables. The proportion of calves that approached and touched the experimenter tended to be higher in the farms in which a stockperson worked longer inside and outside their pens (both ρ = 0.95, P = 0.051). These results were interpreted according to the perspectives of early positive reinforcement with food and the habituation process to humans existing nearby.