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The current study was performed to clarify the effects of GHRH treatment on milk production and plasma hormones and metabolites in lactating Japanese Black cows (a beef breed) under negative energy balance (EB). Ten multiparous lactating beef cows were offered a normal-energy diet daily (110% of ME requirements for maintenance and lactation) until 5 d in milk (DIM) to standardize the cows before dietary treatment. From 6 DIM to the final days (63 DIM) of the experiment, the cows were allotted to experimental dietary treatments: 5 cows were offered a diet formulated for 130% [high-energy diet (HED)] and the remaining 5 cows were offered a diet formulated for 80% [low-energy diet (LED)] of ME requirements for maintenance and lactation. In addition, all cows received daily subcutaneous injections of 3 mg of bovine GHRH from 36 to 56 DIM (GHRH treatment period). Differences in BW of HED- and LED-fed cows at 63 DIM were +28.4 and -7.2 kg compared with BW at 6 DIM, and HED- and LED-fed cows were under positive EB (+23.7 MJ/d) and negative EB (-11.6 MJ/d) throughout the experiment period. Treatment with GHRH increased (P < 0.01) the average daily milk yield to 6.2 kg in HED-fed cows compared with a milk yield of 5.3 kg for 7 d before the GHRH treatment period (pretreatment period); LED-fed cows had no increase in milk production from GHRH treatment. Plasma GH, IGF-1, insulin, and glucose concentrations increased (P < 0.05) after GHRH treatment in both HED- and LED-fed cows; GHRH treatment also induced an increase (P < 0.05) in the net area under the curve of plasma insulin after glucose challenge in both HED- and LED-fed cows. Plasma urea N concentrations were decreased (P < 0.05) by GHRH treatment in HED-fed cows, but not in LED-fed cows. Plasma NEFA concentration was unaffected by GHRH treatment in both HED- and LED-fed cows. We conclude that GHRH treatment of lactating Japanese Black cows stimulates endogenous GH and subsequent IGF-1 secretion and might induce an increase in insulin resistance, irrespective of EB; however, compared with lactating dairy cows, both galactopoietic and lipolytic effects of GHRH might be insufficiently exerted under negative EB in lactating beef cows. Key words: beef breed, energy balance, growth hormone, growth hormone-releasing hormone, insulin, lactation

Urea is an important reutilizable nitrogen source for the ruminant and is mainly synthesized through the urea cycle in the liver. The cycle is undertaken by 5 enzymes: carbamoyl phosphate synthetase (CPS), ornithine transcarbamoylase (OTC), arginino-succinate synthetase (AS), argininosuccinate lyase (AL), and arginase. The purpose of this study was to investigate changes in the activity of the enzymes and mRNA expression, given that previous observations have indicated an increase in plasma urea concentrations with age in Holstein calves. First, plasma concentrations of metabolites and hormones were determined in calves at 1, 3, 8, 13, and 19 wk of age (n = 4, weaned at 6 wk of age). The plasma concentration of urea drastically increased after weaning (P < 0.001). The plasma concentration of glucose was lowest at 8 wk. The plasma concentration of IGF-I gradually increased with age, although those of NEFA, glucagon, and cortisol decreased (P < 0.001). Concentrations of triglyceride, [alpha]-amino nitrogen, growth hormone, and insulin did not change significantly with age of the calf. Next, using the liver tissues taken from calves at 2, 13, and 19 wk of age (n = 4 to 6 at each time point, weaned at 6 wk of age), we measured the activity and mRNA expression of the enzymes by biochemical methods and quantitative reverse transcription-PCR, respectively. The activities of CPS (P < 0.001), OTC (P = 0.001), and AS -(P = 0.015) increased with age, whereas AL (P = 0.003) decreased. Although mRNA expression was decreased with age for AL (P = 0.002) and arginase (P = 0.007), no significant change was observed for CPS, OTC, or AS mRNA expression. We conclude that the increased urea production in the liver may be explained not only by an increase in the activities of the urea cycle enzymes, but also by increased ammonia production by rumen fermentation and gluconeogenesis from amino acids around weaning time. Key words: calf, development, liver, mRNA, urea cycle enzyme, weaning

The current study was conducted to clarify the characteristics of glucose and urea kinetics in male Japanese Black calves, using a glucose and urea dilution method with stable isotopes, at preweaning (3 wk of age) and postweaning (13 and 26 wk of age) stages, in comparison with the kinetics of glucose and urea in male Holstein calves. Six Japanese Black and 6 Holstein calves were arranged in a 2 (breed) x 3 (stage) factorial block design. These 12 suckling calves were fed only whole milk, offered concentrate and orchardgrass hay after 3 wk of age, and weaned at 6 wk of age. Under steady-state conditions, glucose challenges (1.0 mg/kg of BW; [U-[sup.13]C]D-glucose) and urea challenges (both 2.2 mg/kg of BW; [[sup.13]C]urea and [[sup.15][N.sub.2]]urea) were performed at the 3 stages examined. There were no significant differences in plasma glucose concentrations between Japanese Black and Holstein calves at any stage, but the glucose concentrations at 3 wk of age were greater (P < 0.05) than those at 13 wk of age in both breeds. The glucose pool size in Japanese Black calves was smaller (P < 0.05) than that in Holstein calves at all stages. Within each breed, there were no significant differences between the glucose pool sizes at 3 and 13 wk of age, but the glucose pool size was larger (P < 0.05) at 26 wk of age. Moreover, Japanese Black calves had greater glucose irreversible loss rates (P < 0.01) than Holstein calves at 3 wk of age, and the glucose irreversible loss rates were less (P < 0.05) on and after 13 wk of age in both breeds. Japanese Black calves had greater plasma urea N concentrations (P < 0.05) than Holstein calves at all stages, and in both breeds, the urea N concentrations increased (P < 0.05) with age. The urea pool size was smaller (P < 0.01) in Japanese Black calves than in Holstein calves at all stages, and in both breeds, the urea pool size increased (P < 0.05) with age. In comparison with Holstein calves, Japanese Black calves had greater urea irreversible loss rates (P < 0.05) on and after 13 wk of age and greater urea recycling rates (P < 0.05) at 26 wk of age. In addition, in both breeds, urea irreversible loss rates and urea recycling rates increased (P < 0.05) with age. We conclude that Japanese Black calves have partially different glucose and urea kinetics from Holstein calves and that the kinetics of these metabolites in both Japanese Black and Holstein calves are strongly influenced by weaning. Key words: breed, glucose kinetics, irreversible loss rate, Japanese Black calf, recycling rate, urea kinetics

Endotoxin induces marked changes in lipid metabolism via its effects on cytokines. To evaluate the role of tumor necrosis factor-alpha (TNF) in mediating changes of lipid metabolism in ruminants, we performed a crossover saline-controlled study in Holstein heifers (n = 8; 394.0 kg average BW), investigating the metabolic effects of a single intravenous administration of recombinant bovine TNF (rbTNF, 5.0 [micro]g/kg). Blood samples were taken from a jugular vein at 0 (1100, just before injection), 0.5, 6, 12, and 24 h after each treatment. Dry matter intake in the heifers was not affected by single administration of the rbTNF. The rbTNF produced early as well as later hypertriglyceridemia (P < 0.05) in dairy heifers. The rbTNF also induced an early and sustained rise (P < 0.05) in the plasma NEFA concentration. Plasma retinol concentration was decreased (P < 0.05) at 24 h after rbTNF injection, whereas the [alpha]-tocopherol concentration was not significantly affected by rbTNF treatment. At 0.5 and 24 h, there was an increase (P < 0.05) in the plasma concentration of the very-low-density lipoprotein (VLDL) fraction in rbTNF-treated heifers. Between 6 and 24 h after rbTNF treatment, concentration of the low-density lipoprotein fraction declined (P < 0.05) but the high-density lipoprotein fraction was not altered in the rbTNF-treated heifers. These results indicate that TNF produces a hypertriglyceridemic response associated with an increase of the VLDL fraction and a disturbance of retinol metabolism in dairy heifers. Key Words: Dairy Cattle, Lipoproteins, Retinol, Tumor Necrosis Factor, Vitamin E Acetate

Four experiments were conducted to clarify the effect of intravenous (i.v.) administration of recombinant bovine tumor necrosis factor [Alpha] (rbTNF) on selected metabolites and on hormone secretion in Holstein heifers (n = 6; 347.0 kg average BW). In Exp. 1, rbTNF was injected at three dosage levels in a Latin square; 0 (CONT), 2.5 (TNF2.5), or 5.0 (TNF5) [micro]g/kg BW. Plasma glucose and triglyceride concentrations were at first elevated (P [is less than] .05) by rbTNF treatment and then were decreased (P [is less than] .05) by TNF2.5 and TNF5. Plasma NEFA concentrations were increased (P [is less than] .05) in rbTNF-treated groups. The injection of rbTNF resulted in an increase in plasma insulin levels (P [is less than] .05 with TNF5) during the period between 2 and 24 h, except for the period between 6 and 8 h, after the treatment. In Exp. 2, 3, and 4, each heifer received i.v. injections of glucose (.625 mM/kg BW) + rbTNF(5 [micro]g/kg) or glucose + saline (10 mL) (Exp. 2), insulin (0.2 U/kg) + rbTNF or insulin + saline (Exp. 3), and GHRH (0.25 [micro]g/kg) + rbTNF or GHRH + saline (Exp. 4) at 1-wk intervals. In Exp. 2, rbTNF inhibited (P [is less than] .05) glucose-stimulated insulin secretion during the initial phase. Thereafter, plasma insulin was higher (P [is less than] .01) with the glucose + rbTNF treatment than with the glucose + saline treatment. Treatment with rbTNF inhibited the insulin-stimulated glucose utilization (Exp. 3) and GHRH-stimulated GH secretion (Exp. 4) during the initial phase. These results suggest that rbTNF directly and(or) indirectly affects the intermediary metabolism and hormone secretion in Holstein heifers. Key Words: Blood Chemistry, Bovidae, Heifers, Insulin, Tumor Necrosis Factor, Somatotropin