Absorption and metabolism of benzoic acid in growing pigs.

Dietary benzoic acid (BA) supplementation causes a pronounced reduction in urinary pH but only small changes in blood pH. The present study aimed to investigate the portal absorption profile, hepatic metabolism of BA, and renal excretion of hippuric acid (HA) underlying the relatively small impact of BA on systemic acid-base status. Eight growing pigs (BW = 63 +/- 1 kg at sampling) fitted with permanent indwelling catheters in the abdominal aorta, hepatic portal vein, hepatic vein, and mesenteric vein were allocated to 4 sampling blocks and randomly assigned to control (CON; nonsupplemented diet) or BA supplementation (B; control diet + 1% BA top-dressed). Feed intake was restricted to 3.6% of BW and the ration divided into 3 equally sized meals offered at 8-h intervals. Blood pH (7.465 and 7.486 +/- 0.004) and urinary pH (4.99 and 7.01 +/- 0.09) were less (P = 0.03 and P < 0.01) in B compared with CON. The arterial concentration, net portal flux, and net hepatic uptake of BA increased (P < 0.01) in B compared with CON. The net portal flux of BA increased (P < 0.01) after feeding with B, but remained positive (P < 0.01) at all sampling times (n = 8). Recovery of dietary BA as increased net portal flux and hepatic uptake of BA was 87 +/- 5% and 89 +/- 15%, respectively. The recovery of dietary BA as urinary excretion of BA and HA was 0.08 +/- 0.02% and 85 +/- 7%, respectively. It is concluded that the small impact of BA supplementation on systemic acid-base status was caused by a protracted BA absorption and efficient hepatic extraction and glycine conjugation in combination with efficient renal clearance of HA. Together, these physiological mechanisms prevented major BA and HA accumulation in body fluids.

Systemic effects of angiotensin III in conscious dogs during acute double blockade of the renin-angiotensin-aldosterone-system.

AIMS: The study was designed to determine (i) whether the effects of angiotensin III (AngIII) are similar to those of angiotensin II (AngII) at identical plasma concentrations and (ii) whether AngIII operates solely through AT1- receptors. METHODS: Angiotensin II (3 pmol kg(-1) min(-1)-3.1 ng kg(-1) min(-1)) or AngIII (15 pmol kg(-1) min(-1)-14 ng kg(-1) min(-1)) was infused i.v. during acute inhibition of angiotensin converting enzyme (enalaprilate; 2 mg kg(-1)) and of aldosterone (canrenoate; 6 mg kg(-1) plus 1 mg kg(-1) h(-1)). Arterial plasma concentrations of angiotensins were determined by radioimmunoassay using a cross-reacting antibody to AngII. During ongoing peptide infusion, candesartan (2 mg kg(-1)) was administered to block the AT1-receptors. RESULTS: Angiotensin immunoactivity in plasma increased to 60 +/- 10 pg mL(-1) during infusion of AngII or infusion of AngIII. AngII significantly increased mean arterial blood pressure (+14 +/- 4 mmHg) and plasma aldosterone by 79% (+149 +/- 17 pg mL(-1)) and reduced plasma renin activity and sodium excretion (-41 +/- 16 mIU L(-1) and -46 +/- 6 micromol min(-1) respectively). AngIII mimicked these effects and the magnitude of AngIII responses was statistically indistinguishable from those of AngII. All measured effects of both peptides were blocked by candesartan. CONCLUSION: At the present arterial plasma concentrations, AngIII is equipotent to AngII with regard to effects on blood pressure, aldosterone secretion and renal functions, and these AngIII effects are mediated through AT1- receptors. The metabolic clearance rate of AngIII is five times that of AngII.

Effects of sodium intake on plasma potassium and renin angiotensin aldosterone system in conscious dogs.

AIMS: The operating range of the renin-angiotensin-aldosterone system is ill-defined. This study quantifies renin-angiotensin-aldosterone system activity as a function of sodium intake. METHODS: Renin-angiotensin-aldosterone system variables were measured daily after a sudden reduction in sodium intake (3.0-0.5 mmol kg(-1) day(-1)) or at steady states generated by eight levels of sodium intake (0.5-8.0 mmol kg(-1) day(-1)). Potassium intake was 2.79 +/- 0.03 mmol kg(-1) day(-1). Arterial blood pressure was measured invasively. Hormone concentrations were determined by radioimmunoassays. Glomerular filtration rate and plasma volume were determined by standard methods. RESULTS: Sudden sodium intake reduction doubled plasma renin activity and angiotensin II, and tripled aldosterone on day 1 with only small non-significant additional changes on the following days. Different levels of sodium intake did not affect arterial blood pressure, heart rate, and plasma concentrations of sodium, angiotensinogen, atrial natriuretic peptide, vasopressin, glomerular filtration rate and diuresis. With increasing sodium intake, plasma volume increased by 0.47 +/- 0.04 mL (kg body mass)(-1) (unit increase in Na intake)(-1) (P < 0.01), and plasma potassium decreased with the slope -0.038 mm [(mmol Na+ intake) (kg body mass)(-1) day(-1)](-1) (P = 0.001) while plasma renin-activity, angiotensin II, and aldosterone decreased systematically as expected. CONCLUSIONS: A step reduction in sodium intake alters renin-angiotensin-aldosterone system activity on day 1 with little further change the subsequent 4 days. Week-long increases in sodium intake decreases renin-angiotensin-aldosterone system activity, increases plasma volume, and decreases plasma potassium. Isolated decreases in sodium intake increase aldosterone secretion via volume-mediated action on the renin-angiotensin system and via increases in plasma potassium.

Volume natriuresis vs. pressure natriuresis.

Body fluid regulation depends on regulation of renal excretion. This includes a fast vasopressin-mediated water-retaining mechanism, and slower, complex sodium-retaining systems dominated by the renin-angiotensin aldosterone cascade. The sensory mechanisms of sodium control are not identified; effectors may include renal arterial pressure, renal reflexes, extrarenal hormones and other regulatory factors. Since the pioneering work of Guyton more than three decades ago, pressure natriuresis has been in focus. Dissociations between sodium excretion and blood pressure are explained as conditions where regulatory performance exceeds the precision of the measurements. It is inherent to the concept, however, that sudden transition from low to high sodium intake elicits an arterial pressure increase, which is reversed by the pressure natriuresis mechanism. However, such transitions elicit parallel changes in extracellular fluid volume thereby activating volume receptors. Recently we studied the orchestration of sodium homeostasis by chronic and acute sodium loading in normal humans and trained dogs. Small increases in arterial blood pressure are easily generated by acute sodium loading, and dogs appear more sensitive than humans. However, with suitable loading procedures it is possible - also acutely - to augment renal sodium excretion by at least one order of magnitude without any change in arterial pressure whatsoever. Although pressure natriuresis is a powerful mechanism capable of overriding any other controller, it seems possible that it is not operative under normal conditions. Consequently, it is suggested that physiological control of sodium excretion is neurohumoral based on extracellular volume with neural control of renin system activity as an essential component.

Preservation of active incretin hormones by inhibition of dipeptidyl peptidase IV suppresses meal-induced incretin secretion in dogs.

The incretin hormones glucagon-like peptide-1 (GLP-1) and glucose-dependent insulinotropic polypeptide (GIP) are degraded by dipeptidyl peptidase IV (DPP IV), thereby losing insulinotropic activity. DPP IV inhibition reduces exogenous GLP-1 degradation, but the extent of endogenous incretin protection has not been fully assessed, largely because suitable assays which distinguish between intact and degraded peptides have been unavailable. Using newly developed assays for intact GLP-1 and GIP, the effect of DPP IV inhibition on incretin hormone metabolism was examined. Conscious dogs were given NVP-DPP728, a specific DPP IV inhibitor, at a dose that inhibited over 90% of plasma DPP IV for the first 90 min following treatment. Total and intact incretin concentrations increased (P<0.0001) following a mixed meal, but on control days (vehicle infusion), intact peptide concentrations were lower (P<0.01) than total peptide concentrations (22.6 +/- 1.2% intact GIP; 10.1 +/- 0.4% intact GLP-1). Following inhibitor treatment, the proportion of intact peptide increased (92.5 +/- 4.3% intact GIP, P<0.0001; 99.0 +/- 22.6% intact GLP-1, P<0.02). Active (intact) incretins increased after NVP-DPP728 (from 4797 +/- 364 to 10 649 +/- 106 pM x min for GIP, P<0.03; from 646 +/- 134 to 2822 +/- 528 pM x m in for GLP-1, P<0.05). In contrast, total incretins fell (from 21 632 +/- 654 to 12 084 +/- 1723 pM x min for GIP, P<0.002; from 5145 +/- 677 to 3060 +/- 601 pM x min for GLP-1, P<0.05). Plasma glucose, insulin and glucagon concentrations were unaltered by the inhibitor. We have concluded that DPP IV inhibition with NVP-DPP728 prevents N-terminal degradation of endogenous incretins in vivo, resulting in increased plasma concentrations of intact, biologically active GIP and GLP-1. Total incretin secretion was reduced by DPP IV inhibition, suggesting the possibility of a feedback mechanism.

High versus low protein diets to mink--postprandial plasma urea and creatinine response, osmotic load and pattern of nitrogen and electrolyte excretion.

Nitrogen balance, pattern of excretion of nitrogenous end-products, endogenous urinary N excretion, postprandial plasma urea and creatinine, osmotic load, urinary electrolyte excretion and water intake/output relationships were studied in 12 adult female mink fed a high protein diet (HP; n = 6) providing about 155 g protein/kg or a low protein diet (LP; n = 6) providing about 95 g protein/kg. Two balance periods of each 3 d were used and diets were fed raw or cooked. After the last balance period followed a 48 h fasting period. Postprandial plasma urea and creatinine were studied for 48 h following a test meal given after an overnight fast. Osmotic load was determined based on collection of non-acidified urine carried out during 48 h. Level of protein supply did not affect N balance, being close to zero, whereas slightly negative balances were achieved for fasting animals. Protein supply was clearly reflected in excretion of urinary urea and allantoin but not in creatinine and uric acid. Endogenous urinary N excretion was estimated by a second order regression equation giving an intercept of 280 mg/kg0.75. Post-prandial plasma urea concentrations were strongly influenced by protein supply, HP animals having substantially higher peaks than LP animals, but values returned to fasting values within 24 h after the test meal. Plasma creatinine followed a biphasic pattern with a peak about 2 h after feeding and a nadir approximately 6 h after feeding. Physical form of diet influenced postprandial urea, animals fed raw diets having a higher peak, but not creatinine. The HP diet provided almost the double osmotic load of the LP diet and a corresponding increase in urine volume. The resulting water balances were identical irrespective of diet, showing that water intake/output relationships are very accurately regulated.

Energy intake and milk production in mink (Mustela vison)--effect of litter size.

Energy intake and milk production were measured in 12 mink dams raising litters of 3, 6 and 9 kits one to four weeks post partum by means of balance experiments and measurements of milk intake of the kits by the water isotope dilution technique. The dams were fed ad libitum on a conventional wet mink diet (DM: 323 g/kg; CP: 173 g/kg; ME: 4.4 MJ/kg). Milk samples collected from dams with corresponding litter sizes and lactation weeks, and body composition of kits nursed by these dams, were analysed for content of DM, ash, N and fat. The ME and drinking water consumption were higher in dams nursing 9 kits than in dams nursing 3 kits. The N and water balances as well as the live weight of dams were not affected by litter size. Daily milk production was higher in dams nursing 9 kits than in dams nursing 3 kits. The DM, N and fat content of the milk increased during lactation, but were not affected by litter size. Individual kit live weight was higher in litters of 3 than in litters of 6 and 9 kits four weeks post partum. The DM and fat content of the kits were lowest in kits from litters of 9 kits, whereas these kits had the highest protein content. Daily ME for maintenance of kits and the efficiency of utilisation of ME in milk for body gain were estimated to 356 kJ/kg0.75, kp approximately 0.53 and kf approximately 0.71, respectively. In conclusion, daily milk production increased with increasing litter size, but not in proportion to the number of kits, indicating that milk production limits the growth rate of the young. In the fourth week of lactation, milk production was not different between dams nursing 6 or 9 kits, indicating a maximum capacity.

Milk intake in blue fox (Alopex lagopus) and silver fox (Vulpes vulpes) cubs in the early suckling period.

Milk intake of fox cubs (2-16 days of age; body weight, 96-649 g) in ten blue fox litters and ten silver fox litters were measured by the water isotope dilution (WID) technique following a single intraperitoneal injection of tritiated water (3HHO). Litter size varied from four to 14 in blue foxes and from three to eight in silver foxes. Silver fox cubs had higher birth weights than blue foxes. Inter-species body weights and growth rates were apparently dependent on litter size and the dam's constitution. In both species growth rate increased with age and body weight (7-35 g per day). In the cubs, the biological half-life of body water turnover (BWT) rose from 1.5 days at 2-3 days of age to 2.5 days at 13-16 days of age, although a considerable scatter was seen. The mean daily milk intake of the cubs varied with body weight, from 31 to 193 g per day, whereas daily milk intake per unit of body mass remained stable at 30-35 g per 100 g body weight. The ratio of milk intake to body weight gain varied considerably among cubs, averaging 4.5 g/g during the 3-week experimental period. In suckling fox cubs, the calculated daily intake of metabolically energy (ME) corresponded fairly with the estimated energy requirements for growth and maintenance of the young. Finally, the applicability and the accuracy of the WID technique was evaluated in ten 3-week-old fox cubs, by tube-feeding with a milk replacer for 48 h, which documented that the daily rates of milk intake and water turnover can be accurately measured in suckling fox cubs by the WID technique following a single injection of 3HHO.

LH release in mink (Mustela vison). Pattern of the LH surge and effect of metabolic status.

The mink is a seasonal breeder with induced ovulation and delayed implantation. Reproductive processes are strongly influenced by energy supply and body condition. Items for which there is paucity or complete lack of data were the main objectives of this study: the temporal relationship between copulation and the pre-ovulatory LH surge and the influence of energy supply on LH release. A total of 30 yearling female mink with a well defined metabolic status was used. Twelve females kept in the laboratory were measured in six consecutive one-week balance periods each including the measurement of heat production by means of indirect calorimetry, and 18 females were kept under conventional farm conditions. The animals were fed so as to maintain energy balance (CON), flush fed by 2 weeks food restriction followed by 2 weeks refeeding (FLUSH), or kept in a negative energy balance (NEG). Plasma concentrations of the thyroid hormones, IGF-1 and insulin were determined weekly (n = 12), or 1 week after change in energy supply to the FLUSH group (n = 18). On the day of mating, blood samples for LH and oestradiol-17beta (E2) were taken before and immediately after mating and then 4, 8, 12, 24, 30 and 48 h thereafter. Frequent blood samplings, each lasting 60 min, were taken during the LH surge from two other females surgically fitted with venous access ports. Peak concentrations of LH were recorded on the first sampling, an average 16 min after mating. The concentrations remained elevated for 12 h, but almost decreased to basal values 24 h after mating. Plasma E2 was high before mating and peak values were attained 4 h after mating after which it decreased. Energy supply had no significant influence on LH and E2, but there was a tendency for a more sluggish LH release in NEG animals. The lack of response in FLUSH animals was explained by these animals having a lower intake of metabolisable energy than CON animals, the total intake not being significantly different from the NEG group. Plasma concentrations of thyroid hormones, IGF-1 and insulin were not significantly affected by the treatment, but in FLUSH animals the values mirrored energy supply, and in the NEG group, the values tended to decrease during the course of the experiment. It was concluded that the pre-ovulatory LH surge is an immediate response to mating, and that reproductive activity in the mink is maintained over a wide range of energy supply and body condition.

Milk intake of suckling kittens remains relatively constant from one to four weeks of age.

The daily milk intake of 14 domestic short-haired kittens (Felis catus) from five litters was estimated during wk 1-4 postpartum using the isotope dilution technique. Kittens received a single intraperitoneal injection of tritiated water, and blood samples were obtained from the jugular vein for radioactivity measurements at 2 and 96 h after injection. One kitten in each litter was used as a control to allow calculation of recycling of tritiated water. The mean (+/- SEM) biological half-life of tritiated water in the kittens increased from 2.4 +/- 0.1 d in wk 1 to 4.9 +/- 0.2 d in wk 4 postpartum. Recycling of tritiated water accounted for (mean +/- SEM) 5.9 +/- 0.8, 12.0 +/- 0.5, 7.7 +/- 1.3 and 10.0 +/- 1.3% of the kittens' daily water intake during postnatal wk 1-4, respectively. Daily milk intake of the kittens during wk 1-4 postpartum was 47.3 +/- 0.8, 47.4 +/- 1.5, 48.7 +/- 1.6 and 43.7 +/- 2.0 g, respectively. There was no effect of gender on milk intake. The daily metabolizable energy requirement of suckling kittens, estimated by multiple regression analysis, was 356 kJ/kg(0.75), whereas the metabolizable energy required per gram of gain was estimated to be 7. 8 kJ/d. The milk intake of suckling kittens remained relatively constant throughout the first 4 wk of lactation, and during this period, they seemed to have a lower energy requirement for maintenance.

Daily milk intake and body water turnover in suckling mink (Mustela vison) kits.

Daily (24 h) milk intake and body water turnover were measured in eight litters of suckling mink (Mustela vison) kits (6-9 kits litter-1) during weeks 1-4 post partum using the tritiated water (3HHO) dilution technique. The biological half-life of body water turnover in the mink kits increased linearly from 0.9 days in week 1 (3-5 days post partum) to 1.9 days in week 4 (22-24 days post partum). The daily milk intake varied markedly among the mink kits within a litter and increased significantly with increasing body mass from (mean +/- SEM) 10.9 +/- 0.4 g per kit during week 1 to 27.7 +/- 1.0 g per kit during week 4. Throughout the study, male kits were approximately 10% heavier and had a significantly higher milk intake than female kits. The results were corrected for water recycling between the dam and her kits, ranging from approximately 4 to 15% of the daily milk water intake, and the calculated daily milk yield of the 2 year old lactating mink dams increased from 87 +/- 7 g day-1 in week 1 to 190 +/- 15 g day-1 in week 4 post partum. The average body growth rate of the mink kits ranged from 2.9 g kit-1 per day in week 1 to 5.4 g kit-1 per day in week 4, and the calculated mean intake of mink milk per unit of body weight gain was remarkably stable at 1.0 (g g-1) during weeks 1-3 post partum, but increased to 5.6 (g g-1) in week 4 post partum. The amount of metabolizable energy supplied to the kits by the daily milk yield of the dam increased from approximately 450 to approximately 990 kJ day-1, which corresponded well with the calculated daily energy requirements of the kits. The tritiated water dilution technique was found feasible and reliable for repeated measurements of milk intake in suckling mink kits up to 4 weeks of age.

Nitrogen balance in adult female mink (Mustela vision) in response to normal feeding and short-term fasting.

Ten adult female mink (Mustela vison) were studied in a 7 d balance experiment consisting of a 2 d pre-surgery feeding period, followed by surgery, 1 d of recovery 4 d of ad libitum feeding, and a 2 d fasting period. In this experiment (Expt A) the animals had osmotic pumps implanted for continuous release of radioactively-labelled p-aminohippuric acid (p-aminobenzoyl-2-[3H]glycine; [3H]PAH; n 10) and 14C-labelled inulin ([14C]IN; n 5). Repeated 24 h collections of urine, corrected to 100% [3H]PAH or [14C]IN recovery, were used for accurate determination of N balances, 24 h urinary excretion of urea, creatinine, and total N, and calculation of mean 24 h renal clearance rates for endogenous creatinine and inulin. N balances were slightly below zero, but not significantly different between feeding and fasting periods, indicating that correction to 100% [3H]PAH recovery resulted in slight overestimation of the final balances. During fasting, withdrawal of the dietary water and protein loads resulted in a dramatic decline in 24 h urinary volume, and urea and creatinine excretion. Large individual variations in 24 h urinary creatinine excretion (with relative variation coefficients up to 30%) confirmed that this is an unreliable index of the completeness of urine collection. In this respect, recovery rates of [3H]PAH proved far more consistent. Renal clearance values obtained in fed mink were in fair agreement with published data from cats, dogs and ferrets (Mustela putorius furo). Inulin clearance was about 30% higher than endogenous creatinine clearance, although its decline in response to fasting was not significant. In a separate study (Expt B) another ten female mink were equipped with osmotic pumps containing [3H]PAH for determination of 24 h excretion rates of purine derivatives. During feeding, allantoin accounted for more than 97% of the excretion of purine derivatives in urine, uric acid making up less than 2.5%, xanthine and hypoxanthine less than 1%. In fasted animals, urinary excretion of each of these purine derivatives declined to less than 50% of the feeding value. In conclusion, an experimental technique is presented for efficient and accurate measurements of daily urinary excretion of nitrogenous constituents, which allows for correct determination of N balances in adult mink and, presumably, in other mammalian species.

Effects of feeding and short-term fasting on water and electrolyte turnover in female mink (Mustela vison).

Daily (24 h) rates of water and electrolyte turnover were measured in a conventional balance study in ten adult female pastel mink (Mustela vison) given free access to a standard mink feed for a 1-week conditioning period, followed by a 4 d experimental period and a 2 d fasting period. Drinking water was available throughout. In addition, the completeness of urine collection and the fraction of urine collected with the faeces were determined using a new experimental technique based on 24 h recoveries of specific urinary markers such as tritiated p-aminohippuric acid ([3H]PAH) or 14C-labelled inulin ([14C]IN) continuously delivered by small Alzet osmotic pumps implanted intraperitoneally. During feeding the mean individual percentage recovery in urine of [3H]PAH released from the osmotic pumps ranged from 68 to 88% (median 78%). The mean percentage of urinary [3H]PAH recovered from faecal collections was 6% (range 3-12%). In response to fasting the mean individual percentage recovery of [3H]PAH in urine ranged from 62 to 78% (median 68%). For urinary [14C]IN the mean percentage recoveries in fed and fasted animals were 79 and 63% respectively. Furthermore, during fasting, withdrawal of the supplies of dietary water caused a slight but insignificant (P = 0.17) increase in the daily intake of drinking water and, hence, the animals maintained their normal water balance by a dramatic reduction in urine excretion (P < 0.001). At the same time urinary solute excretion declined significantly (P < 0.001), due in part to the cessation of dietary electrolyte intake and in part to reduced formation of urea, whereas urinary osmolality decreased only moderately. The mean 24 h balances of Na, K, Ca, Mg, Cl and P were close to zero and only minor differences between the feeding and fasting periods were observed. When corrected for the measured inaccuracies in urine collection the balance data obtained in the present study represent useful reference standards for normally fed and fasted non-growing mink and, to some extent, useful guidelines for future studies in experimental animals.

Incidence of nursing sickness and biochemical observations in lactating mink with and without dietary salt supplementation.

The impact of dietary sodium on the incidence of nursing sickness in mink dams and on the average litter biomass of 28 and 42 day old kits was studied. One group (n = 115 including 12 barren females) was given a standard feed mixture with a natural content of 0.53 g NaCl/MJ and another group (n = 115 including 8 barren females) was given the same feed mixture supplemented with NaCl to a final content of 1.00 g/MJ. The average dam weight at weaning was significantly lower (P < 0.001) and the incidence of nursing sickness during the last part of the lactation period 3 times higher in the nonsupplemented group. The average litter biomass at weaning did not differ between the 2 experimental groups. A number of biochemical markers of preclinical nursing sickness, e.g. plasma aldosterone and osmolality, Na+ and Cl concentrations in plasma and urine, were studied during the last part of the lactation period and at weaning in 20 dams of the nonsupplemented group, in 10 dams of the salt supplemented group and, for comparison, in 5 + 5 barren females on the day corresponding to day 34 after parturition in nursing mink. The nonsupplemented group had significantly lower concentrations of sodium and chloride in plasma and urine and a significantly higher concentration of plasma aldosterone as compared to the salt supplemented group. Distinct signs of relative salt deficiency and preclinical nursing sickness thus characterized the nonsupplemented group throughout this period, while more blurred hints of electrolyte imbalances were noticed in the sodium chloride supplemented group at weaning. A beneficial effect of salt supplementation on the incidence of nursing sickness was shown; however, it remains unclear whether salt deficiency can cause nursing sickness or whether salt acts as an appetite stimulant preventing inanition and the development of the disorder.