A. C. Storm

Effect of dietary nitrogen content and intravenous urea infusion on ruminal and portal-drained visceral extraction of arterial urea in lactating Holstein cows

Urea extraction across ruminal and portal-drained visceral (PDV) tissues were investigated using 9 rumen-cannulated and multi-catheterized lactating dairy cows adapted to low-N (12.9% crude protein) and high-N (17.1% crude protein) diets in a crossover design. The interaction between adaptation to dietary treatments and blood plasma concentrations of urea was studied by dividing samplings into a 2.5-h period without urea infusion followed by a 2.5-h period with primed continuous intravenous infusion of urea (0.493+/-0.012 mmol/kg of BW per h). Cows were sampled at 66+/-14 and 68+/-12 d in milk and produced 42+/-1 and 36+/-1 kg of milk/d with the high-N and low-N diets, respectively. The arterial blood urea concentration before urea infusion was 1.37 and 4.09+/-0.18 mmol/L with low-N and high-N, respectively. Dietary treatment did not affect the urea infusion-induced increase in arterial urea concentration (1.91+/-0.13 mmol/L). Arterial urea extraction across the PDV and rumen increased from 2.7 to 5.4+/-0.5% and from 7.1 to 23.8+/-2.1% when cows were changed from high-N to low-N, respectively. Urea infusion did not decrease urea extractions, implying that urea transport rates were proportional to arterial urea concentrations. Urea extraction increased more across the rumen wall than across the total PDV for low-N compared with high-N, which implies that a larger proportion of total PDV uptake of arterial urea is directed toward the rumen with decreasing N intake. The ruminal vein - arterial (RA) concentration difference for ammonia increased instantly (first sampling 15 min after initiation of infusion) to the primed intravenous infusion when cows were adapted to the low-N diet. The RA difference for ammonia correlated poorly to the ventral ruminal concentration of ammonia (r=0.55). Relating the RA difference for ammonia to a function of both ruminal ammonia concentration and the RA difference for urea markedly improved the fit (r=0.85), indicating that a large fraction of ammonia released to the ruminal vein is absorbed from an epithelial ammonia pool not in equilibrium with the ventral ruminal ammonia pool. Changing cows from high-N to low-N affected the relative blood urea clearance by kidneys and PDV. The clearance by the kidneys decreased from 41 to 27+/-2 L/h and the clearance by the PDV increased from 52 to 105+/-12 L/h when the diet was changed from high-N to low-N. In conclusion, urea transport across gut epithelia in cattle is adapting to N status and driven by mass action. Data are commensurable with a model for urea transport across gut epithelia based on regulated expression or activity of facilitative urea transporters.

Effects of ruminal ammonia and butyrate concentrations on reticuloruminal epithelial blood flow and volatile fatty acid absorption kinetics under washed reticulorumen conditions in lactating dairy cows

The effect of reticuloruminal epithelial blood flow on the absorption of propionate as a volatile fatty acid (VFA) marker in 8 lactating Holstein cows was studied under washed rumen conditions. The cows were surgically prepared with ruminal cannulas and permanent catheters in an artery and mesenteric, right ruminal, and hepatic portal veins. The experiment was designed with 2 groups of cows: 4 cows adapted to high crude protein (CP) and 4 to low CP. All cows were subjected to 3 buffers: butyric, ammonia, and control in a randomized replicated 3 × 3 incomplete Latin square design. The buffers (30 kg) were maintained in a temporarily emptied and washed rumen for 40 min. The initial concentration of VFA was 84.2 mmol/L. Butyrate was increased from 4 to 36 mmol/L in butyric buffer by replacement of acetate, and ammonia (NH(3)) was increased from 2.5 to 22.5 mmol/L in ammonia buffer by replacement of NaCl. Increasing amounts of deuterium oxide (D(2)O) were added to the buffers as the order of buffer sequence increased (6, 12, and 18 g of D(2)O). Ruminal clearance of D(2)O was used to estimate epithelial blood flow. To increase accuracy of the epithelial blood flow estimates, data of ruminal liquid marker (Cr-EDTA), and initial and final buffer volumes were fitted to a dynamic simulation model. The model was used to estimate ruminal liquid passages, residual liquid, and water influx (saliva and epithelia water) for each combination of cow and buffer (n=24). Epithelial blood flow increased 49±11% for butyric buffer compared with control. The ruminal disappearance of propionate (marker VFA) was affected by buffer and followed the same pattern as for epithelial blood flow. The correlation between ruminal disappearance of propionate and epithelial blood flow (r=0.56) indicates that the removal of propionate can be limited by epithelial blood flow. The ruminal disappearance of propionate increased 30±12% for the butyric compared with ammonia buffer and 12.5±8% when compared with control. The net portal flux of propionate increased 32±6% in butyric compared with control. In conclusion, rumen epithelial blood flow is positively correlated with ruminal disappearance of propionate and affects the kinetics of ruminal VFA absorption.

Effects of particle size and dry matter content of a total mixed ration on intraruminal equilibration and net portal flux of volatile fatty acids in lactating dairy cows.

Effects of physical changes in consistency of ruminal contents on intraruminal equilibration and net portal fluxes of volatile fatty acids (VFA) in dairy cows were studied. Four Danish Holstein cows (121+/-17 d in milk, 591+/-24 kg of body weight, mean+/-SD) surgically fitted with a ruminal cannula and permanent indwelling catheters in the major splanchnic blood vessels were used. The experimental design was a 4x4 Latin square with a 2x2 factorial design of treatments. Treatments differed in forage (grass hay) particle size (FPS; 3.0 and 30 mm) and feed dry matter (DM) content of the total mixed ration (44.3 and 53.8%). The feed DM did not affect chewing time, ruminal variables, or net portal flux of VFA. However, decreasing the FPS decreased the overall chewing and rumination times by 151+/-55 and 135+/-29 min/d, respectively. No effect of the reduced chewing time was observed on ruminal pH or milk fat percentage. Cows maintained average ventral ruminal pH of 6.65+/-0.02, medial ruminal pH of 5.95+/-0.04, and milk fat of 4.42+/-0.12% with chewing time of 28.0+/-2.1 min/kg of DM when fed short particles. The medial ruminal pool of wet particulate matter was decreased by 10.53+/-2.29 kg with decreasing FPS, thereby decreasing the medial pool of total VFA, acetate, propionate, butyrate, isobutyrate, and isovalerate by 1,143+/-333, 720+/-205, 228+/-69, 140+/-51, 8.0+/-2.3, and 25.2+/-5.6 mmol, respectively. Ventral pool variables were not affected by treatments. Relatively large intraruminal differences of VFA concentrations and pH between the ventral and medial pools were observed, VFA concentrations being largest and pH being the lowest medially. This indicates that the ruminal mat acts as a barrier retaining VFA. The effects of reduced FPS were limited to the VFA pool sizes of the mat, leaving ruminal pH, ruminal VFA concentrations, and net portal flux of VFA unaffected. Consequently reduced FPS affected the intraruminal equilibration of VFA between mat and ventral rumen with an estimated turnover rate of isobutyrate increasing from 66+/-3%/h with long particles to 81+/-3%/h [corrected] with short particles. The estimated ruminal fluid flow and therefore intraruminal VFA transport between medial and ventral phase was not affected by the FPS. In conclusion, the ruminal mat pool of VFA was proportional to the size of the mat and the only detected effects of decreasing FPS were decreasing the mat size and an increasing turnover of the mat pool of VFA.

A model of ruminal volatile fatty acid absorption kinetics and rumen epithelial blood flow in lactating Holstein cows

Ruminal absorption of volatile fatty acids (VFA) is quantitatively the most important nutrient flux in cattle. Historically, VFA absorption models have been derived primarily from ruminal variables such as chemical composition of the fluid, volume, and pH. Recently, a mechanistic model incorporated the control of VFA absorption from epithelial surface area of the reticulorumen. In the present study, we hypothesized that ruminal absorption of VFA was controlled through epithelial permeability to VFA and rumen epithelial capillary blood flow. The objective of the study was to construct a model of VFA exchange across the rumen wall that incorporates epithelial blood flow as a driving force for ruminal VFA removal. The bidirectional fluxes between the ruminal and epithelial pool of VFA were assumed mass action driven, given that passive diffusion of nonionized VFA is the dominant transmembrane VFA flux. Parameter estimates were derived by fitting the model to observed data. The model provided reliable unbiased estimates of ruminal VFA absorption and rumen epithelial blood flow. Blood flow was modeled using an equation that considered the effect of butyrate and dietary crude protein intake per kilogram of body weight. The rate constants related to the flux from ruminal fluid to epithelium were in the order isobutyrate < acetate < propionate < butyrate (0.32 ± 0.02, 0.72 ± 0.2, 0.91 ± 0.06, and 0.97 ± 0.02 /h, respectively). The rate constants for fluxes of isobutyrate, acetate, propionate, and butyrate from the rumen epithelium to the ruminal fluid, relative to the pool size of the epithelium, were 4.78, 10.6, 13.4, and 14.3 /h, respectively. Ruminal concentrations of acetate, propionate, butyrate, and isobutyrate were predicted with root mean square prediction errors as percentage of the observed means (RMSPE) of 5.86, 5.75, 11.3, and 4.12, respectively. The epithelial blood flow was predicted with 26.3% RMSPE. Sensitivity analyses indicated that when ruminal butyrate concentration increased from 4.0 to 37.4 mmol/L, blood flow of the epithelium increased 47% and the ruminal disappearance rate of propionate increased 11%. The concentration gradient of propionate between ruminal fluid and epithelium was no more than 3:1 and increased with increasing blood flow. In conclusion, a dynamic model based on rumen epithelial blood flow and bidirectional fluxes of VFA between ruminal fluid and epithelium gave unbiased predictions with low residual error of ruminal VFA absorption under washed rumen conditions. The model indicates that the effect of varying epithelial blood flow on the control of ruminal VFA absorption is related to the concentration gradient of individual VFA between ruminal fluid and epithelial blood. Epithelial blood flow may be an important determinant of ruminal absorption of VFA, a result that has not been evaluated on independent data.

Technical note: A method for quantification of saliva secretion and salivary flux of metabolites in dairy cows

Salivary flow and net jugular flux of metabolites were studied during resting and rumination in 3 lactating dairy cows (BW 548 ± 17.2 kg, days in milk 113 ± 4 d). The method was based on the concentration difference between arterial and jugular blood, and jugular blood flow measured by downstream dilution of p-aminohippuric acid (pAH). Cows were surgically prepared with a permanent arterial catheter in A. intercostales dorsales before the trial. On sampling days, cows were prepared with left and right side jugular, and ear vein catheters for blood sampling and infusion of pAH, respectively. Blood was sampled simultaneously from the 2 jugular veins and artery during periods of rest and rumination. Secretion of saliva was set equal to the net water extraction calculated from the increased hemoglobin concentration in jugular blood compared with arterial blood. Arterial and jugular blood flow summed for both sides of the head doubled (P < 0.001) during rumination (437 ± 19, 424 ± 18 L/h, respectively) compared with resting (210 ± 19, 202 ± 18 L/h, respectively), consequently doubling the saliva secretion (P < 0.001, resting = 7.6 ± 0.8 L/h, rumination = 13.8 ± 0.8 L/h). The extraction of inorganic phosphate (Pi) from arterial blood during resting periods was greater compared with rumination (P = 0.004; resting = 21.7% ± 0.9%; rumination = 15.6% ± 0.9%), resulting in a greater Pi concentration in saliva secreted during resting. The concentrations of Pi in saliva were 4.5 ± 0.3 and 3.7 ± 0.3 times the arterial concentration during resting and rumination (P = 0.09), respectively. The urea concentration in saliva was 0.63 ± 0.04 times the arterial level, showing that urea is less efficiently transferred from blood than water, resulting in a greater numerical urea concentration in jugular compared with arterial blood. The water extraction method presented in the present paper offers an alternative way of estimating saliva secretion without the chewing activity constraints associated with other methods, for example, allowing for determination of saliva flow during rumination.

Mammary nutrient uptake in multiparous sows fed supplementary arginine during gestation and lactation1

Arginine is the precursor for the synthesis of nitric oxide and may increase mammary plasma flow (MPF), which may in turn increase mammary nutrient uptake. Quantifying mammary nutrient uptake improves our understanding of mammary nutrient metabolism and may potentially allow identification of limiting nutrients for colostrum and milk production. Thus, the objectives of the present study were 1) to study the impact of 25 g/d of crystalline Arg (ARG) on MPF and uptake of nutrients by the mammary glands compared with an isonitrogenous supply of Ala (51 g/d; control [CON]) fed to a total of 8 sows from d 30 of gestation until weaning on d 28 of lactation and 2) to quantify mammary nutrient uptake in late gestation and in early and at peak lactation. Sows were surgically fitted with indwelling catheters on d 76 ± 2 SEM of gestation. -amino hippuric acid (AH) was infused (3.0 mmol/h) in the infusion catheter inserted in the mammary vein, initiated 1 h before the first blood sample at -10, -3, 3, and 17 d in milk (DIM). Blood samples were simultaneously drawn from catheters inserted in the femoral artery and the mammary vein, and the samples were collected in hourly intervals from 0.5 h before to 6.5 h after feeding. Sow milk production was assessed at 3 and 17 DIM. Arterial plasma concentrations of Arg and Ala were increased in ARG and CON sows, respectively ( < 0.01), whereas we did not succeed in detecting a greater MPF in ARG sows ( = 0.30). Arterial-venous differences ( = 0.03) and net mammary flux ( = 0.01) of Ala were increased in CON sows, while the net flux of most other metabolites ( > 0.05) was unaffected by treatment. The mammary extraction of all essential AA was below 13% in late gestation. The average mammary extraction of essential AA at peak lactation was greatest for Leu (51%), while the preprandial extraction was greatest for Lys (57%). The mammary carbon balance (input-output) was negative (-39 ± 12 mol C/d) in early lactation but almost balanced at peak lactation (-13 ± 14 mol C/d), suggesting that mammary fat depots contributed to milk synthesis. In conclusion, we failed to observe an increased MPF and mammary uptake of AA and energy metabolites in ARG-supplemented sows. The mammary extraction rate of essential AA indicated that AA were not limiting for the mammary glands in late gestation, while Lys and Leu appeared to be the 2 most limiting essential AA for milk production at peak lactation.

Forage fiber effects on particle size reduction, ruminal stratification, and selective retention in heifers fed highly digestible grass/clover silages 1

The objective of this study was to investigate the effect of NDF content in highly digestible grass/clover silage on particle size reduction, ruminal stratification, and selective retention in dairy heifers. The reduction in particle size from feed to feces was evaluated and related to feed intake, chewing activity, and apparent digestibility. Four grass/clover harvests (Mixtures of Lolium perenne, Trifolium pratense, and Trifolium repens) were performed from early May to late August at different maturities, at different regrowth stages, and with different clover proportions, resulting in silages with NDF contents of 312, 360, 371, and 446 g/kg DM, respectively, and decreasing NDF digestibility with greater NDF content. Four rumen-fistulated dairy heifers were fed silage at 90% of ad libitum level as the only feed source in a 4 × 4 Latin square design. Silage, ingested feed boluses, medial and ventral ruminal digesta, and feces samples were washed with neutral detergent in nylon bags of 10-μm pore size, freeze dried, and divided into small (<0.212 mm), medium (0.212 to 1 mm), and large : LP; >1 mm) particles by dry-sieving. Chewing activity, rumen pool size, and apparent digestibility were measured. Intake of NDF increased linearly from 2.3 to 2.8 kg/d with greater NDF content of forages (P = 0.01), but silages were exposed to similar eating time (P = 0.55) and rumination time per kg NDF (P = 0.35). No linear effect of NDF content was found on proportion of LP in ingested feed boluses (P = 0.31), medial rumen digesta (P = 0.95), ventral rumen digesta (P = 0.84), and feces (P = 0.09). Greater proportions of DM (P < 0.001) and particulate DM (P = 0.008) were found in medial ruminal digesta compared with ventral rumen, and differences in DM proportion increased with greater NDF content (P = 0.02). Particle size distributions were similar for digesta from the medial and ventral rumen regardless of NDF content of the silages (P > 0.13). The LP proportion was >30% of particles in the ventral and medial rumen, whereas in the feces, the LP proportion was <2%. Particle size stratification of the rumen was undetectable regardless of NDF content of the silages, stressing that the retention mechanism of large undigested particles lies elsewhere than with particle entrapment in the rumen mat. In this study, forage particle breakdown, ruminal stratification, and retention of particles in the rumen were not affected by NDF content of highly digestible grass/clover silages.

Technical note: Measurement of mammary plasma flow in sows by downstream dilution of mammary vein infused para-aminohippuric acid1

The objectives of the present study were to design a method to estimate mammary plasma flow (MPF) in lactating sows using downstream dilution of -aminohippuric acid (AH) and to compare these estimates with MPF estimates based on specific AA as internal markers (MPF-AA). A permanent indwelling catheter was surgically implanted in the femoral artery, and another 2 were inserted in the right cranial mammary vein of 8 second- and third-parity sows on d 76 ± 2 SEM of gestation. On the 3rd and 17th days in milk, arterial and venous blood samples were drawn in hourly intervals from 0.5 h before until 6.5 h after feeding. The MPF in the right cranial mammary vein was measured by downstream dilution of infused AH (3.0 mmol/h). Total MPF-AH was calculated assuming that the measured flow constituted the flow from 5 out of 14 suckled glands on the basis of the anatomical structure of the mammary vascular system. Total MPF-AA was estimated on the basis of the output of the specific AA marker in milk and the arteriovenous differences of the marker as free AA in plasma, assuming a direct transfer of AA from plasma to milk protein. Total MPF-AH was 6,860 L/d in early lactation and increased to 8,953 L/d at peak lactation ( = 0.003). In early lactation, MPF-AA estimates were greater or tended to be greater (132% to 175%; < 0.10) than MPF-AH estimates for all internal markers, except Met (119%). Moreover, MPF-AH was correlated with MPF-AA only for MET as an internal marker ( = 0.74; = 0.03) in early lactation. In contrast, MPF-AH and MPF-AA estimates did not differ and were well correlated at peak lactation with the strongest correlation observed when Met ( = 0.84; = 0.009) and Phe + Tyr ( = 0.82; = 0.01) were used as the internal AA markers. Litter gain increased from d 3 to 17 of lactation (2.13 vs. 3.46 g/d; = 0.001) and was correlated with MPF-AH during lactation ( = 0.74; < 0.001), whereas no correlation between litter gain and MPF-AA was observed ( > 0.10). These results suggest that downstream dilution of infused AH and the AA methods are applicable methods to estimate MPF at peak lactation. The reason for the observed discrepancy in early lactation between MPF- AH and MPF-AA is not obvious but might be related to the rapid metabolic changes observed in early lactation. In conclusion, MPF measured by downstream dilution of mammary infused AH was higher at peak compared to early lactation, which the internal AA marker approach failed to show.

Effect of propylene glycol on adipose tissue mobilization in postpartum over-conditioned Holstein cows

Our objective was to investigate the quantitative and qualitative effects of propylene glycol (PG) allocation on postpartum adipose tissue mobilization in over-conditioned Holstein cows. Nine ruminally cannulated and arterially catheterized cows were, at parturition, randomly assigned to a ruminal pulse dose of either 500g of tap water (n=4) or 500g of PG (n=5) once a day. The PG was given with the morning feeding for 4 wk postpartum (treatment period), followed by a 4-wk follow-up period. All cows were fed the same prepartum and postpartum diets. At -16 (±3), 4 (±0), 15 (±1) and 29 (±2) days in milk (DIM), body composition was determined using the deuterium oxide dilution technique, liver and subcutaneous adipose tissue biopsies were collected, and mammary gland nutrient uptake was measured. Weekly blood samples were obtained during the experiment and daily blood samples were taken from -7 to 7 DIM. Postpartum feed intake and milk yield was not affected by PG allocation. The body content of lipid was not affected by treatment, but tended to decrease from 4 to 29 DIM with both treatments. Except for the first week postpartum, no difference in plasma nonesterified fatty acids concentration was noted between treatments in the treatment period. Yet, PG allocation resulted in decreased plasma concentrations of β-hydroxybutyrate (BHB) and increased plasma concentrations of glucose. In the follow-up period, plasma concentrations of nonesterified fatty acids, glucose, and BHB did not differ between treatments. Additionally, the change in abundance of proteins in adipose tissue biopsies from prepartum to 4 DIM was not affected by treatment. In conclusion, the different variables to assess body fat mobilization were concurrent and showed that a 4-wk postpartum PG allocation had limited effect on adipose tissue mobilization. The main effect was an enhanced glucogenic status with PG. No carry-over effect of PG allocation was recorded for production or plasma metabolites, and, hence, a new period of metabolic adaption to lactation seemed to occur with PG treatment after ceasing PG allocation. Thus, PG seemed to induce a 2-step adaption to lactation, reducing the immediate postpartum nadir and peak of plasma concentration of glucose and BHB, respectively; which is beneficial for postpartum cows at high risk of lipid-related metabolic diseases.

Effect of postpartum propylene glycol allocation to over-conditioned Holstein cows on concentrations of milk metabolites.

The objective of the study was to investigate the effect of propylene glycol (PG) allocation on concentrations of milk metabolites with potential use as indicators of glucogenic status in high yielding postpartum dairy cows. At time of calving, nine ruminally cannulated Holstein cows were randomly assigned to ruminal dosing of 500 g/d tap water (CON, n = 4) or 500 g/d PG (PPG, n = 5). The PG was given with the morning feeding week 1-4 postpartum (treatment period) and cows were further followed during week 5-8 postpartum (follow-up period). All cows were fed the same postpartum diet. Milk samples were obtained at each milking (3 times/d) in the treatment period, and at morning milking during the follow-up period. Weekly blood samples were obtained from -4 to +8 weeks relative to calving and daily blood samples from -7 until +7 d relative to calving. The main effect of PG allocation was an increased glucogenic status, e.g. visualised by a prompt marked increase in blood fructosamine. During the treatment period, milk concentration of free glucose tended to be greater, whereas milk concentrations of isocitrate and BHBA were lower for PPG compared with CON. It is proposed that the ratio between free glucose and isocitrate in milk may be a potential biomarker for glucogenic status in the vulnerable early postpartum period. We will pursue this issue in the future.