Feed restriction of lactating cows triggers acute downregulation of mammary mammalian target of rapamycin signaling and chronic reduction of mammary epithelial mass.

While there is generally no consensus about how nutrients determine milk synthesis in the mammary gland, it is likely that the mechanistic target of rapamycin (mTOR) complex 1 (mTORC1) plays a role as a key integrator of nutritional and mitogenic signals that can influence a multitude of catabolic and anabolic pathways. The objectives of this study were to evaluate acute changes (<24 h) in translational signaling, in addition to chronic changes (14 d) in mammary gland structure and composition, in response to a severe feed restriction. Fourteen lactating Holstein dairy cows were assigned to either ad libitum feeding (n = 7) or a restricted feeding program (n = 7). Feed-restricted cows had feed removed after the evening milking on d 0. Mammary biopsies and blood samples were collected 16 h after feed removal, after which cows in the restricted group were fed 60% of their previously observed ad libitum intake for the remainder of the study. On d 14, animals were slaughtered and their mammary glands dissected. In response to feed removal, an acute increase in plasma nonesterified fatty acid concentration was observed, concurrent to a decrease in milk yield. In mammary tissue, we observed downregulation of the mTORC1-S6K1 signaling cascade, in addition to reductions in mRNA expression of markers of protein synthesis, endoplasmic reticulum biogenesis, and cell turnover (i.e., transcripts associated with apoptosis or cell proliferation). During the 14 d of restricted feeding, animals underwent homeorhetic adaptation to 40% lower nutrient intake, achieving a new setpoint of 14% reduced milk yield with 18% and 29% smaller mammary secretory tissue DM and CP masses, respectively. On d 14, no treatment differences were observed in markers of protein synthesis or mammary cell turnover evaluated using gene transcripts and immunohistochemical staining. These findings implicate mTORC1-S6K1 in the early phase of the adaptation of the mammary gland's capacity for milk synthesis in response to changes in nutrient supply. Additionally, changes in rates of mammary cell turnover may be transient in nature, returning to basal levels following brief alterations that have sustained effects.

Relationship between rate of glucose or propionate infusion and milk protein yield and concentration in dairy cows: A meta-regression.

Although postruminal glucose infusion into dairy cows has increased milk protein yield in some past experiments, the same trend has not been observed in others. A meta-regression of 64 sets of observations from 29 previously published glucose and propionate infusion studies in dairy cattle, treating study and experiment (study) as random effects, was performed to establish the general effects of glucose equivalent (GlcE) infusion rate on milk true protein (MTP) yield and content, if any, and to identify independent, fixed-effect variables that accounted for the changes in MTP yield and content that were observed. Candidate explanatory variables included rate and site of infusion, diet composition and intake, body weight and lactation stage of the cows, and the change in nutrient intake between GlcE and control treatments. Across all studies, according to a model containing only the random effects of study and experiment, GlcE infusion at an average of 954 g/d increased MTP yield by 26 g/d, on average, whereas mean MTP content was not affected. Backward stepwise elimination of potential explanatory variables from a full mixed model produced a final, reduced model for MTP yield that retained a positive, second-order quadratic effect of infusion rate of GlcE and a positive, linear effect of the change in crude protein intake (CPI) between GlcE treatment and control. This change in CPI due to GlcE infusion ranged from -0.546 to 0.173 kg/d in the dataset. The model fit indicated that when CPI was allowed to drop during GlcE infusion, the effect of GlcE on MTP yield was smaller than when CPI was maintained or increased, in a manifestation of the classic protein:energy interaction. The final reduced model for MTP content contained the same explanatory variables as for MTP yield, plus a negative effect of intravenous compared with gastrointestinal infusion. Overall, the meta-analysis revealed that both MTP yield, and content were positively related to GlcE infusion rate and to the change in CPI between glucose treatment and control.

Behavioral and physiological responses to an inspired-air supplemental cooling system for dairy cows in free-stall housing.

During heat stress, dairy cows spend less time lying down to dissipate heat. Heat stress abatement strategies generally target cows outside of their resting areas. However, cooling cows while in their stalls could help alleviate heat stress without compromising lying behavior. The objective of this study was to assess the effects of an inspired-air supplemental cooling system (SCS) on respiration rate, rectal temperatures, lying behavior, rumination time, and milk production (energy-corrected milk, ECM) of lactating dairy cows. A free-stall pen was retrofitted with custom stall partitions to deliver cooled air and mist. The pen, including the stall platform, was divided into two separate sides. Twenty-eight lactating Holstein cows were randomly sorted into two groups, each housed on one side of the experimental pen. Cows experienced four treatments (control, CTRL; cooled air, AIR; mist, MIST; cooled air and mist, AIR + MIST) in a four-treatment, four-period, two-sequence crossover design, with each period lasting seven days. Cooled air was provided continuously, and mist was cycled 3 min on, 12 min off from 0900 to 2100 h. Respiration rates were observed hourly between 0900 and 1500 h, and only measurements recorded while cows were lying down were used in the analysis. Rectal temperatures of 16 focal cows were recorded at 1545 h once per day. Lying behavior and rumination were recorded continuously, and milk yields recorded twice daily were used to calculate ECM. Throughout the experimental period, the average temperature-humidity index was 66.4 ± 6.07. During the MIST treatment, the respiration rate was lower than the CTRL (45.7 vs 49.0 ± 1.92 breaths/min) and AIR (45.7 vs 48.7 ± 1.92 breaths/min). CTRL and AIR did not differ (48.7 vs 49.0 ± 1.92 breaths/min), and MIST and AIR + MIST (45.7 vs 47.1 ± 1.92 breaths/min) did not differ. All other variables were not significantly different between treatments. In conclusion, the SCS appeared to be tolerated by cows and shows the potential to assist in alleviating heat stress. The cooling capacity needs to be evaluated under more extreme environmental conditions causing heat stress than those experienced during this study. Further testing is required to determine the cooled air temperature specifications and mist delivery frequency.

Review: Influence of postabsorptive metabolism on essential amino acid partitioning in lactating dairy cows.

In the lactating cow, essential amino acids (EAAs) absorbed from the gut are partitioned to mammary and extra-mammary tissues via blood plasma circulation. There is also entry of EAA into plasma from the breakdown of proteins in the cow's body. A balance model across plasma was solved to integrate entry rates of branched-chain (BCAA) and non-branched-chain EAA (NBAA) with their corresponding rate constants for clearance by mammary glands and the remainder of the body, for selected glucose and fat infusion experiments. Endogenous EAA entry from whole-body proteolysis was reduced by glucose and unchanged or increased by fat, the efficiency of net plasma BCAA clearance by mammary and extra-mammary tissues was elevated by glucose but slightly reduced by fat, and the efficiency of extra-mammary NBAA clearance may have decreased during glucose infusion but it was not affected or slightly increased by fat. These differences between glucose and fat responses can be accounted for by insulin and glucagon. Insulin suppresses endogenous EAA entry through mechanistic target of rapamycin complex 1, integrated stress response, and glycogen synthase kinase 3 signaling networks in skeletal muscle. While these networks can also regulate protein synthesis rates in muscle and the extra-mammary body, they exhibit low sensitivities to insulin in lactating ruminants. However, in the mammary glands, via these same networks, insulin stimulates clearance of EAA from plasma, although the drive to maintain a set point for milk protein yield takes precedence over nutritional signals. The glucose-induced increase in mammary BCAA clearance without an effect on NBAA clearance is due to a pronounced decrease in plasma BCAA concentrations. Because NBAAs do not experience a similar decline in concentration, the BCAA effect must be due to their metabolic transformation as opposed to sequestration in proteins. In adipose, the products of BCAA catabolism are lipogenic precursors. We propose that faster lipogenesis in adipose tissue, stimulated by glucose infusion, also promotes the uptake of precursor BCAA from plasma, causing a drop in their circulating concentrations. In addition, insulin stimulates BCAA oxidation in muscle as an alternative fuel to fatty acids. A lower efficiency of extra-mammary NBAA clearance during glucose infusion may be the consequence of decreased hepatic expression of AA-catabolizing enzymes in response to low glucagon concentration. The proportion of EAA entry partitioned to the mammary glands is a culmination of regulatory shifts at all of the points discussed above according to a regulated or unfair competition between mammary and extra-mammary processes.

Development of a novel stall design for dairy cattle: Part II. The effect of minimal stall partitioning on lying behavior, rumination, stall cleanliness, and preference.

Free-stalls for dairy cows promote cleanliness, provide cows with a defined space to lie down, and decrease labor and bedding required for maintenance. However, current stall features can restrict behavior and reduce stall use. The objective of this study was to assess the short-term effect of a novel free-stall design (flexible single-bar partition, no neck rail, increased slope) on stall cleanliness and the lying behavior, rumination, milk production, and preferences of dairy cows in comparison to standard free-stalls (metal loop partitions, neck rail). In the first experiment, 60 Holstein cows were randomly divided into two groups and switched between standard and novel stalls in a replicated crossover design with four 7-day periods. Lying behavior and rumination were recorded continuously. Milk yields were recorded 2x/day, and stall cleanliness scores were collected on the last four days of weeks 3 and 4. The second experiment was a 6-day preference test where 14 cows from experiment 1 were given free access to 16 standard and 16 novel stalls. Lying behavior was recorded continuously. On day 6, standing, perching, and intention, lying, and rising movements were recorded. In experiment 1, cows spent 12 min less time lying down (14.1 vs 14.3 ± 0.48 h/d), had one fewer lying bouts (8.8 vs 9.8 ± 0.23 bouts/d), and had longer lying bouts by 6 min (1.4 vs 1.5 ± 0.05 h/bout) in novel stalls. Rumination time (547.3 vs 548.9 ± 4.66 min/d) and milk yield (35.0 vs 35.2 ± 0.51 kg/d) did not differ between stall types. However, novel stalls were more than twice as soiled as standard stalls (32.8 vs 14.2 ± 1.38% of rear half soiled). In experiment 2, the cows most recently housed in the novel stalls showed a clear preference for lying down in those stalls, whereas cows most recently housed in standard stalls showed no preference. Intention and lying down movements were longer in standard stalls when other cows were present in diagonally opposite stalls, but were similar between stall types when not occupied. All cows preferred standing in novel stalls. Novel stalls may have improved stall use compared to standard free-stalls, and the design requires further development to address cleanliness concerns. Future work is required to provide comfortable stalls without sacrificing cleanliness.

Development of a novel stall design for dairy cattle: Part I. The effect of an increased slope on lying behavior, rumination, cleanliness, and preference.

The longitudinal slope of a stall is important for positioning cows in the stall and maintaining cleanliness of the stall surface. The objectives of this study were to determine the short-term effect of increasing free-stall slope from 4.5% to 9.3% on dairy cow lying behavior, rumination, milk production, cleanliness, and preference. In experiment 1, 60 multiparous Holstein cows were divided randomly into two groups and exposed to a 4.5% stall slope (standard in the research facility) and a 9.3% stall slope in a replicated crossover design with four 7-day periods. Each group of cows were housed in a pen with 30 free-stalls and switched between treatments weekly. Lying behavior and rumination were recorded continuously, milk yields were recorded twice per day, and cleanliness scores were collected on the last day of each week. Experiment 2 consisted of an 8-day preference test conducted with 14 cows from experiment 1 in an experimental pen with 32 stalls (16 stalls with a 4.5% slope and 16 stalls with a 9.3% slope). Continuous video monitoring was used to record time spent lying, standing, and perching in each treatment during the last 5 days, and lying time was used to assess preference. In experiment 1, cows spent 12 min/day less time lying down (12.8 vs 12.6 ± 0.28 h/d) and had 0.6 more bouts/day (9.5 vs 10.1 ± 0.38 no./d) with a shorter duration by 6 min/day (1.4 vs 1.3 ± 0.03 h/bout) in stalls with a 9.3% slope. Rumination was 5.5 min less per day in stalls with a 9.3% slope (578.4 vs 572.9 ± 16.56 min/d) and milk yield did not differ between treatments (33.6 vs 33.4 ± 0.78 kg/d). Stall slope did not affect cow cleanliness (1.6 vs 1.6 ± 0.05 points on a 1-5 scale). In experiment 2, cows showed no clear preference for lying, standing, or perching in either stall slope (4.5% and 9.3%) when given a choice. These results indicate that increasing stall slope from 4.5% to 9.3% marginally altered lying behavior, but did not interfere with stall use or influence cow preferences.

Associations between feed efficiency and aspects of lactation curves in primiparous Holstein dairy cattle.

Genetic selection for improved feed efficiency in dairy cattle has received renewed attention over the last decade to address the needs of a growing global population. As milk yield is a critical component of feed efficiency metrics in dairy animals, our objective was to evaluate the associations between feed efficiency in primiparous Holstein cattle and parameters of a mathematical model describing individual lactation curves. The Dijkstra lactation curve model was fit to individual lactation records from 34 Holstein heifers with previously estimated measures of feed efficiency. We found that the optimal fit of the Dijkstra model was achieved using daily milk yield records up to 21 d in milk to capture the rise to peak milk yield and using monthly dairy herd improvement records for the remainder of lactation to accurately characterize lactation persistency. In the period of lactation before peak milk yield, improved feed efficiency was associated with a faster increase in daily milk yield over a shorter period of time at the expense of increased mobilization of body reserves; this serves to reinforce the concept that dairy cattle are primarily capital breeders versus income breeders. Feed efficiency in the period following peak lactation, as measured by gross feed efficiency, return over feed costs, and net energy efficiency of lactation, was positively associated with higher peak milk yield. The findings in early lactation suggest that estimates of feed efficiency could be improved by evaluating feed efficiency relative to conception, rather than parturition and lactation, to better account for the energy stored and released from body reserves in capital breeding.

Effects of energy source in milk replacer on glucose metabolism of neonatal dairy calves.

Milk replacers (MR) formulated to contain more lactose than whole milk could potentially reduce insulin sensitivity in dairy calves. This study evaluated how partially replacing lactose in MR with fat on a weight/weight basis affects glucose-insulin kinetics in neonatal dairy calves fed high quantities of MR. Thirty-four Holstein bull calves were blocked by dam parity (1.6 ± 0.3) and randomly assigned to 1 of 2 treatments (n = 17): a high-lactose MR (HL; 46.1% lactose, 18.0% crude fat, and 23.9% crude protein on a dry matter basis) or a high fat MR (HF; 39.9% lactose, 24.6% crude fat, and 24.0% crude protein on a dry matter basis). Calves were individually housed and fed pooled colostrum at 1.5 h and 12 h postnatal at 18 and 9% of metabolic body weight (BW0.75), respectively. From 24 h postnatal until the end of the study (d 7), calves were transitioned to MR (prepared at 150 g/L) at 18% of BW0.75 twice daily at 0700 and 1900 h. On d 4, an insulin-modified intravenous glucose tolerance test was conducted 8 h after the morning meal by intravenous infusion of glucose (0.54 g/kg of BW0.75, min 0) and insulin (3.9 µg/kg of BW0.75, min 20). During postprandial sampling on d 6, abomasal emptying was assessed by dosing acetaminophen (0.13 g/kg of BW0.75) with MR and measuring its appearance within plasma. Sequential blood samples were collected and analyzed for plasma glucose, insulin, and acetaminophen concentrations. Calves were killed on d 7 and liver and pancreatic tissues were collected for histomorphological analysis. Partially replacing lactose in MR with fat increased body weight gain (679.6 vs. 462.0 ± 69.68 g/d) and gain:metabolizable energy intake (186.6 vs. 120.5 ± 20.21 g of body weight/Mcal) compared with feeding HL MR. Furthermore, abomasal emptying rate was reduced in calves fed HF MR. Calculated variables based on postprandial glucose and insulin concentrations, such as maximum concentration or area under the curve, were or tended to be lower in HF than in HL calves. Postprandial insulin sensitivity tended to be reduced in HL compared with HF, although insulin sensitivity estimated from the insulin-modified intravenous glucose tolerance test was not different. Calves fed HF MR showed enhanced glucose-stimulated insulin secretion. In conclusion, partially replacing lactose in MR with fat resulted in smaller fluctuations in postprandial glucose and insulin concentrations and tended to increase postprandial but not fasting insulin sensitivity in neonatal dairy calves.

Review: Synergy between mechanistic modelling and data-driven models for modern animal production systems in the era of big data.

Mechanistic models (MMs) have served as causal pathway analysis and 'decision-support' tools within animal production systems for decades. Such models quantitatively define how a biological system works based on causal relationships and use that cumulative biological knowledge to generate predictions and recommendations (in practice) and generate/evaluate hypotheses (in research). Their limitations revolve around obtaining sufficiently accurate inputs, user training and accuracy/precision of predictions on-farm. The new wave in digitalization technologies may negate some of these challenges. New data-driven (DD) modelling methods such as machine learning (ML) and deep learning (DL) examine patterns in data to produce accurate predictions (forecasting, classification of animals, etc.). The deluge of sensor data and new self-learning modelling techniques may address some of the limitations of traditional MM approaches - access to input data (e.g. sensors) and on-farm calibration. However, most of these new methods lack transparency in the reasoning behind predictions, in contrast to MM that have historically been used to translate knowledge into wisdom. The objective of this paper is to propose means to hybridize these two seemingly divergent methodologies to advance the models we use in animal production systems and support movement towards truly knowledge-based precision agriculture. In order to identify potential niches for models in animal production of the future, a cross-species (dairy, swine and poultry) examination of the current state of the art in MM and new DD methodologies (ML, DL analytics) is undertaken. We hypothesize that there are several ways via which synergy may be achieved to advance both our predictive capabilities and system understanding, being: (1) building and utilizing data streams (e.g. intake, rumination behaviour, rumen sensors, activity sensors, environmental sensors, cameras and near IR) to apply MM in real-time and/or with new resolution and capabilities; (2) hybridization of MM and DD approaches where, for example, a ML framework is augmented by MM-generated parameters or predicted outcomes and (3) hybridization of the MM and DD approaches, where biological bounds are placed on parameters within a MM framework, and the DD system parameterizes the MM for individual animals, farms or other such clusters of data. As animal systems modellers, we should expand our toolbox to explore new DD approaches and big data to find opportunities to increase understanding of biological systems, find new patterns in data and move the field towards intelligent, knowledge-based precision agriculture systems.

Effects of jugular infused methionine, lysine, and histidine as a group or leucine and isoleucine as a group on production and metabolism in lactating dairy cows.

Essential AA (EAA), particularly leucine, isoleucine, methionine, and histidine, possess signaling properties for promoting cellular anabolic metabolism, whereas methionine, lysine, and histidine are considered also to be substrate limiting AA. The objective of this study was to evaluate production responses to supplementation of 2 AA groups in a 2 × 2 factorial design. Eight cows (99 ± 18 days in milk) were assigned to 4 jugular infusion treatments consisting of saline (CON), methionine plus lysine plus histidine (MKH), isoleucine plus leucine (IL), or MKH plus IL, in a replicated 4 × 4 Latin square design. Periods were 18 d in length, comprising 8 d of rest followed by 10 d of jugular infusion. Daily infusion amounts were 21 g of methionine, 38 g of lysine, 20 g of histidine, 50 g of leucine, and 22 g of isoleucine. Cows were ad libitum fed a common diet consisting of 15.2% crude protein and 1.61 Mcal/kg NEL on a dry matter basis that was predicted to meet rumen degradable protein requirements but was 15% deficient in metabolizable protein. Milk and energy-corrected milk yields increased by 2.3 kg/d and 1.9 kg/d, respectively, with infused IL, and no change was observed for MKH. Milk protein concentration increased by 0.13 percentage units for MKH, whereas milk protein yield increased for both MKH and IL by 84 g/d and 64 g/d, respectively. The milk protein yield increase for MKH+IL was 145 g/d versus CON. Gross feed efficiency tended to increase with IL infusion, and N efficiency tended to increase with MKH infusion. Aggregate arterial EAA concentrations less Met, Lys, and His declined by 7.2% in response to MKH infusion. Arterial EAA less Ile and Leu also declined by 6.2% in response to IL infusion. Net total AA (TAA) and EAA uptake by the udder tended to increase in response to MKH infusion, whereas mammary blood flow increased in response to IL infusion, but TAA and EAA net uptakes were unaffected. Apparent udder affinity increased for TAA and EAA less Met, Lys, and His in response to MKH infusion, whereas affinity for EAA less Ile and Leu increased for IL infusion. Venous Met and Leu concentrations increased by 192% and 35% from the MKH and IL infusions, respectively, compared with CON, which indicates that intracellular concentration of these EAA changed substantially. Increases in milk protein yield were observed from 2 groups of amino acids independently and additively, which contradicts the single limiting amino acid theory that a single EAA will limit milk protein yield.

The dynamic behavior of feed efficiency in primiparous dairy cattle.

A variety of measures are currently used to evaluate feed efficiency in livestock, with some of the most popular being the linear regression of nutrient intake on the various energy sinks of the cohort being evaluated. The adoption of novel on-farm technologies has greatly increased the breadth and depth of data being collected on a daily basis, which creates the opportunity for the implementation of more advanced feed efficiency models. The objectives of the current study were to leverage data collected using high-throughput biometric technologies to evaluate changes of early-lactation feed efficiency on a daily basis and to develop a novel measure of feed efficiency based on the NRC net energy model. Daily records of nutrient intake, fat- and protein-corrected milk yield, body weight, and body condition score were used to model 7 different indices of feed efficiency (gross feed efficiency, return over feed costs, residual feed intake, residual net energy intake, and net energy efficiencies of lactation, maintenance, and body reserve flux) in primiparous Holstein heifers (n = 40) from 0 to 150 d in milk. We observed that feed efficiency was dynamic and fluctuated over the course of the experimental period regardless of the efficiency measure used, but most measures followed similar trends in changes of efficiency on a day-to-day basis. Of the measures evaluated, return over feed costs would be most suited for management purposes due to practicality and simplicity, whereas other measures, such as the efficiency of net energy flux to and from body reserves, present potential targets for inclusion in breeding goals. Net energy efficiency of lactation was estimated to have higher repeatability compared with residual feed or energy intake, which may reduce the time required to estimate whole-lactation efficiency. Because decreased net energy balance was strongly correlated with increased efficiency for all indices evaluated, future work investigating the associations of feed efficiency with the magnitude and duration of negative energy balance is warranted.

Single-step genome-wide association for longitudinal traits of Canadian Ayrshire, Holstein, and Jersey dairy cattle.

Estimating single nucleotide polymorphism (SNP) effects over time is essential to identify and validate candidate genes (or quantitative trait loci) associated with time-dependent variation of economically important traits and to better understand the underlying mechanisms of lactation biology. Therefore, in this study, we aimed to estimate time-dependent effects of SNP and identifying candidate genes associated with milk (MY), fat (FY), and protein (PY) yields, and somatic cell score (SCS) in the first 3 lactations of Canadian Ayrshire, Holstein, and Jersey breeds, as well as suggest their potential pattern of phenotypic effect over time. Random regression coefficients for the additive direct genetic effect were estimated for each animal using single-step genomic BLUP, based on 2 random regression models: one considering MY, FY, and PY in the first 3 lactations and the other considering SCS in the first 3 lactations. Thereafter, SNP solutions were obtained for random regression coefficients, which were used to estimate the SNP effects over time (from 5 to 305 d in lactation). The top 1% of SNP that showed a high magnitude of SNP effect in at least 1 d in lactation were selected as relevant SNP for further analyses of candidate genes, and clustered according to the trajectory of their SNP effects over time. The majority of SNP selected for MY, FY, and PY increased the magnitude of their effects over time, for all breeds. In contrast, for SCS, most selected SNP decreased the magnitude of their effects over time, especially for the Holstein and Jersey breeds. In general, we identified a different set of candidate genes for each breed, and similar genes were found across different lactations for the same trait in the same breed. For some of the candidate genes, the suggested pattern of phenotypic effect changed among lactations. Among the lactations, candidate genes (and their suggested phenotypic effect over time) identified for the second and third lactations were more similar to each other than for the first lactation. Well-known candidate genes with major effects on milk production traits presented different suggested patterns of phenotypic effect across breeds, traits, and lactations in which they were identified. The candidate genes identified in this study can be used as target genes in studies of gene expression.

Genome-wide association for milk production traits and somatic cell score in different lactation stages of Ayrshire, Holstein, and Jersey dairy cattle.

We performed genome-wide association analyses for milk, fat, and protein yields and somatic cell score based on lactation stages in the first 3 parities of Canadian Ayrshire, Holstein, and Jersey cattle. The genome-wide association analyses were performed considering 3 different lactation stages for each trait and parity: from 5 to 95, from 96 to 215, and from 216 to 305 d in milk. Effects of single nucleotide polymorphisms (SNP) for each lactation stage, trait, parity, and breed were estimated by back-solving the direct breeding values estimated using the genomic best linear unbiased predictor and single-trait random regression test-day models containing only the fixed population average curve and the random genomic curves. To identify important genomic regions related to the analyzed lactation stages, traits, parities and breeds, moving windows (SNP-by-SNP) of 20 adjacent SNP explaining more than 0.30% of total genetic variance were selected for further analyses of candidate genes. A lower number of genomic windows with a relatively higher proportion of the explained genetic variance was found in the Holstein breed compared with the Ayrshire and Jersey breeds. Genomic regions associated with the analyzed traits were located on 12, 8, and 15 chromosomes for the Ayrshire, Holstein, and Jersey breeds, respectively. Especially for the Holstein breed, many of the identified candidate genes supported previous reports in the literature. However, well-known genes with major effects on milk production traits (e.g., diacylglycerol O-acyltransferase 1) showed contrasting results among lactation stages, traits, and parities of different breeds. Therefore, our results suggest evidence of differential sets of candidate genes underlying the phenotypic expression of the analyzed traits across breeds, parities, and lactation stages. Further functional studies are needed to validate our findings in independent populations.

Invited review: Determination of large-scale individual dry matter intake phenotypes in dairy cattle.

Feed efficiency has been widely studied in many areas of dairy science and is currently seeing renewed interest in the field of breeding and genetics. A critical part of determining how efficiently an animal utilizes feed is accurately measuring individual dry matter (DM) intake. Currently, multiple methods are used to measure feed intake or determine the DM content of that feed, resulting in different levels of accuracy of measurement. Furthermore, the scale at which data need to be collected for use in genetic analyses makes some methodologies impractical. This systematic review aims to provide an overview of the current methodologies used to measure both feed intake in ruminants and DM content of feedstuffs, current methods to predict individual DM intake, and applications of large-scale intake measurements. Overall, advances in milk spectral data analysis present a promising method of estimating individual DM intake on a herd scale with further validation of prediction models. Although measurements of individual feed intake rely on the same underlying principle, the methods selected are largely dictated by the costs of capital, labor, and necessary analyses. Finally, DM methodologies were synthesized into a comprehensive protocol for use in a variety of feedstuffs.

Expression of genes related to energy metabolism and the unfolded protein response in dairy cow mammary cells is affected differently during dietary supplementation with energy from protein and fat.

Secretory capacity of bovine mammary glands is enabled by a high number of secretory cells and their ability to use a range of metabolites to produce milk components. We isolated RNA from milk fat to measure expression of genes involved in energy-yielding pathways and the unfolded protein response in mammary glands of lactating cows given supplemental energy from protein (PT) and fat (FT) tested in a 2 × 2 factorial arrangement. We hypothesized that PT and FT would affect expression of genes in the branched-chain AA catabolic pathway and tricarboxylic acid (TCA) cycle based on the different energy types (aminogenic versus lipogenic) used to synthesize milk components. We also hypothesized that the response of genes related to endoplasmic reticulum (ER) homeostasis via the unfolded protein response would reflect the increase in milk production stimulated by PT and FT. Fifty-six multiparous Holstein-Friesian dairy cows were fed a basal total mixed ration (34% grass silage, 33% corn silage, 5% grass hay, and 28% concentrate on a dry matter basis) for a 28-d control period. Experimental rations were then fed for 28 d, consisting of (1) low protein, low fat (LP/LF); (2) high protein, low fat (HP/LF); (3) low protein, high fat (LP/HF); or (4) high protein and high fat (HP/HF). To obtain the high-protein (HP) and high-fat (HF) diets, intake of the basal ration was restricted and supplemented isoenergetically (net energy basis) with 2.0 kg/d rumen-protected protein (soybean + rapeseed, 50:50 mixture on dry matter basis) and 0.68 kg/d hydrogenated palm fatty acids on a dry matter basis. RNA from milk fat samples collected on d 27 of each period underwent real-time quantitative PCR. Energy from protein increased expression of BCAT1 (branched-chain amino acid transferase 1) mRNA, but only at the LF level, and tended to decrease expression of mRNA encoding the main subunit of the branched-chain keto-acid dehydrogenase complex. mRNA expression of malic enzyme, a proposed channeling route for AA though the TCA cycle, was decreased by PT, but only at the LF level. Expression of genes associated with de novo fatty acid synthesis was not affected by PT or FT. Energy from fat had no independent effect on genes related to ER homeostasis. At the LF level, PT activated XBP1 (X-box binding protein 1) mRNA. At the HF level, PT increased mRNA expression of the gene encoding GADD34 (growth arrest and DNA damage-inducible 34). These findings support our hypothesis that mammary cells use aminogenic and lipogenic precursors differently for milk component production when dietary intervention alters AA and fatty acid supply. They also suggest that mammary cells respond to increased AA supply through mechanisms of ER homeostasis, dependent on the presence of FT.

Addition of glycerol to lactating cow diets stimulates dry matter intake and milk protein yield to a greater extent than addition of corn grain.

The objective of this study was to determine if the addition of glycerol to the diet of dairy cows would stimulate milk protein yield in the same manner as the addition of corn grain. Twelve multiparous lactating dairy cows at 81 ± 5 d in milk were subjected to 3 dietary treatments in a replicated 3 × 3 Latin square design for 28-d periods. The diets were a 70% forage diet considered the basal diet, the basal diet with 19% ground and high-moisture corn replacing forages, and the basal diet with 15% refined glycerol and 4% added protein supplements to be isocaloric and isonitrogenous with the corn diet. Cows were milked twice a day and samples were collected on the last 7 d of each period for compositional analysis. Within each period, blood samples were collected on d 26 and 27, and mammary tissue was collected by biopsy on d 28 for Western blot analysis. Dry matter intake increased from 23.7 kg/d on the basal diet to 25.8 kg/d on the corn diet and 27.2 kg/d on the glycerol diet. Dry matter intake tended to be higher with glycerol than corn. Milk production increased from 39.2 kg/d on the basal diet to 43.8 kg/d on the corn diet and 44.2 kg/d on the glycerol diet. However, milk yield did not differ between corn and glycerol diets. Milk lactose yields were higher on the corn and glycerol diets than the basal diet. Milk fat yield significantly decreased on the glycerol diet compared with the basal diet and tended to decrease in comparison with the corn diet. Mean milk fat globule size was reduced by glycerol feeding. Milk protein yield increased 197 g/d with addition of corn to the basal diet and 263 g/d with addition of glycerol, and the glycerol effect was larger than the corn effect. The dietary treatments had no effects on plasma glucose concentration, but plasma acetate levels decreased 27% on the glycerol diet. Amino acid concentrations were not affected by dietary treatments, except for branched-chain amino acids, which decreased 22% on the glycerol diet compared with the corn diet. The decreases in plasma acetate and branched-chain amino acid concentrations with glycerol and the larger effects of glycerol than corn on milk protein and fat yields suggest that glycerol is more glucogenic for cows than corn grain.

Exogenous essential amino acids stimulate an adaptive unfolded protein response in the mammary glands of lactating cows.

The phosphorylation of mammalian target of rapamycin complex 1 (mTORC1) components and integrated stress response networks in the mammary glands of lactating cows have not accounted for the stimulation of milk protein yield by chronic supplementation with AA or glucose. Faster milk protein synthesis could be a consequence of increased milk protein mRNA per cell, the number of ribosomes per cell, the secretory capacity of cells, or the mammary cell number. To investigate these 4 possibilities using a translational and transcriptional approach, we performed protein and gene expression analyses of mammary and longissimus dorsi tissue collected from lactating dairy cows after 5 d of abomasal infusion with saline or 844 or 1,126 g/d of an essential AA (EAA) mixture, with and without 1,000 g/d glucose. Infusion with EAA increased milk protein yield but did not affect the phosphorylation of mTORC1-related proteins in the mammary gland. In skeletal muscle, phosphorylation of 4EBP1 (eIF4E-binding protein 1) increased in response to both EAA and glucose, and phosphorylated S6K1 (70-kDa ribosomal protein S6 kinase) increased with glucose. In response to EAA, mammary mRNA expression of the marker genes for milk proteins, ribosome biogenesis, and cell proliferation were not upregulated. Instead, reciprocal regulation of 2 arms of the unfolded protein response occurred. Infusion of EAA for 5 d activated XBP1 (X-box binding protein 1) mRNA, encoding a transcription factor for endoplasmic reticulum biogenesis, and it decreased the mRNA expression of genes encoding pro-apoptotic protein CHOP (C/EBP homologous protein) and downstream GADD34 (growth arrest and DNA damage-inducible 34). These findings implicate non-stress-related, adaptive capabilities of the unfolded protein response in the long-term nutritional regulation of milk protein yield in lactating dairy cows.

Short communication: Parameters of abomasal emptying and glucose-insulin dynamics in Holstein-Friesian calves at 2 ages and 2 levels of milk replacer intake.

Elevated planes of nutrition in the preweaning period of dairy calf management can increase average daily gain, shorten age at puberty, and increase milk yield. In a previous study, 12 Holstein calves were fed 2 meals/d of 4 or 2 L milk replacer up to 7 wk of age. The objective of the current study was to estimate parameters of abomasal emptying and glucose-insulin dynamics in these calves by fitting a mechanistic model to postprandial appearances of plasma glucose, insulin, and the abomasal emptying marker acetaminophen measured at 4 and 7 wk of age. Higher intake of milk replacer resulted in longer bouts of abomasal emptying at a slower rate. Parameters of glucose and insulin dynamics were not affected by milk replacer intake. However, older calves had decreased insulin-stimulated glucose utilization indicating impaired insulin sensitivity, as well as increased pancreatic responsiveness. Neither of these effects were apparent from i.v. glucose tolerance tests on the calves and may have been related to postprandial gut hormone release. Effects of age on parameters of glucose-insulin dynamics were larger than effects of milk replacer intake. Conversely, effects of milk replacer intake on abomasal emptying were larger than effects of age.

Effect of plane of milk replacer intake and age on glucose and insulin kinetics and abomasal emptying in female Holstein Friesian dairy calves fed twice daily.

The objective of this study was to investigate how preweaning plane of milk replacer intake and age can affect insulin and glucose kinetics as well as abomasal emptying rate in dairy calves fed twice a day. A total of 12 female Holstein Friesian calves were blocked by cow parity, paired by colostrum origin, and were randomly assigned to a high plane of milk replacer intake (8 L/d, 1.2kg of milk replacer/d; n=6) or a low plane of nutrition (4 L/d, 0.6kg of milk replacer/d; n=6). All calves received 4 L of colostrum over 2 meals (1 and 6h after birth) and were then directly transferred to their assigned feeding plans until they were stepped-down from milk by 50% during wk 7 and weaned on wk 8. Milk replacer (24% crude protein, 18% crude fat) was fed at 150g/L twice daily (0700 and 1700h) and all calves had ad libitum access to pelleted calf starter, chopped wheat straw, and water. Jugular catheters were placed in all calves at 4, 7, and 10wk of age. Then, postprandial response to plasma glucose, insulin, and acetaminophen (supplied with the meal) were determined to measure abomasal emptying. The next day, a glucose tolerance test was conducted by infusing glucose via the jugular catheter. At 4 and 7wk of age, the rate constant (%/h) for abomasal emptying of the meal was lower in high calves (0.21±0.02 in wk 4; 0.27±0.02 in wk 7) compared with low (0.34±0.02 in wk 4; 0.47±0.02 in wk 7). The postprandial plasma insulin area under the curve over 420min was greater in high calves (18,443±7,329; low=5,834±739 µU/mL) compared with low. We found no differences in glucose tolerance test kinetics between the high and low dairy calves at 4, 7, or 10wk of age. The findings from this study suggest that feeding dairy calves an elevated plane of nutrition in 2 meals of milk replacer per day does not decrease insulin sensitivity.

Predicting extraction and uptake of arterial energy metabolites by the mammary glands of lactating cows when blood flow is perturbed.

Previous work shows that mammary uptake of milk precursors from blood can be affected by the rate of blood flow (F) to the glands. The purpose of the current work was to test the ability of compartmental and cylindrical capillary models to account for the variation in mammary extraction and net uptake of plasma metabolites produced by perturbation of mammary F. The data for model fitting were obtained from a previous experiment in which mammary arteriovenous differences of acetate + ß-hydroxybutyrate (2C), glucose, triacylglycerol (TAG), and long-chain fatty acids (LCFA) were measured in 4 cows before, during, and after intraarterial infusion of inhibitors of endothelial nitric oxide synthase and cyclooxygenase, which are 2 major systems of F control in the mammary glands. The 4 models tested were (1) constant extraction within each cow, (2) clearance from an extracellular compartment is a linear function of F with an intercept, (3) total capillary volume in a cylindrical representation is a linear function of F with an intercept, and (4) uptake from an extracellular compartment obeys Henri-Michaelis-Menten kinetics, where maximum velocity (Vmax) is a linear function of F with an intercept. According to prediction errors, model 4 fitted 2C extraction data best, accounting for 82% of the observed variation. The estimated Km (Henri-Michaelis-Menten constant) for venous 2C was 0.4 mM. For glucose clearance, a variant of model 2 with a positive effect of 2C uptake on clearance was identified as best, producing a coefficient of determination (R(2)) of 0.31. For TAG, model 2 with a positive effect of arterial TAG concentration on TAG clearance was best, with an R(2) of 0.22. For LCFA, model 2 with a positive effect of arterial LCFA on LCFA clearance was best, with an R(2) of 0.29. Models 2 and 3 fitted the extraction data with the same R(2)-values and prediction errors, so both compartmental and cylindrical approaches to describing the vascular bed were equally capable of describing the effect of F on mammary uptakes. A combined fit of all best-fit models to extraction data for all 4 metabolites at once explained 52, 42, 73, and 77% of variation in net uptakes of 2C, glucose, TAG, and LCFA, respectively. According to the fitted model, each 1 L/min increase in F increased the mammary volumes of distribution of 2C, glucose, TAG, and LCFA by 13, 14, 18, and 7%, respectively.