Milk protein yield responses to changes in the profile of essential amino acids absorbed by the gastrointestinal tract or circulating in blood plasma do not follow the classic limiting amino acid response, in part because of an ability of the mammary glands to modify their blood flow rate and net clearance of amino acids out of plasma. The hypothesis that mammary blood flow is locally regulated to maintain ATP balance accounts for observed changes in flow due to postruminal glucose, insulin, and essential amino acid (EAA) infusions. An additional hypothesis that net mammary uptakes of metabolites from blood are affected by perturbations in their respective arterial concentrations and the rate of mammary blood flow also appears to hold for the energy metabolites glucose, acetate, β-hydroxybutyrate, and fatty acids. However, net EAA uptakes by the mammary glands are poorly predicted by models considering arterial concentrations and blood flow rates only. Evidence points to intramammary protein synthesis and secretion as the determinant of net EAA uptake. The intracellular signaling network anchored by the mechanistic target of rapamycin complex 1 stands as an excellent candidate to explain nutritional effects on milk protein synthesis because it integrates information on physiological and nutritional state to affect protein synthesis and cell metabolism, growth, proliferation, and differentiation in many cell types. In mammary cells in vitro and in vivo, the mechanistic target of rapamycin complex 1, integrated stress response, and glycogen synthase kinase-3 networks that contribute to regulation of initiation of mRNA translation are responsive to acute changes in nutrient supply and EAA profile. However, after several days of postruminal infusion of balanced and imbalanced EAA profiles, these signaling networks do not appear to continue to account for changes in milk protein yields. Gene expression evidence suggests that regulation of components of the unfolded protein response that control biogenesis of the endoplasmic reticulum and differentiation of a secretory phenotype may contribute to effects of nutrition on milk protein yield. Connections between early signaling events and their long-term consequences should be sought.

The purposes of this study were 1) to determine the compartmentation of body water in horses by using indicator dilution techniques and 2) to simultaneously measure bioelectrical impedance to current flow at impulse current frequencies of 5 and 200 kHz to formulate predictive equations that could be used to estimate total body water (TBW), extracellular fluid volume (ECFV), and intracellular fluid volume (ICFV). Eight horses and ponies weighing from 214 to 636 kg had catheters placed into the left and right jugular veins. Deuterium oxide, sodium thiocyanate, and Evans blue were infused for the measurement of TBW, ECFV, and plasma volume (PV), respectively. Bioelectrical impedance was measured by using a tetrapolar electrode configuration, with electrode pairs secured above the knee and hock. Measured TBW, ECFV, and PV were 0.677 +/- 0.022, 0.253 +/- 0.006, and 0.040 +/- 0.002 l/kg body mass, respectively. Strong linear correlations were determined among measured variables that allowed for the prediction of TBW, ECFV, ICFV, and PV from measures of horse length or height and impedance. It is concluded that bioelectrical impedance analysis (BIA) can be used to improve the predictive accuracy of noninvasive estimates of ECFV and PV in euhydrated horses at rest.

To test which, if any, of the major milk precursors can elicit a rapid change in the rate of mammary blood flow (MBF) and to define the time course and magnitude of such changes, 4 lactating cows were infused with glucose, amino acids, or triacylglycerol into the external iliac artery feeding one udder half while iliac plasma flow (IPF) was monitored continuously by dye dilution. Adenosine and saline were infused as positive and negative controls, respectively, and insulin was infused to characterize the response to a centrally produced anabolic hormone. To test the roles of cyclooxygenase, NO synthase and ATP-sensitive K (K ATP ) channels in nutrient-mediated changes in blood flow, their respective inhibitors-indomethacin, N ω -nitro-Larginine methyl ester hydrochloride (L-NAME), and glibenclamide-were infused simultaneously with glucose. Each day, 1 infusate was given twice to each cow, over a 20-min period each time, separated by a 20-min washout period. In addition, each treatment protocol was administered on 2 separate days. A 73% increase in IPF during adenosine infusion showed that the mammary vasodilatory response was quadratic in time, with most changes occurring in the first 5 min. Glucose infusion decreased IPF by 9% in a quadratic manner, most rapidly in the first 5 min, indicating that a feedback mechanism of local blood flow control, likely through adenosine release, was operative in the mammary vasculature. Amino acid infusion increased IPF 9% in a linear manner, suggesting that mammary ATP utilization was stimulated more than ATP production. This could reflect a stimulation of protein synthesis. Triacylglycerol only tended to decrease IPF and insulin did not affect IPF. A lack of IPF response to glibenclamide indicates that K ATP channels are not involved in MBF regulation. Indomethacin and L-NAME both depressed IPF. In the presence of indomethacin, glucose infusion caused a quadratic 9% increase in IPF. Indomethacin is an inhibitor of mitochondrial function, so the glucoseinduced increase in IPF was interpreted as feedback on mammary adenosine release from an anabolic response to glucose. Because NO synthase was not inhibited during indomethacin infusion, the feedback system is postulated to act through endothelial NO synthase. In the presence of L-NAME, glucose infusion had no effect on IPF, indicating that endothelial cyclooxygenase is not involved in glucose-induced changes in MBF.

To test the effect of mammary blood flow on net uptakes of milk precursors by the mammary glands, inhibitors of nitric oxide synthase (NOS) and cyclooxygenase (COX) were infused into the mammary circulation of 4 lactating cows. Inhibitors were infused in a 4×4 Latin square design, where treatments were infusion for 1 h of saline, NOS inhibitor (Nω-nitro-l-arginine methyl ester hydrochloride), COX inhibitor (indomethacin), or both NOS + COX inhibitors into one external iliac artery. Para-aminohippuric acid was also infused to allow for estimation of iliac plasma flow (IPF), of which approximately 80% flows to the mammary glands. Blood samples were collected before, during, and after inhibitor infusion from the contralateral external iliac artery and ipsilateral mammary vein. Inhibition of COX and NOS each produced a decrease in IPF, although the NOS effect was smaller and IPF continued to be depressed throughout the recovery period. The combination of COX and NOS inhibition produced a 50% depression in IPF and there was no carryover into the recovery period. Treatments that depressed IPF also increased arterial concentrations of acetate, β-hydroxybutyrate (BHBA), and glucose. Similarly, arteriovenous differences of acetate, BHBA, and glucose were all increased during IPF depression. To correct for a potential effect of arterial concentration, arteriovenous differences were normalized to arterial concentration, producing an extraction percentage. Inhibition of COX increased glucose extraction and tended to increase acetate and BHBA extraction. Dual inhibition only increased BHBA extraction and had no effect on mammary extraction of other metabolites. These extractions did not increase because clearances of glucose and TAG decreased as IPF decreased, and clearances of acetate and BHBA tended to decrease. Net uptake of TAG was depressed by dual NOS/COX inhibition, whereas uptakes of acetate, BHBA, and glucose were not affected by any of the treatments. To separate effects of flow from effects of arterial concentration, uptakes were regressed against IPF and arterial concentration simultaneously. According to the slopes of the regressions, a 10% decrease in IPF from the mean observed during saline infusion resulted in 3.8, 7.3, and 10.4% decreases in uptakes of acetate, glucose, and triacylglycerol, respectively. These findings indicate that mammary blood flow affects milk precursor uptake, and that clearance should not be assumed constant to predict mammary uptakes of milk precursors in situations where blood flow is changing.

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