Intravenous infusion of 5-hydroxytryptophan to mid-lactation Holstein cows transiently affects milk production and circulating amino acid concentrations.

In dairy cows, the lactating mammary glands synthesize serotonin, which acts in an autocrine-paracrine manner in the glands and is secreted into the periphery. Serotonin signaling during lactation modulates nutrient metabolism in peripheral tissues such as adipose and liver. We hypothesized that the elevation of circulating serotonin during lactation would increase nutrient partitioning to the mammary glands, thereby promoting milk production. Our objective was to elevate circulating serotonin via intravenous infusion of the serotonin precursor 5-hydroxytryptophan (5-HTP) to determine its effects on mammary supply and extraction efficiency of AA, and milk components production. Twenty-two multiparous mid-lactation Holstein cows were intravenously infused with 5-HTP (1 mg/kg body weight) or saline, in a crossover design with two 21-d periods. Treatments were infused via jugular catheters for 1 h/d, on d 1 to 3, 8 to 10, and 15 to 17 of each period, to maintain consistent elevation of peripheral serotonin throughout the period. Milk and blood samples were collected in the last 96 h of each period. Whole-blood serotonin concentration was elevated above saline control for 96 h after the last 5-HTP infusion. Dry matter intake was decreased for cows receiving 5-HTP, and on average they lost body weight over the 21-d period, in contrast to saline cows who gained body weight. Milk production and milk protein yield were lower in cows receiving 5-HTP during the 3 infusion days, but both recovered to saline cow yields in the days after. Although milk fat yield exhibited a day-by-treatment interaction, no significant difference occurred on any given day. Milk urea nitrogen concentration was lower in 5-HTP cows on the days following the end of infusions, but not different from saline cows on infusion days. Meanwhile, plasma urea nitrogen was not affected by 5-HTP infusion. Circulating concentrations of AA were overall transiently decreased by 5-HTP, with concentrations mostly returning to baseline within 7 h after the end of 5-HTP infusion. Mammary extraction efficiency of AA was unaffected by 5-HTP infusion. Overall, both lactation performance and circulating AA were transiently reduced in cows infused with 5-HTP, despite sustained elevation of circulating serotonin concentration.

Mechanisms by which feeding synthetic zeolite A and dietary cation-anion difference diets affect mineral metabolism in multiparous Holstein cows: Part I.

The periparturient period is characterized by the increased demand for calcium (Ca) in dairy cows. This has resulted in the use of several different prepartal nutritional strategies to prevent hypocalcemia postpartum. The objective of our study was to determine the effects of feeding synthetic zeolite A (XZ), a diet with negative dietary cation-anion difference (-DCAD), or a positive-DCAD diet (CON) during the close-up period on peripartal mineral dynamics and hormones involved in calcium metabolism. To this end, 121 multiparous Holstein cows, blocked by lactation number and expected due date, were enrolled at 254 d of gestation and randomly assigned to 1 of 3 prepartum diets: CON (+190 mEq/kg; n = 40), -DCAD (-65 mEq/kg; n = 41), or a diet supplemented with sodium aluminum silicate (XZ; +278 mEq/kg, fed at 3.3% DM, targeting 500 g/d; n = 40; Protekta Inc.). Blood, urine, and saliva samples were collected from enrollment until parturition, with data analyzed and presented beginning 14 d before parturition (d -14) until parturition (d 0), and on d 1, 2, 3, 6, 9, 12, 15, 18, 21, 35, and 49 postpartum, to assess mineral and hormone dynamics. Total fecal collections were performed in a subset of 8 cows per treatment group to assess fecal mineral loss. Data were analyzed as a randomized complete block design in SAS. Cows fed XZ and -DCAD had higher blood Ca concentrations compared with CON-fed cows, with XZ-fed cows exhibiting the highest blood Ca concentrations pre- and postpartum. Cows fed XZ had decreased blood and salivary phosphorus (P), increased fecal water-extractable phosphate, and the highest blood calcium concentrations pre- and postpartum. Parathyroid hormone was unaffected by diet but was increased at parturition in all treatments. Serotonin concentrations were increased in -DCAD and XZ cows compared with CON during the prepartum period. Our data indicate that the XZ group's improvement in blood Ca concentrations pre- and postpartum is most likely regulated by a dietary P restriction. Taken together, these data suggest that XZ and -DCAD diets improve postpartum calcium metabolism; however, they appear to work through different mechanisms.

Serotonin alters the response to a glucose challenge in lactating cows.

Energy partitioning in lactating cows affects milk production, feed efficiency, and body reserves, with the latter having health implications for the transition into the following lactation. One molecule likely involved in the regulation of energy partitioning is serotonin. The objective of this experiment was to explore how increasing circulating serotonin, by intravenous infusion of the serotonin precursor 5-hydroxytryptophan (5-HTP), affects metabolic responses to a glucose challenge in midlactation cows as a means to manipulate energy partitioning. We intravenously infused Holstein cows with 5-HTP (1 mg/kg bodyweight dissolved in saline, n = 11) or saline alone as control (n = 9) over 1 h/day for 3 days. Cows were fasted overnight on day 2. On day 3, fasted cows were given an intravenous bolus of glucose (0.092 g/kg bodyweight). Blood samples were collected for the following 120 min for metabolic and hormonal analysis. Infusion of 5-HTP elevated circulating concentrations of serotonin and free fatty acids, reduced the concentration of insulin and amino acids, and did not affect the concentration of glucose and glucagon before the glucose challenge. Surrogate insulin sensitivity indices indicated improved insulin sensitivity in 5-HTP cows, but due to the unique metabolism of lactating ruminants, these index changes may instead reflect effects in insulin-independent glucose disposal, like milk synthesis. Challenging 5-HTP-treated cows with a glucose bolus reduced the insulin spike and blunted the decrease in free fatty acids, compared to saline cows, without changing glucose dynamics. Overall, these results suggest that serotonin stimulates insulin-independent glucose disposal, requiring less insulin to maintain normoglycemia.

The antidepressant fluoxetine (Prozac®) modulates serotonin signaling to alter maternal peripartum calcium homeostasis.

Antidepressant use is two-fold greater in women compared to men; however, most studies have been performed in male subjects. We aimed to understand the impact of selective serotonin reuptake inhibitors (SSRI, most used antidepressants) on calcium homeostasis and steroid metabolism during the peripartum period. Pregnant sheep (n = 10/group) were treated with vehicle or fluoxetine (most common SSRI) during the last month of gestation. Fluoxetine treatment decreased circulating calcium prior to parturition (8.7 ± 0.1 mg/dL vs 8.2 ± 0.1 mg/dL; P = 0.07). In the control group, total calcium decreased after parturition corresponding to the onset of lactogenesis followed by increase in calcium by day 2 postpartum. Interestingly, this normal transient decrease in circulating calcium was absent in fluoxetine-treated ewes. The steroids cortisol and progesterone were not altered by fluoxetine treatment whereas estradiol was decreased after the onset of treatment (12.4 ± 1.3 vs 9.1 ± 1.2 pg/mL, P = 0.05) and prior to parturition (38.1 ± 8.1 vs 22.3 ± 4.2 pg/mL, P = 0.03). Our hypothesis was supported that fluoxetine treatment alters circulating concentrations of calcium in the peripartum period; however, we surprisingly observed a decrease in estradiol concentrations contrary to reports in in vitro studies.

Fluoxetine-induced perinatal morbidity in a sheep model.

Selective serotonin reuptake inhibitors (SSRI) are the most common antidepressants used by pregnant women. However, adverse pregnancy outcomes have been described in women taking SSRI during pregnancy-placental lesions, premature birth, poor neonatal adaptation. We aimed to investigate the effects of fluoxetine (Prozac® most commonly used SSRI) treatment during the last month of gestation on pregnancy complications, placental and neonatal health in a non-depressed sheep model. On day 119 ± 1 postbreeding (experimental day 0; E0) of a 151-day expected gestation, Hampshire ewes were randomly assigned to receive fluoxetine (n = 9 ewes, 15 lambs; daily intravenously treatment with 10 mg/kg on E0 and E1 and 5 mg/kg daily thereafter until parturition) or to a control group (n = 10; 14 lambs; vehicle only). Blood samples from ewes were collected throughout the experimental period and postpartum; blood from lambs were collected postpartum. Analysis of variance was used for statistical analysis. Fluoxetine treatment reduced placentome growth during the last month of pregnancy. Gestation length was decreased by 4.5 days in fluoxetine-treated ewes. Birthweight was reduced in lambs exposed to fluoxetine in utero; weights remained decreased until postnatal day 3. Placentome diameter by birthweight ratio was not different between groups suggesting that the decreased placentome diameter was accompanied by decreased lamb birthweight. During the first week postnatal, lambs exposed to fluoxetine in utero had decreased blood pH and decreased total carbon dioxide, bicarbonate, and base excess and increased lactate (days 3-6), collectively indicative of metabolic acidemia. Additionally, ionized calcium was decreased between postnatal days 0 to 4 in lambs exposed to fluoxetine in utero. Using a non-depressed animal model clearly defines a role for SSRI on the occurrence of perinatal complications and neonatal morbidity. The decreased placentome diameter, shortened gestation, decreased birthweight, decreased calcium levels, and neonatal acidemia suggest the occurrence of intrauterine growth restriction. The persistence of neonatal acidemia for several days postpartum suggests poor neonatal adaptation to extrauterine environment.

Effects of methionine, leucine, and insulin on circulating concentrations and mammary extraction of energy substrates and amino acids in lactating dairy cows.

The aim of this experiment was to test whether insulin potentiates the effects of two abomasally infused amino acids (AA), leucine and methionine (LM), on mammary extraction efficiency of energetic and nitrogenous nutrients. Six lactating Holstein cows (155 ± 9 DIM) were ruminally-cannulated and had the right carotid artery subcutaneously transposed. Cows were fed a 20% metabolizable protein-restricted diet and abomasally infused with water (8 L/d) or AA (Met 26 g/d, Leu 70 g/d) for 8 h/d, for 7 days. On the last day of each period, cows were intravenously infused with saline (0.9% NaCl, 110 mL/h) or subjected to 8 h hyperinsulinemic clamp (IC) alongside abomasal infusions. For IC, insulin was infused at 1 µg/kg/h. Normoglycemia was maintained by varying glucose (50% w/v in water) infusion rate based on coccygeal vein glucose concentration. Carotid arterial and subcutaneous abdominal (mammary) vein blood samples were collected at 0, 1, 2, 4, and 6 h from the start of infusions. Milk weights and samples for baseline measurements of production were taken on day 5 PM, day 6 AM and PM, and day 7 AM of the experimental period. A final milk weight and sample was taken immediately after abomasal and intravenous infusions on day 7 PM for assessing the interaction between insulin and the infused AA. The experiment had an incompletely replicated Latin square design with a 2 × 2 factorial arrangement of treatments (abomasal and intravenous infusion). Baseline milk production when cows were only receiving abomasal infusions was largely unaffected by LM, but milk protein yield tended to be decreased. On day 7, LM tended to positively increase milk fat and de novo fatty acid content, and IC tended to decrease milk protein content. Both milk urea nitrogen and plasma urea nitrogen were decreased by IC. Circulating AA concentrations in plasma were decreased by both LM and IC, but mammary extraction efficiency was affected by neither. Infusion of LM had no effect on any energy metabolite analyzed. Circulating non-esterified fatty acid concentration was decreased by IC, with no effect on mammary extraction efficiency. Mammary extraction efficiency of both acetate and ß-hydroxybutyrate were decreased by IC. Overall, while both circulating concentrations of energy metabolites and amino acids were decreased in response to treatments, this was not due to improved mammary extraction efficiency.

Elevated serotonin coordinates mammary metabolism in dairy cows.

Serotonin plays a diverse role in maternal and mammary metabolism. Recent research in the dairy cow has shown a relationship between serotonin and calcium, with increased serotonin concentrations improving calcium homeostasis in the peri-partum dairy cow. Therefore, the objective was to elucidate how administration of 5-hydroxy-l-tryptophan (5-HTP), the immediate precursor to serotonin, altered serotonin and calcium metabolism in lactating dairy cows. Twelve mid-late lactation multiparous cows were blocked by parity, production and days in milk and allocated to a daily intravenous infusion of (i) 1.5 mg/kg of 5-HTP (n = 6) or (ii) saline (n = 6) for 3 consecutive days. Milk samples were collected daily. Blood samples were collected before and after each infusion with mammary biopsies and blood samples collected at 48, 56, and 72 h relative to termination of first infusion. Infusion of 5-HTP increased (p = 0.001) circulating serotonin concentrations and decreased blood calcium via a transient hypocalcemia immediately after each infusion (p = 0.02). Treatment with 5-HTP increased milk calcium concentrations (p = 0.02) and calcium release-activated channel protein 1 (ORAI1) mRNA at 56 h and protein at 48 h relative to termination of first infusion (p = 0.008 and p = 0.09, respectively). Fifty-six hours from termination of the first infusion mRNA of parathyroid hormone-related protein and mammary serotonin content were increased relative to control (p = 0.03 and p = 0.05, respectively). These findings demonstrate the ability of 5-HTP infusion to increase circulating serotonin concentrations and alter endocrine and mammary autocrine/paracrine calcium and serotonin metabolism in the lactating dairy cow.

The effects of incomplete milking and increased milking frequency on milk production rate and milk composition1.

Increased milking frequency and incomplete milking have differential effects on milk yield and mammary gland physiology that are important for optimization of milking practices in dairy herds. The objectives of this experiment were to determine the effects of increased milking frequency and incomplete milking on milk production rate (MPR) and milk composition and to determine if milking 3 times daily (3×) could rescue the negative production effects of incomplete milking. Twenty-two multiparous cows were enrolled onto this experiment beginning at 5 days in milk (DIM) and continuing through 47 DIM. A split-plot design was used to randomize the 2 treatments, which were milking frequency and incomplete milking. Eleven cows were randomly assigned to be milked 2 times (2×) daily and 11 cows were randomly assigned to be milked 3×. Within each cow, a contralateral half-udder was randomly assigned to be incompletely milked (30% milk remaining in the gland; IM), and the other half-udder was randomly assigned to be milked completely (CM). Quarter-level milk yields were recorded at each milking session. Milk samples from all quarters were collected twice weekly at the beginning of the morning milking for analysis. Cows milked 2× tended to have reduced MPR compared with 3× milked cows (1.81 ± 0.06 vs. 1.97 ± 0.06 kg milk/h; P = 0.06). Half-udders that were CM and IM produced 1.09 ± 0.03 and 0.80 ± 0.03 kg milk/h, respectively. There was an interaction between incomplete milking treatment and week of lactation (P = 0.04). No interaction was detected between milking frequency and incomplete milking for MPR or milk components. Cows milked 3× had increased milk fat percent (1.93 ± 0.09% vs. 1.65 ± 0.09%, P = 0.047), decreased milk lactose percent (4.80 ± 0.04% vs. 4.93 ± 0.04%, P = 0.04), and exhibited no differences in milk protein percent or milk somatic cell count (SCC) compared with cows milked 2×. Half-udders that were IM had increased milk fat percent (2.15 ± 0.07% vs. 1.43 ± 0.07%, P < 0.0001), decreased lactose percent (4.75 ± 0.03% vs. 4.99 ± 0.03%, P < 0.0001), increased milk log10SCC (4.22 ± 0.05 vs. 4.41 ± 0.05, P = 0.0004), and no differences in milk protein percent compared with CM half-udders. These results indicate that a 3× milking frequency in IM half-udders was not able to improve milk production compared with IM half-udders milked 2×. Our results indicate that 30% milk remaining in the gland had an irreversible impact on milk yield as increased milking frequency was not able to reverse the milk yield lost.