G.E. Dahl

Invited review: Heat stress effects during late gestation on dry cows and their calves

In dairy cattle, late gestation is a critical period for fetal growth and physiological transition into the next lactation. Environmental factors, such as temperature and light, exert dramatic effects on the production, health, and well-being of animals during this period and after parturition. The aim of this review was to introduce effects of heat stress during late gestation on dairy cattle, and discuss the biological mechanisms that underlie the observed production and health responses in the dam and her fetus. Relative to cooled cows, cows that are heat stressed during late gestation have impaired mammary growth before parturition and decreased milk production in the subsequent lactation. In response to higher milk yield, cows cooled prepartum undergo a series of homeorhetic adaptations in early lactation to meet higher demand for milk synthesis compared with heat-stressed cows, but no direct effect of environmental heat stress on metabolism exists during the dry period. Prepartum cooling improves immune status of transition cows and evidence suggests that altered prolactin signaling in immune cells mediates the effects of heat stress on immune function. Late-gestation heat stress compromises placental development, which results in fetal hypoxia, malnutrition, and eventually fetal growth retardation. Maternal heat stress may also have carryover effects on the postnatal growth of offspring, but direct evidence is still lacking. Emerging evidence suggests that offspring from prepartum heat-stressed cows have compromised passive immunity and impaired cell-mediated immune function compared with those from cooled cows.

Evidence for short or ultrashort loop negative feedback of gonadotropin-releasing hormone secretion.

The present studies tested the hypothesis that either short or ultrashort loop negative feedback regulation of gonadotropin-releasing hormone (GnRH) secretion occurs in the ewe. As part of ongoing studies investigating the regulation of follicle-stimulating-hormone secretion, we obtained the unexpected result that a GnRH antagonist (Nal-Glu) may stimulate GnRH secretion. In that experiment, hypophyseal portal blood was collected from five short-term ovariectomized ewes at 5-min intervals for 6 h before and 6 h after intravenous injection of Nal-Glu (10 micrograms/kg body weight). An increase in GnRH pulse frequency in association with the blockade of luteinizing hormone (LH) release was evident in 3 of the 5 animals. To determine if an effect of Nal-Glu on episodic GnRH secretion would be more evident in an animal model in which low-frequency pulses of GnRH prevail, the study was repeated in six ewes in the midluteal phase of the estrous cycle and six ovariectomized ewes bearing estradiol and progesterone implants to suppress GnRH release (artificial luteal model). In luteal-phase ewes, administration of Nal-Glu was followed by an increase in GnRH pulse frequency, pulse size and the secretion of GnRH between pulses, and by a blockade of LH release. In ovariectomized ewes treated with estradiol and progesterone, Nal-Glu administration also stimulated GnRH and inhibited LH secretion. Our finding that the GnRH antagonist stimulated GnRH secretion is consistent with the hypothesis that endogenous GnRH may influence its own release via either a short or ultrashort loop feedback mechanism.

Effect of late-gestation maternal heat stress on growth and immune function of dairy calves

Heat stress during the dry period affects the cows mammary gland development, metabolism, and immunity during the transition period. However, the effect of late-gestation heat stress on calf performance and immune status is unknown. Our objective was to evaluate the effect of heat stress during the final ~45 d of gestation on growth and immune function of calves. Calves (17/treatment) were born to cows that were exposed to cooling (CL) or heat stress (HT) during the dry period. Only heifer calves (CL, n=12; HT, n=9) were used in measurements of growth and immune status after birth. Heifer calves were managed under identical conditions. All were fed 3.78 L of colostrum from their respective dams within 4 h of birth and were weaned at 2 mo of age (MOA). Body weight (BW) was obtained at weaning and then monthly until 7 MOA. Withers height (WH) was measured monthly from 3 to 7 MOA. Hematocrit and plasma total protein were assessed at birth, 1, 4, 7, 11, 14, 18, 21, 25, and 28 d of age. Total serum IgG was evaluated at 1, 4, 7, 11, 14, 18, 21, 25, and 28 d of age, and apparent efficiency of absorption was calculated. Peripheral blood mononuclear cells were isolated at 7, 28, 42, and 56 d of age, and proliferation rate was measured by (3)H-thymidine incorporation in vitro. Blood cortisol concentration was measured in the dams during the dry period and in calves in the preweaning period. Gestation length was 4d shorter for HT cows compared with CL cows. Calves from CL cows had greater BW than calves from HT cows at birth (42.5 vs. 36.5 kg). Compared with CL heifers, HT heifers had decreased weaning BW (78.5 vs. 65.9 kg) but similar BW (154.6 vs. 146.4 kg) and WH (104.8 vs. 103.4 cm) from 3 to 7 MOA. Compared with CL, heifers from HT cows had less total plasma protein (6.3 vs. 5.9 g/dL), total serum IgG (1,577.3 vs. 1,057.8 mg/dL), and apparent efficiency of absorption (33.6 vs. 19.2%), and tended to have decreased hematocrit (33 vs. 30%). Additionally, CL heifers had greater peripheral blood mononuclear cell proliferation relative to HT heifers (23.8 vs. 14.1 fold). Compared with CL, late-gestation HT did not affect the blood cortisol concentration of dams during the dry period or that of the calves in the preweaning period, but CL calves tended to have increased circulating cortisol at birth (7.6 vs. 5.7 µg/dL). We conclude that heat stress of the dam during the dry period compromises the fetal growth and immune function of offspring from birth through weaning.

Heat-stress abatement during the dry period: Does cooling improve transition into lactation?

Environmental factors, especially temperature and light exposure, influence the health and productivity of dairy cows during lactation, possibly via similar physiological mechanisms. For example, heat stress is a critical component of decreased milk yield during summer. However, less is known about the effect of heat stress during the dry period. The objective of this study was to evaluate the effects of heat stress prepartum under a controlled photoperiod on lactation performance and hepatic metabolic gene expression of periparturient multiparous Holstein cows (n = 16). Cows were dried off approximately 46 d before expected calving date and assigned to treatment randomly after blocking by mature equivalent milk production and parity. Treatments consisted of either heat stress (HT) or cooling (CL) with fans and sprinklers, both under a photoperiod of 14L:10D. Rectal temperature was measured twice daily during the dry period. After calving, cows were housed in a freestall barn with cooling devices, and milk yield was recorded daily up to 210 d in milk. Blood samples were taken from dry off until +42 d relative to calving for metabolites and from -2 until +2 d relative to calving for hormone analysis. Daily dry matter intake was measured from -35 to +42 d relative to calving. Liver biopsies were collected at dry off, -20, +2, and +20 d relative to calving for cows on HT (n = 5) and CL (n = 4) to measure mRNA expression of suppressors of cytokine signaling-2 (SOCS-2), insulin-like growth factor binding protein-5 (IGFBP-5), a key transcription factor in lipid biosynthesis (SREBP-1c), and enzymes of lipid metabolism (FASN, ACACA, and ACADVL) by real-time quantitative PCR. Heat stress increased rectal temperatures (39.2 vs. 38.8 degrees C), plasma prolactin concentrations at -1 (171 vs. 79 ng/mL) and 0 d (210 vs. 115 ng/mL) relative to calving, and decreased dry matter intake at 0 and +14 d relative to calving and 3.5% fat-corrected milk postpartum (26.1 vs. 35.4 kg/d) compared with CL cows. Relative to CL cows, hepatic mRNA expression of SOCS-2 and IGFBP-5 was downregulated in HT cows. Expression of ACADVL was upregulated in CL cows at d +2 but downregulated at d +20 relative to HT cows. Concentrations of C16:0 and cis C18:1 were greater in the milk and liver of CL cows compared with HT cows, which reflects greater lipid mobilization. These results suggest that heat-stress abatement in the dry period improves subsequent lactation, possibly via suppression of plasma prolactin surge around calving, SOCS-2 expression, and regulation of hepatic lipid metabolism.

Heat stress abatement during the dry period influences metabolic gene expression and improves immune status in the transition period of dairy cows

Heat stress (HT) and photoperiod affect milk production and immune status of dairy cows. The objective was to evaluate the effects of HT abatement prepartum under controlled photoperiod on hepatic metabolic gene expression and cellular immune function of periparturient Holstein cows (n=21). Cows were dried off 46 d before expected calving date and assigned to treatments by mature equivalent milk production. The treatments were 1) HT and 2) cooling (CL), both imposed during a photoperiod of 14L:10D. Rectal temperature was measured twice daily, whereas respiration rate was measured 3 times/wk at 1500 h during the entire dry period. After calving, cows were housed in a freestall barn with cooling, and milk yield was recorded daily up to 140 d in milk. Liver samples were taken at dry off, -20, 2, and 20 d relative to calving by biopsy. Under a similar schedule, neutrophil function was determined in blood of cows on HT (n=12) and CL (n=9). Blood samples were taken on -46, -32, -18, 0, 14, 28, and 42 d relative to calving for measurement of metabolites and were collected twice daily from -7 to 2 d relative to calving for prolactin (PRL) analysis. The HT cows had greater concentrations of PRL at 0 d relative to calving (150 vs. 93; SEM=11 ng/mL) and had higher afternoon rectal temperatures (39.4 vs. 39.0; SEM=0.04°C) and elevated respiration rates (78 vs. 56; SEM=2 breaths/min) during the prepartum period compared with CL cows. Relative to HT cows, CL cows had greater hepatic expression of PRL-R, SOCS-3, and CAV-1 mRNA. Neutrophil oxidative burst was greater in CL cows relative to HT cows at 2 d (61 vs. 42; SEM=6%) and at 20 d (62 vs. 49; SEM=5%) relative to calving, and phagocytosis was greater in CL cows at 20 d (47 vs. 33; SEM=4%) relative to calving compared with HT cows. Humoral response, as measured by IgG secretion against ovalbumin challenge, was greater for CL cows at -32 d (0.44 vs. 0.33; SEM=0.05 OD) and -21 d (0.60 vs. 0.50±0.04 OD) relative to calving compared with HT cows. These results suggest that HT abatement during the dry period improved innate and acquired immune status as measured by neutrophil function and immunoglobulin secretion against ovalbumin challenge, and altered hepatic gene expression related to PRL signaling in the periparturient period or subsequent lactation.

Effect of heat stress during the dry period on mammary gland development

Heat stress during the dry period negatively affects hepatic metabolism and cellular immune function during the transition period, and milk production in the subsequent lactation. However, the cellular mechanisms involved in the depressed mammary gland function remain unknown. The objective of the present study was to determine the effect of heat stress during the dry period on various indices of mammary gland development of multiparous cows. Cows were dried off approximately 46 d before expected calving and randomly assigned to 2 treatments, heat stress (HT, n=15) or cooling (CL, n=14), based on mature equivalent milk production. Cows in the CL treatment were provided with sprinklers and fans that came on when ambient temperatures reached 21.1°C, whereas HT cows were housed in the same barn without fans and sprinklers. After parturition, all cows were housed in a freestall barn with cooling. Rectal temperatures were measured twice daily (0730 and 1430 h) and respiration rates recorded at 1500 h on a Monday-Wednesday-Friday schedule from dry off to calving. Milk yield and composition were recorded daily up to 280 d in milk. Daily dry matter intake was measured from dry off to 42 d relative to calving. Mammary biopsies were collected at dry off, -20, 2, and 20 d relative to calving from a subset of cows (HT, n=7; CL, n=7). Labeling with Ki67 antigen and terminal deoxynucleotidyl transferase deoxyuridine triphosphate nick-end labeling were used to evaluate mammary cell proliferation and apoptosis, respectively. The average temperature-humidity index during the dry period was 76.6 and not different between treatments. Heat-stressed cows had higher rectal temperatures in the morning (38.8 vs. 38.6°C) and afternoon (39.4 vs. 39.0°C), greater respiration rates (78.4 vs. 45.6 breath/min), and decreased dry matter intake (8.9 vs. 10.6 kg/d) when dry compared with CL cows. Relative to HT cows, CL cows had greater milk production (28.9 vs. 33.9 kg/d), lower milk protein concentration (3.01 vs. 2.87%), and tended to have lower somatic cell score (3.35 vs. 2.94) through 280 d in milk. Heat stress during the dry period decreased mammary cell proliferation rate (1.0 vs. 3.3%) at -20 d relative to calving compared with CL cows. Mammary cell apoptosis was not affected by prepartum heat stress. We conclude that heat stress during the dry period compromises mammary gland development before parturition, which decreases milk yield in the next lactation.

Recombinant bovine soluble CD14 sensitizes the mammary gland to lipopolysaccharide.

Standard therapies including administration of potent antibiotics, aggressive fluid resuscitation and metabolic support have not been successful in relieving symptoms and reducing mortality associated with acute coliform mastitis. It is important to understand the pathophysiological response of the mammary gland to coliform infections when designing preventive or therapeutic regimens for controlling coliform mastitis. Our laboratory has previously shown that macrophages and polymorphonuclear neutrophils in milk express CD14 on their cell surface. In this study, we found that soluble CD14 (sCD14) is present in milk whey as a 46kDa protein reacted with anti-ovine CD14 antibody. Additional functional studies found that: (1) under serum-free condition, complexes of LPS-recombinant bovine soluble CD14 (rbosCD14) induced activation of mammary ductal epithelial cells (as measured by changes in interleukin-8 (IL-8) mRNA level by competitive RT-PCR) at low concentrations of LPS after 6 or 24h incubation (1-1000ng/ml), whereas LPS alone did not induce activation of mammary ductal epithelial cells at the same concentrations, and (2) intramammary injection of low concentrations of LPS did not increase concentration of leukocytes in milk. In contrast, LPS-rbosCD14 complex containing the same concentration of LPS increased the concentration of leukocytes in the injected mammary gland at 12 and 24h post-injection. These results indicate that rbosCD14 sensitizes mammary epithelial cells to low concentrations of LPS in vitro and in vivo. Endogenous sCD14 in milk may be important in initiating host responses to Gram-negative bacterial infections.

Sexual Differentiation of the Surge Mode of Gonadotropin Secretion: Prenatal Androgens Abolish the Gonadotropin-Releasing Hormone Surge in the Sheep

In sheep, the surge mode of gonadotropin secretion is sexually differentiated, i.e. the LH surge is present in the female, but not in the male. The present study tested the hypothesis that sexual differentiation of the LH surge mechanism reflects a sex difference in the pattern of GnRH, and that prenatal androgens abolish the surge mode of GnRH secretion. We monitored the pattern of GnRH secretion in pituitary portal blood after acute treatment with estradiol in gonadectomized postpubertal males (n=6), females (n=6), and androgenized females (exposed prenatally to testosterone from day 30–90 in gestation, n=7). Four capsules, each containing a 30‐mm column of estradiol were implanted s.c. into each lamb to produce high physiologic concentrations of the hormone. Beginning 7 h later, portal and peripheral blood samples were collected hourly for 48 h for measurement of GnRH and LH, respectively. All females exhibited a GnRH surge beginning 13.0±0.4 h after estradiol treatment; this was accompanied by an LH surge. By contrast, only one male produced a small surge in GnRH (1.7 pg/min) with a latency of 32 h; a corresponding increase in LH occurred in this male. Likewise, among the androgenized females, only one exhibited GnRH and LH surges which began at about 22 h after estradiol treatment. Some of the androgenized females had sporadic increases in GnRH which were of lower amplitude than in the control females, and were unaccompanied by rises in LH. These findings provide the first direct evidence that the sex difference in the surge mode of LH secretion results from the sexual differentiation of the pattern of GnRH release. The study also suggests that androgens during prenatal development abolish the GnRH surge and subsequently, the generation of the LH surge.

Heat stress abatement during the dry period influences prolactin signaling in lymphocytes

Heat stress perturbs prolactin (PRL) release and affects dairy cow lactational performance and immune cell function. We hypothesized that greater PRL concentration in plasma of heat-stressed cows relative to cooled cows would decrease expression of prolactin receptor (PRL-R) mRNA and increase mRNA expression of suppressors of cytokine signaling (SOCS) in lymphocytes, altering their cytokine production. To test this hypothesis, multiparous Holstein cows were dried off 46 d before their expected calving date and assigned randomly to heat stress (HT; n=9) or cooling (CL; n=7) during the entire dry period. A second study was conducted the following year with an additional 21 cows (12 HT; 9 CL). Lymphocytes were isolated from cows at -46, -20, +2, and +20 d relative to expected calving date and mRNA expression of PRL-R, SOCS-1, SOCS-2, SOCS-3, cytokine-inducible SH2-containing protein (CIS), and heat shock protein 70 KDa A5 (HSPA5), and housekeeping genes hydroxymethylbilane synthase (HMBS), ATP synthase, H+ transporting mitochondrial F1 complex, beta subunit (ATP5B), and ribosomal protein S9 (RPS9) was analyzed by quantitative real-time reverse transcriptase polymerase chain reaction (RT-PCR). Cows exposed to HT had greater PRL concentration in plasma compared with CL cows. Measurement of lymphocyte proliferation indicated that lymphocytes of CL cows proliferated more than those from HT cows and exressed more PRL-R mRNA and less SOCS-1 and SOCS-3 mRNA relative to HT cows. Further, lymphocytes from CL cows produced more tumor necrosis factor-alpha (TNF-alpha) than those from HT cows. These results suggest that changes in PRL-signaling pathway genes during heat stress are associated with differential cytokine secretion by lymphocytes and may regulate lymphocyte proliferation in dairy cows.

Effect of colostral volume, interval between calving and first milking, and photoperiod on colostral IgG concentrations in dairy cows

OBJECTIVE To identify cow and management factors associated with colostral IgG concentration in dairy cows. DESIGN Prospective observational study. ANIMALS 81 multiparous Holstein-Friesian cows from a single herd. PROCEDURES Serum was obtained at the start of the nonlactating period, and cows were assigned to 1 of 4 photoperiod groups: natural day length (n = 22 cows), long days (16 h of light/d [21]) or short days (8 h of light/d [20]) for the entire nonlactating period, or natural day length followed by short days for the last 21 days of the nonlactating period (18). Serum and colostrum were collected at the first milking after calving. Regression analysis was used to investigate associations between colostral IgG concentration and the interval between calving and first milking, colostral volume, photoperiod, length of the nonlactating period, and season of calving. RESULTS Colostral IgG concentration decreased by 3.7% during each subsequent hour after calving because of postparturient secretion by the mammary glands. The interval between calving and first milking and the colostral volume were significantly and negatively associated with colostral IgG concentration, with the former effect predominating. Photoperiod had no effect on colostral IgG concentration or volume. Serum protein concentration at calving correlated poorly with colostral IgG concentration. CONCLUSIONS AND CLINICAL RELEVANCE Dairy producers should harvest colostrum as soon as possible after calving to optimize transfer of passive immunity in neonatal calves. Photoperiod can be manipulated without adversely affecting colostral IgG concentration.