Effect of consuming endophyte-infected fescue seed on transcript abundance in the mammary gland of lactating and dry cows, as assessed by RNA sequencing

Abstract

Ergot alkaloids in endophyte-infected grasses inhibit prolactin secretion and reduce milk production in lactating cows. However, we previously showed that prepartum consumption of infected seed throughout the dry period did not inhibit subsequent milk production and prior exposure to bromocriptine (ergot peptide) actually increased production in the next lactation. To identify changes in the transcriptome and molecular pathways mediating the mammary gland's response to ergot alkaloids in the diet, RNA sequencing (RNAseq) was performed on mammary tissues obtained from 22 multiparous Holstein cows exposed to 1 of 3 treatments. Starting at 90 ± 4 d prepartum, cows were fed endophyte-free fescue seed (control; CON), endophyte-free fescue seed plus 3×/wk subcutaneous injections of bromocriptine (BROMO; 0.1 mg/kg of BW), or endophyte-infected fescue seed (INF) as 10% of the diet. Cows were dried off 60 ± 2 d prepartum. Mammary biopsies from 4 (BROMO, INF) or 5 (CON) cows/treatment at each of the 3 phases were obtained: 7 d before dry off during the initial lactation (L1), mid-dry period (D), and 10 d postpartum (L2). Although tissue from the same cow was preferentially used at 3 phases (L1, D, L2), tissue from additional cows were used to as necessary to provide RNA of sufficient quality. Individual samples were used to generate individual RNA-seq libraries. Normalized reads of the RNA-seq data were organized into technical and biological replicates before processing with the RSEM software package. Each lactation phase was processed separately and genes that differed between any of 3 treatments were identified. A large proportion of genes differentially expressed in at least 1 treatment (n = 866) were found to be similarly expressed in BROMO and INF treatments, but differentially expressed from CON (n = 575, total for 3 phases). Of genes differentially expressed compared with CON, 104 genes were common to the L1 and L2 phases. Consistent with the production findings, networks most affected by treatments in L1 and L2 included lipid metabolism, small molecule biochemistry, and molecular transport, whereas networks related more to developmental and cellular functions and maintenance were evident during D phase. Similar patterns of expression in BROMO and INF during late and early lactation suggest involvement of similar cell signaling pathways or mechanisms of action for BROMO and INF and the importance of prolactin messaging pathways.

Synthetic Alkaloid Treatment Influences the Intestinal Epithelium and Mesenteric Adipose Transcriptome in Holstein Steers

“…Based on the current dataset and previous work (15,20) in dairy cows BROMO impacted the intestinal transcriptome less than that observed in the transcriptome of mesenteric adipose and the mammary gland. We cannot eliminate the possibility that the intestinal transcriptome could be differentially influenced by luminal vs. systemic administration of ergot alkaloids, however, previous studies have shown that subcutaneous injections of bromocriptine alter intestinal transit in mice (27) and both intravenous and intraluminal bromocriptine stimulated small intestinal absorption of nutrients in multiple mammalian species (28).…”

“…Bromocriptine, a synthetic ergopeptine and dopamine receptor agonist, shares a high degree of structural homology with ergovaline and has also been shown to reduce circulating prolactin concentration in cattle (14,20). More recently we have shown that 90% of DEG (n = 866) in mammary tissue respond similarly for both fed fescue-derived alkaloids and BROMO treated cows (15); indicating that BROMO provides an adequate model for studying physiological changes in cattle grazing endophyte-infected tall fescue. In the current study we extend our finding concerning the effects of BROMO on the transcriptome to the intestinal epithelium and mesenteric adipose in growing ruminant animals.…”

“…The endophytic infection creates a source of ergot alkaloids of which ergovaline is found in the greatest concentrations ( 26 ). Consumption of these associated ergot alkaloids in sufficient amounts to cause fescue toxicosis in cattle results in a dramatic reduction in circulating prolactin concentration; a diagnostic tool used to confirm exposure to detrimental ergot alkaloids ( 5 , 12 – 15 , 20 ). Bromocriptine, a synthetic ergopeptine and dopamine receptor agonist, shares a high degree of structural homology with ergovaline and has also been shown to reduce circulating prolactin concentration in cattle ( 14 , 20 ).…”

“…Bromocriptine was reconstituted as working stock in 95% ethanol and diluted with saline so no more than 40% ethanol was given in a single injection. The concentration of BROMO was determined in a previous study where BROMO give at this dosage elicited the same gene expression response in 90% of differentially expressed genes in mammary tissue (n = 866) identified comparing the BROMO treatment response with the response observed in cows fed endophyte-infected fescue seed (15). Body weights were taken weekly to ensure feed was provided at 1.5 × maintenance and injection volumes calculated properly.…”

“…Bromocriptine is a dopamine receptor agonist that effectively inhibits prolactin secretion ( 14 ) similar to ergot alkaloids. Lactating cows fed endophyte-infected fescue or treated with bromocriptine had over 90% of differentially expressed genes (>850 genes) influenced in a similar manner when compared with control cows ( 15 ). Doses of bromocriptine have ranged from 15 to 80 mg/animal, and treatment frequencies have ranged from two times a day to three times per week ( 16 – 20 ) but provide a model to decrease prolactin and initiate fescue toxicosis symptoms in cattle with much greater control by not relying on animal intake.…”