Increased liver blood flow (LBF) resulting from elevated feed intake in lactating dairy cows may increase steroid metabolism. Continuous infusion of bromosulphthalein (BSP; specifically metabolized in liver) was used to measure LBF. Similarly, progesterone (P4) and estradiol-17beta (E2) were administered by continuous infusion. Circulating concentrations at steady state were used to calculate the metabolic clearance rate (MCR) of BSP, P4, and E2. Experiment 1: Variation in LBF was determined in thee nonlactating and four lactating cows over 3 d at 3 to 5 h after feeding. Coefficients of variation ranged from 14 to 31% among cows within day and from 4 to 8% within cows across days. Experiment 2: Six nonlactating cows were used in a 3 x 3 Latin-square design with three feed regimens: no feed, 0.5 maintenance diet (M), and 1.5 M. Experiment 3: Eight lactating cows were used in a 4 x 4 Latin-square design with four feed regimens: no feed, 0.5 M, 1.5 M, and 2.2 M. In experiments 2 and 3, LBF and MCR of P4 increased immediately after feed consumption and increases persisted longer at higher intakes. The LBF reached a maximum at 2 h after feeding and MCR of P4 reached maximum at 3 h after feeding with a positive correlation (r = 0.92) between LBF and MCR for P4. Experiment 4: A crossover design was used to determine MCR of E2 in unfed or full-fed lactating dairy cows. The MCR of E2 increased immediately after feeding and stayed elevated throughout the 4.5-h infusion period. Thus, LBF and steroid metabolism were acutely elevated by feed consumption in lactating and nonlactating cows. Higher rates of LBF and steroid metabolism in lactating than in nonlactating cows may indicate chronic effects of higher feed intakes as well.

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This review integrates information on follicular and hormonal physiology and epidemiology into a novel physiological model for regulation of the ovulation rate in lactating dairy cows. First, the basic mechanisms that produce a single ovulation are examined. Follicular deviation is a critical new concept in our understanding of selection of a single dominant follicle. Follicular deviation is characterized by an abrupt deviation in the growth rates between the two largest follicles when the future dominant follicle reaches a diameter of 8.5+/-1.2 mm (mean and SD). The mechanisms involved in this selection process are not completely defined but appear to involve acquisition of LH receptors on granulosa cells of the dominant follicle, increased estradiol production by the dominant follicle, and inhibition of circulating FSH concentrations. Second, lactation number and milk production were found to be critical epidemiological factors associated with increased ovulation rate and twinning in dairy cattle. Finally, high steroid metabolism is proposed as the critical link between high milk production and double ovulation. It is proposed that high milk production increases steroid metabolism due to increased blood flow to the digestive tract and subsequently to the liver. The liver represents the primary site of steroid metabolism, and blood entering the liver is cleared of steroids. At the time of selection of the dominant follicle, the normal increase in circulating estradiol concentrations and subsequent depression in circulating FSH is blunted due to estradiol metabolism. Thus, FSH remains elevated for a time sufficient to allow follicles to undergo the physiological changes necessary to proceed to ovulation.

Some studies have reported improved reproductive performance with dietary fat supplementation. This study examined effects of fatty acids with different lengths, or desaturation, or both, on metabolism of estradiol (E2) and progesterone (P4) in bovine liver slice incubations (experiments 1 and 2) and in vivo (experiment 3). In experiment 1, effects of fatty acids C16:0 (palmitic acid), C16:1 (palmitoleic acid), C18:1 (oleic acid), and C18:3 (linolenic acid) were evaluated at 30, 100, and 300 microM on P4 and E2 metabolism in vitro. In experiment 2, stearic acid (C18:0) and C18:3 were evaluated in the same incubation conditions. In experiment 1, all of the fatty acids had some significant inhibitory effect on metabolism of P4, E2, or both (300 microM C16:0 on E2; 100 microM C16:1 on E2; 300 microM C16:1 on both P4 and E2; 300 microM C18:1 on P4; and 100 and 300 microM C18:3 on both P4 and E2). In experiment 2, C18:3 (100 and 300 microM) but not C18:0 decreased P4 and E2 metabolism. Overall, the most profound increase (approximately 60%) in half-life of P4 and E2 was observed with incubations of 300 microM C18:3 in both in vitro experiments. Based on these in vitro results, in experiment 3 linseed oil (rich in C18:3) was supplemented into the abomasum and acute effects on metabolism of E2 and P4 were evaluated. Cows (n=4) had endogenous E2 and P4 minimized (corpus luteum regressed, follicles aspirated) before receiving continuous intravenous infusion of E2 and P4 to analyze metabolic clearance rate for these hormones during abomasal infusion of saline (control) or 70 mL of linseed oil every 4h for 28h. Linseed oil infusion increased C18:3 in plasma by 46%; however, metabolic clearance rate for E2 and P4 were similar for control cows compared with linseed-treated cows. Thus, in vitro experiments indicated that E2 and P4 metabolism can be inhibited by high concentrations of C18:3. Nevertheless, in vivo, linseed oil did not acutely inhibit E2 and P4 metabolism, perhaps because insufficient C18:3 concentrations (increased to approximately 8 microM) were achieved. Further research is needed to determine the mechanism(s) of fatty acid inhibition of P4 and E2 metabolism and to discover practical methods to mimic this effect in vivo.

This study was designed to compare the efficiency of the Cryotop method and that of two methods that employ a micro volume air cooling (MVAC) device by analyzing the survival and development of bovine oocytes and blastocysts vitrified using each method. In experiment I, in vitro-matured (IVM) oocytes were vitrified using an MVAC device without direct contact with liquid nitrogen (LN2; MVAC group) or directly plunged into LN2 (MVAC in LN2 group). A third group of IVM oocytes was vitrified using a Cryotop device (Cryotop group). After warming, vitrified oocytes were fertilized in vitro. There were no significant differences in cleavage and blastocyst formation rates among the three vitrified groups, with the rates ranging from 53.1% to 56.6% and 20.0% to 25.5%, respectively; however, the rates were significantly lower (P < 0.05) than those of the fresh control group (89.3% and 43.3%, respectively) and the solution control group (87.3% and 42.0%, respectively). In experiment II, in vitro-produced (IVP) expanded blastocysts were vitrified using the MVAC, MVAC in LN2 and Cryotop methods, warmed and cultured for survival analysis and then compared with the solution control group. The rate of development of vitrified-warmed expanded blastocysts to the hatched blastocyst stage after 24 h of culture was lower in the MVAC in LN2 group than in the solution control group; however, after 48–72 h of culture, the rates did not significantly differ between the groups. These results indicate that the MVAC method without direct LN2 contact is as effective as the standard Cryotop method for vitrification of bovine IVM oocytes and IVP expanded blastocysts.

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Gender selection is important in livestock industries; for example, female calves are required in the dairy industry. Sex-sorted semen is commonly used for the production of calves of the desired gender. However, assessment of the sex ratio of the sorted semen is tedious and expensive. In this study, a rapid, cost effective and reliable method for determining the sex ratio was developed using a multiplex real-time polymerase chain reaction (PCR) assay. In this assay, the X and Y chromosome-specific markers, i.e., bovine proteolipid protein (PLP) gene and sex-determining region Y (SRY) were simultaneously quantified in a single tube. The multiplex real-time PCR assay was shown to have high amplification efficiencies (97% to 99%) comparable to the separated-tube simplex real-time PCR assay. The results obtained from both assays were not significantly different (p>0.05). The multiplex assay was validated using reference DNA of known X ratio (10%, 50%, and 90%) as templates. The measured %X in semen samples were the same within 95% confidence intervals as the expected values, i.e., >90% in X-sorted semen, <10% in Y-sorted semen and close to 50% in the unsorted semen. The multiplex real-time PCR assay as shown in this study can thus be used to assess purity of sex-sorted semen.