Reproductive studies of Brahman cattle V. The effect of various dose levels of estradiol-17β upon serum luteinizing hormone in ovariectomized Brahman cows

Twelve mature chronically-ovariectomized Brahman cows were randomly assigned to receive three of six different dosages of estradiol-17_b (E2) at three different time periods such that at the termination of the trial six animals received each E2 dosage. The E2 was suspended in 0, 1, 2.5, 5, 10 and 20 milligrams. A two week period was maintained between injections. The cows were bled via coccygeal vessel puncture immediately before E2 injection, every 2 hr from 0 to 18 hr, every hr from 18 to 42 hr and every 2 hr from 42 to 48 hr postinjection. Blood was processed to yield serum and stored at ?20 Celsius. Serum luteinizing hormone (LH) was quantitated by validated radioimmunoassay. An LH surge was defined as a sustained elevation of LH at least two standard deviations above the level of LH prior to the rise and was observed in $\text{0}\text{6}$, $\text{3}\text{6}$, $\text{5}\text{6}$, $\text{5}\text{6}$, $\text{5}\text{6}$, and $\text{6}\text{6}$ cows administered 0, 1, 2.5, 5, 10, and 20 mg of E2, respectively. All animals injected with E2 responded with a significant initial decrease (independent of E2 dosage) in LH that persisted from 2 through 12 hr post E2 injection. No significant decrease in LH levels was recorded in animals injected with the corn oil vehicle. The time to the LH surge differed (P<.05) between 1 mg E2 (10 hr) vs 20 mg E2 (19.5 hr), 1 mg E2 vs 10 mg E2 (16.2 hr), and 2.5 mg E2 (12.4 hr) vs 20 mg estradiol-17_β. Luteinizing hormone concentrations at the onset of the surge did not differ (P>.10) between E2 dosages. The elapsed time from E2 injection to the peak LH value differed (P<.05) between 1 mg E2 (20.3 hr) vs 20 mg E2 (26.8 hr), and 2.5 mg E2 (21.2 hr) vs 20 mg estradiol-17_β. The peak LH value, the area under the LH curve and the duration of the LH surge did not differ (P>.10) with E2 dosage. The time to the end of the LH surge differed (P<.05) between 1 mg E2 (25.3 hr) vs 2.5 mg E2 (31.6 hr), 1 mg E2 vs 5 mg (34.4 hr), 1 mg E2 vs 10 mg E2 (34.8 hr), 1 mg E2 vs 20 mg E2 (37.3 hr), and 2.5 mg E2 vs 20 mg estradiol-17_β. Luteinizing hormone values at the termination of the surge did not differ (P>.10) between dosages nor did the LH values at the termination of the surge differ (P>.10) from LH concentrations observed at the onset of the LH surge.     

Effect of variable suckling intensity and estrogen administration upon serum luteinizing hormone in Brahman cows

Ten mature Brahman cows were randomly allotted within calving intervals to either a suckled (S) or nonsuckled (NS) treatment group. All cows received a 20 mg intramuscular injection of estradiol-17beta (E2), suspended in 2 ml of corn oil, to determine the effect of suckling on the estrogen induced LH surge. Starting on day 21 postpartum the S cows were suckled at six hour intervals for 24 hours, at which time they were challenged with a 20 mg E2 injection. The suckling regimen was continued for 48 hours postinjection. The NS cows were separated from their calves on day 21 postpartum and received no suckling stimulus for 72 hours. At 24 hours after calf separation, the NS cows were challenged with a 20 mg E2 injection. Blood samples were removed at two hour intervals beginning 10 hours post E2 injection until 36 hours postinjection, at which time blood samples were removed at four hour intervals until 48 hours postinjection. Blood samples were processed to yield serum and assayed for luteinizing hormone (LH) via radioimmunoassay. The injection of a 20 mg dose of E2 induced an LH surge in all cows. The NS cows were found to exhibit a longer (P<.05) duration of the estrogen induced LH surge than the S cows, 15.6 +/- .98 and 12.4 +/- .75 hours, respectively. The timing parameters (time to start of LH surge, time to peak LH value and time to end of surge) and LH concentration parameters (LH concentration at start of LH surge, peak value of LH surge and LH concentration at end of LH surge) were not different between suckling regimens. No blockage of the LH response to estrogen challenge was found on day 22 postpartum. Suckling did depress the duration of the LH surge indicating some blockage due to suckling stimuli.     

Amniotic fluid and plasma glycine/valine ratios in substrate deprived growth retarded fetal rats.

Characteristic profiles of the free amino acid concentration in umbilical cord blood of growth retarded newborns have been observed. We hypothesized that the amniotic fluid of growth retarded fetal rats would show an increase in the ratio between glycine and valine which would parallel the pattern observed in the cord blood of growth retarded neonates, thus providing an index for the antepartum identification of the substrate deprived growth retarded fetus. Six test and 6 control dams were tested. Four fetuses per dam, matched for uterine location were examined. Test animals were fasted for 72 hours. Sampling was performed on day 21 under anaesthesia. Fetal size was significantly reduced (P < 0.0001) in the test group. [T = 2.68 gs. +/- 0.28 vs. C = 3.67 gs. +/- 0.25]. Fetal plasma concentrations of glycine showed an increase in test animals (P < 0.01) while valine showed a significant reduction (P < 0.0001). Glycine (pm/microliters) T = 308 +/- 64 vs. C = 269 +/- 47, valine (pm/microliters) T = 424 +/- 79 vs. C = 671 +/- 218]. Amniotic fluid concentrations for both glycine and valine were significantly decreased (P < 0.0001) in test animals. [Glycine (pm/microliters) T = 710 +/- 124 vs. C = 931 +/- 178; valine (pm/microliters) T = 845 +/- 169 vs. C = 1,339 +/- 234]. The glycine/valine ratio was significantly increased (P < 0.01) in both fetal plasma and amniotic fluid in test animals [Plasma T = 0.74 +/- 0.18 vs. C = 0.43 +/- 0.13. Amniotic fluid T = 0.85 +/- 0.08 vs. C = 0.69 +/- 0.09]. Consistent with our hypothesis, the amniotic fluid concentrations generally parallel the observations made in the plasma. This finding could enhance the antepartum identification of the substrate deprived growth retarded fetus.     

Anaerobic Growth and Fermentation Characteristics of Paecilomyces lilacinus Isolated from Mullet Gut

The anaerobic growth and fermentation of a marine isolate of Paecilomyces lilacinus is described. The fungus was isolated from mullet gut and grew optimally at 30°C and at a salinity of ≥10%. The best growth was obtained with glucose or laminarin as substrate, and the growth yield was 5.0 g (dry weight of fungus) per mol of hexose fermented. Moles of products as a percentage of moles of hexose fermented were acetate, 29.0%; ethanol, 156.6%; CO₂, 108.0%; and lactate, 4.3%. Together these products accounted for >80% of hexose carbon. Hydrogen and formate were not detectable as fermentation end products (<0.5%). Other substrates utilized for growth, although less effectively than laminarin or glucose, included the monosaccharides galactose, fructose, arabinose, and xylose and the disaccharides maltose and cellobiose. No growth of the fungus occurred on cellulose, and of a variety of other polysaccharides tested only xylan supported growth.     

Gas Chromatography-Mass Spectrometry of N- Heptafluorobutyryl Isobutyl Esters of Amino Acids in the Analysis of the Kinetics of [N]H(4) Assimilation in Lemna minor L

Rapid, sensitive, and selective methods for the determination of the ¹⁵N abundance of amino acids in isotopic tracer experiments with plant tissues are described and discussed. Methodology has been directly tested in an analysis of the kinetics of [¹⁵N]H₄⁺ assimilation in Lemna minor L. The techniques utilize gas chromatography-mass spectrometry selected ion monitoring of major fragments containing the N moiety of N-heptafluorobutyryl isobutyl esters of amino acids. The ratio of selected ion pairs at the characteristic retention time of each amino acid derivative can be used to calculate ¹⁵N abundance with an accuracy of ±1 atom% excess ¹⁵N using samples containing as little as 30 picomoles of individual amino acids. Up to 11 individual amino acid derivatives can be selectively monitored in a single chromatogram of 30 minutes. It is suggested that these techniques will be useful in situations where the small quantities of N available for analysis have hitherto hindered the use of ¹⁵N-labeled precursors.     

Cysteine transport and sulfite reductase activity in a germination-defective mutant of Histoplasma capsulatum.

A mutant of the dimorphic, zoopathogenic fungus Histoplasma capsulatum, defective in the ability of its blastospores to germinate, was used to show that the nutritional requirement for cysteine and the expression of sulfite reductase activity are temperature dependent and not related to the growth form the fungus, whereas the kinetics of cysteine transport depend on both temperature and the form of the fungus.     

Successful superovulation, nonsurgical collection and transfer of embryos from Brahman cows

Nonsurgical recoveries and transfers of embryos were performed at the McKellar Embryo Transplant Center from 122 superovulated Brahman cows. FSH-P (Armour) was used to superovulate all cows at dose levels ranging from 36 to 48 mg total FSH-P. Luteal regression was induced by use of 40 mg PGF(2(alpha)) in all 122 cows. Embryos were transferred into recipients 6, 7 or 8 days after observed estrus. Embryos were successfully collected from 82% of the FSH-P treated cows. The dose level of FSH-P affected numbers of embryos collected (P<.05). Numbers of embryos collected from cows superovulated with 36-38, 40, 42, 43, 44, 45, 46 and 47-48 mg FSH-P were 2.8 +/- 1.0, 6.8 +/- 1.1, 9.4 +/- 1.4, 10.0 +/- 2.7, 7.1 +/- 1.6, 6.8 +/- 2.0, 5.0 +/- 1.7 and 4.6 +/- 2.0 embryos, respectively. The dose level of FSH-P also affected numbers of embryos transferred (P<.10). Number of embryos transferred from cows superovulated with 36-38, 40, 42, 43, 44, 45, 46 and 47-48 mg FSH-P were 2.8 +/- 1.9, 5.2 +/- 0.9, 6.9 +/- 1.2, 6.7 +/- 2.1, 4.8 +/- 1.3, 5.1 +/- 1.4, 3.4 +/- 1.2 and 3.2 +/- 2.1 embryos, respectively. The developmental stage (D) of the embryo was also a factor in pregnancy rate of recipients (morula = 13.8%, blastocyst = 22.1% and expanded blastocyst = 29.9%; P<.005). The skill of the technician (T) transferring the embryo had a dramatic effect upon subsequent pregnancy rate of the recipients (T 1 = 46.0% vs T 2 = 22.6% pregnancy rate; P<.005). Pregnancy rate of recipients was also affected by the stage postestrus (S) at which the embryo was transferred (day 6 = 23.5%, day 7 = 25.5% and day 8 = 42.3% pregnancy rate; P<.05). Interactions were found between T x S, T x D, S x D and T x S x D (P<.05). These data indicate that use of 40, 42, or 43 mg total doses of FSH-P were quite effective in superovulating the Brahman cow. Recipients transferred on day 8 postestrus achieved higher pregnancy rates than recipients transferred on days 6 or 7 postestrus. Embryos transferred in the expanded blastocyst stage of development proved to yield the highest pregnancy rates in recipients.