Biomedical subjects

F C Gwazdauskas

Publications and source records attributed to F C Gwazdauskas.

At least 19 recordsLinked to original sources

PubMed2006

Dopamine antagonist alters serum cortisol and prolactin secretion in lactating Holstein cows.

The role of dopamine in regulating glucocorticoid and prolactin secretion was investigated in lactating Holstein cows by characterizing serum cortisol and prolactin responses to fluphenazine, a dopamine receptor antagonist. Twelve anovulatory cows received an intravenous bolus injection of either saline (n = 6) or 0.3 mg of fluphenazine/kg of body weight (n = 6) in wk 2 postpartum. Blood samples were collected every 30 min for 4 h before and 4 h after saline or fluphenazine injection. Serum progesterone concentration was 0.13 +/- 0.1 ng/mL and did not differ between groups. No difference in serum cortisol concentrations was detected between groups before treatments. Fluphenazine increased serum cortisol concentrations within 30 min after fluphenazine administration (>30 ng/mL) and concentrations remained elevated throughout the sampling period. Cortisol remained unchanged in saline-treated cows (<10 ng/mL). Prolactin concentrations also increased after fluphenazine administration (103.1 +/- 3.1 ng/mL), but were unaffected by saline (18 +/- 3.1 ng/mL). Prolactin concentrations remained elevated throughout the sampling period in fluphenazine-treated cows. Our results indicated that a dopamine antagonist increased cortisol, suggesting that endogenous dopamine, at least in part, regulates cortisol and prolactin secretion. These effects are regulated through dopamine receptors in anovulatory lactating dairy cows during the early postpartum period.

Animals↗

PubMed2005

The incidence of apoptosis after IVF with morphologically abnormal bovine spermatozoa.

Normal embryonic development depends on the maintenance of a population of normal healthy cells within each embryo. The aim of this study was to use a combination of apoptotic measures to assess differences in embryo quality after IVF with semen samples with high percentages of abnormal spermatozoa. Semen samples were obtained and cryopreserved from four Holstein bulls before (5 day prior) and after (2 week-post-insult; 2 week-PI and day 20; 3 week-PI) a scrotal insulation period of 48 h (day 0). The swim-up sperm separation method was used. The post-thaw morphology revealed a decrease (P < 0.01) in the percentages of normal spermatozoa in the 3 week-PI samples in comparison with the pre-insult samples for Bulls I and III (74-22.3 and 67.7-0.5%, respectively). The percentage of vacuolated spermatozoa increased significantly for Bull II. After 18 h of sperm-oocyte co-incubation, zygotes were cultured and subpopulations were removed from culture at day 8 and subjected to either the TUNEL or caspase assay. On day 8, caspase intensity increased significantly for both Bull I (217+/-147) and Bull III (229+/-98) for the 3 week-PI embryo groups compared to the equivalent embryo groups for Bull II (98+/-115) and Bull IV (90+/-111). In conclusion, the inability to consistently measure apoptosis with TUNEL alone complicated the assessment of differences in embryo quality. Thus, it is uncertain exactly when during early pre-implantation development the differences in embryo quality are first manifest. Despite discrepancies, our results clearly indicated a difference in the embryo quality between embryos obtained after IVF with semen samples from bulls that had an intense response to scrotal insulation.

Animals↗

PubMed2005

Assessment of pronuclear formation following in vitro fertilization with bovine spermatozoa obtained after thermal insulation of the testis.

This study was conducted to follow the chronology of pronuclear formation in bovine zygotes after in vitro insemination with a population of spermatozoa having abnormal morphology. Semen samples were obtained and cryopreserved from four Holstein bulls before and after a scrotal insulation period of 48 h (Day 0). A pre-insult (Day 5) and a Day 20 post-insult semen sample were evaluated for morphology and used for IVF after standard swim-up sperm separation protocols. Pronuclear formation was scored on subpopulations of presumptive zygotes after they were fixed and stained at 3-h time intervals from 6 to 18 h post in vitro insemination (hpi). Post-thaw morphological evaluation of semen samples revealed a decrease in the percentages of normal spermatozoa in the post-insult samples compared with the pre-insult samples for Bull I (74-22%) and Bull III (68-1%). The sperm penetration rate decreased (P<0.05) between the pre- and post-insult samples for Bulls I (90-76%) and III (92-70%), but was not different for Bulls II (92-90%) and IV (78-85%). The pronuclear formation rates for post-insult zygotes for Bulls II and IV had comparable increases in development over time, whereas there was no increase in the pronuclear development for the zygotes from the post-insult samples for Bulls I and III, and generally a condensed sperm head was observed in the oolemma. At 18 hpi the fertilization rate between the pre- and post-insult samples for Bulls I (51-4%), II (88-75%) and III (94-2%) decreased (P<0.01), but there was no change for Bull IV (66%). In conclusion, we inferred that the failure in normal pronuclear formation was associated with an absence of normal decondensation of the penetrating spermatozoon; this suggested that the effect of morphologically abnormal spermatozoa occurred prior to cleavage, thus limiting early development.

Animals↗

PubMed2005

Bovine embryo development after IVF with spermatozoa having abnormal morphology.

The study was conducted to evaluate the effects of scrotal insulation on semen samples collected from bulls on embryonic development after IVF. Semen samples were obtained and cryopreserved from four Holstein bulls before and after a scrotal insulation period of 48 h (Day 0). Three types of samples were used for IVF: (1) semen from the test bulls collected 5 d prior to scrotal insulation (pre-insult); (2) semen from Day 13 (2-week post-insult; 2-week PI); and (3) semen from Day 20 (3-week PI). After 18 h of sperm-oocyte co-incubation, the zygotes were cultured for 8 d when a developmental score (0=degenerate, 1=2-cell embryo through 5=blastocyst) was assigned to each embryo. The post-thaw morphological evaluation of sperm samples revealed a decrease (P<0.01) in the percentages of normal spermatozoa in the 3-week PI samples in comparison with the pre-insult samples for Bulls I and III (74-22.3% and 67.7-0.5 %, respectively). The percentage of vacuolated spermatozoa increased significantly for Bull II. The cleavage and blastocyst formation rates and embryo development scores were affected (P<0.01) by the interaction of bull by sample collection time. For Bulls I and III (severe responders) the scrotal insulation effects persisted from the time of cleavage through blastocyst formation. In contrast, the cleavage and blastocyst formation rates for Bulls II and IV were unaffected, despite high percentages of vacuolated spermatozoa present in the post-insult samples for Bull II. In conclusion, the use of scrotal insulation to elevate scrotal temperature was an effective method to obtain semen samples with high percentages of abnormal spermatozoa. The decrease in embryonic development after IVF when using spermatozoa with morphological abnormalities seemed to be multifaceted and related to changes in head morphology.

Acrosome↗

PubMed2004

Xenogenous fertilization of equine oocytes following recovery from slaughterhouse ovaries and in vitro maturation.

The in vitro production (IVP) of equine embryos using currently available protocols has met limited success; therefore investigations into alternative approaches to IVP are justified. The objective of this study was to evaluate the feasibility of xenogenous fertilization and early embryo development of in vitro matured (IVM) equine oocytes. Follicular aspirations followed by slicing of ovarian tissue were performed on 202 equine ovaries obtained from an abattoir. A total of 667 oocytes (3.3 per ovary) were recovered from 1023 follicles (recovery rate, 65%). Oocytes underwent IVM for 41 +/- 2 h (mean +/- S.D.), before being subjected to xenogenous gamete intrafallopian transfer (XGIFT). An average of 13 +/- 0.8 oocytes and 40x10(3) spermatozoa per oocyte were transferred into 20 oviducts of ewes. Fourteen percent of transferred oocytes (36/259) were recovered between 2 and 7 days post-XGIFT and 36% of those recovered displayed embryonic development ranging from the 2-cell to the blastocyst stage. Fertilization following XGIFT was also demonstrated by the detection of zinc finger protein Y (ZFY) loci. Ligation of the uterotubal junction (UTJ), ovarian structures, or the duration of oviductal incubation did not significantly affect the frequency of embryonic development or recovery of oocytes/embryos after XGIFT. In conclusion, equine embryos can be produced in a smaller non-equine species that is easier for handling.

Animals↗

PubMed2003

Development of murine embryos following electroporation.

PURPOSE: The objective was to establish the parameters for reversible electroporation of murine embryos. METHODS: In Trial 1, murine presumptive zygotes received an electrical pulse of 5, 10, or 20-micros duration, and one of five voltages (100, 200, 250, 300, or 400 V). In Trial 2, embryo orientation within the electroporation chamber was evaluated with 250 or 400 V at a pulse period of 10 micros. RESULTS: Presumptive zygotes that received 400 V at each pulse length and zygotes exposed to 20 micros at each voltage had the lowest embryonic development (P < 0.05). Presumptive zygotes that received 250 V had higher development compared to 400 V, irrespective of orientation (P < 0.01), but development was lower than the controls (P < 0.01). CONCLUSIONS: Electrical stimulation of presumptive zygotes can have a detrimental impact on early embryo development, but low amounts of stimulation may allow for potential gene transfer in transgenic experimentation.

Animals↗

PubMed2002

Milk yield, energy balance, hormone, follicular and oocyte measures in early and mid-lactation Holstein cows.

The purpose of the study was to determine the influence of energy status on metabolic and endocrine measures, follicular development, and the quality of oocytes obtained from cows during early and mid-lactation (ML). We selected Holstein cows at calving to be assigned to the early lactation (EL) group (n = 8), while we assigned cows at about day 90 postpartum to the ML group (n = 7). We obtained blood samples twice weekly from 4 weeks before aspiration to the aspiration periods for metabolite and hormone determinations. We performed ultrasound-guided transvaginal follicular aspiration (TVFA) twice weekly on all cows for a 10-week period. We obtained follicular fluid from the largest follicle > 10 mm in diameter for hormone determinations. We analyzed data by ANOVA, using the general linear model (GLM) procedures. Energy balance was positive (2.43 +/- 0.32 Mcal/kg) for ML cows and negative (-1.55 +/- 0.33 Mcal/kg) for EL cows. Serum progesterone (P4) for ML cows decreased rapidly from the first aspiration session (2.7 +/- 0.1 ng/ml) and reached a nadir at Week 8 (0.33 +/- 0.1 ng/ml), while follicular fluid P4 increased from 0.9 +/- 0.5 to 5.6 +/- 0.05 ng/ml. Serum and follicular fluid P4 remained relatively constant over the entire aspiration period for EL cows. Follicular fluid insulin-like growth factor I (TGF-I) concentrations increased linearly for EL and ML cows, but the increase was more rapid (159 +/- 36 to 200 +/- 36 ng/ml) for ML cows than for EL cows (145 +/- 36 to 164 +/- 36 ng/ml). Serum IGF-I followed the same pattern for ML cows but declined for EL cows. Early lactation cows experienced a rapid decrease in serum nonesterified fatty acids (NEFA; 0.32 +/- 0.2 to 0.22 +/- 0.2 meq/l), while serum NEFA concentrations were relatively stable (0.19 +/- 0.2 to 0.22 +/- 0.2 meq/l) for ML cows over the aspiration period. The number of follicles obtained from the twice weekly aspiration sessions increased linearly for both EL and ML cows (P < 0.05) over the 10-week period. However, the number of follicles increased from 14.2 +/- 0.5 (Day 119) to 18.1 +/- 0.5 (Day 190) in the ML cows, compared to the changes from 14.9 +/- 0.3 (Day 32) to 15.7 +/- 0.5 (Day 90) for the EL cows. These results indicate that cows are physiologically under more production stress during EL, but increasing follicular fluid and serum IGF-I throughout ML may reflect potential differences in follicle and oocyte measures, compared to cows in EL.

Animals↗

PubMed2002

Parity-related changes in bovine follicle and oocyte populations, oocyte quality, and hormones to 90 days postpartum.

Ultrasound-guided transvaginal follicular aspiration was used to obtain oocytes from Holstein cows to study follicular development and oocyte morphology during early lactation. A single aspiration was conducted on each of the 120 cows during wk 1 to 12 postpartum. On the day of aspiration, the number of follicles and their sizes were recorded. The collected oocytes were morphologically classified into four grades. Blood and follicular fluid samples were collected for hormone and metabolite assays. Greater serum nonesterified fatty acids concentrations for first- and second-parity cattle on d 7 implied a more negative energy balance status than for third-parity cattle. The numbers of follicles and oocytes retrieved were affected by linear days postpartum x parity interactions, with second-parity cattle having increases compared with decreases for third-parity cattle. Oocyte quality score was affected by the quadratic days postpartum x parity interaction. First- and second-parity cattle had greater oocyte quality scores from d 20 to 70 postpartum compared with third-parity cattle, but third-parity cattle had greater quality oocytes near the end of the 12-wk period. In contrast to cattle subjected to multiple aspirations, first- and second-parity cattle had higher estradiol and insulin-like growth factor I concentrations in follicular fluid associated with greater numbers of oocytes retrieved than third-parity cattle. This study demonstrated that conditions related to early lactation have a negative effect on oocyte quality and endocrine measures of dairy cattle and that animals of various parities may be differentially affected.

Animals↗

PubMed2001

Recombinant human protein C expression in the milk of transgenic pigs and the effect on endogenous milk immunoglobulin and transferrin levels.

Colostrum and milk are natural vehicles for acquiring passive immunity and are valuable tools for decreasing neonatant mortality from diarrheal disease. The effects of recombinant human protein C (rhPC) expression levels on endogenous immunoglobulin and transferrin content of the milk of different lineages of transgenic pigs were studied. The levels of rhPC in the milk ranged from 40 to 1200 microg/ml. Transgenic pigs with rhPC expression levels less than 500 microg/ml had no significant differences in milk protein composition with respect to nontransgenic pigs. A line of transgenic pigs having rhPC expression levels of 960-1200 microg/ml had two- to three-fold higher IgG, IgM, and secretory IgA concentrations compared to other transgenic and nontransgenic pig groups (P < 0.05), and four- to five-fold higher transferrin levels than nontransgenic pigs (P < 0.05). Changes in milk protein composition were not associated with mastitis or other pathologic disruption of epithelial cell junctions as indicated by normal casein and albumin levels in milk. Since IgG, IgM, secretory IgA, and transferrin are transported into the milk by transcytosis, higher levels of these proteins indicate that transcyctosis in the mammary epithelial cell was likely upregulated in pigs having high rhPC expression levels. This study is the first that shows a statistically significant example that mammary tissue specific expression of a heterologous protein can enhance endogenous phenotypic characteristics of milk.

Albumins↗

PubMed2001

Comparison of carbohydrate and milk-based beverages on muscle damage and glycogen following exercise.

This study examined effects of carbohydrate (CHO), milk-based carbohydrate-protein (CHO-PRO), or placebo (P) beverages on glycogen resynthesis, muscle damage, inflammation, and muscle function following eccentric resistance exercise. Untrained males performed a cycling exercise to reduce muscle glycogen 12 hours prior to performance of 100 eccentric quadriceps contractions at 120% of 1-RM (day 1) and drank CHO (n = 8), CHO-PRO (n = 9; 5 kcal/kg), or P (n = 9) immediately and 2 hours post-exercise. At 3 hours post-eccentric exercise, serum insulin was four times higher for CHO-PRO and CHO than P (p < .05). Serum creatine kinase (CK) increased for all groups in the 6 hours post-eccentric exercise (p < .01), with the increase tending to be lowest for CHO-PRO (p < .08) during this period. Glycogen was low post-exercise (33+/-3.7 mmol/kg ww), increased 225% at 24 hours, and tripled by 72 hours, with no group differences. The eccentric exercise increased muscle protein breakdown as indicated by urinary 3-methylhistidine and increased IL-6 with no effect of beverage. Quadriceps isokinetic peak torque was depressed similarly for all groups by 24% 24 hours post-exercise and remained 21% lower at 72 hours (p < .01). In summary, there were no influences of any post-exercise beverage on muscle glycogen replacement, inflammation, or muscle function.

Adult↗

PubMed2000

Impact of follicular aspiration on folliculogenesis as influenced by dietary energy and stage of lactation.

Prolonged dietary energy restriction has been shown to influence the secretion of ovarian hormones and follicular development. Energy balance affects the ability of the cow to support a functional reproductive system during early lactation. Follicular aspiration is a tool that can be used to assess the impact of dietary energy and stage of lactation on follicle development in dairy cattle. Follicular aspiration was conducted twice weekly from d 30 to 100 postpartum in cows fed either 104 or 80% of NRC energy requirements. Oocyte numbers recovered increased linearly from d 30 to 100 postpartum. Cows on high energy diets produced more high quality oocytes, but also produced more poor quality oocytes than cows fed the lower energy diet. Follicles < 5 mm predominated throughout. Cows in early (d 28) and mid (d 117) lactation were used to assess stage of lactation effects on serum and follicular fluid hormones, and quality of oocytes recovered by follicular aspiration procedures for 10 wk. Serum IGF-1 decreased in early lactation cows, but increased in midlactation cows. Follicular fluid progesterone in early lactation cows decreased from 275 to 235 ng/ml, whereas in midlactation cows it increased from 65 to 550 ng/ml at aspiration wk 10. The percentage of high quality ova was higher in early lactation cows (11%) than midlactation cows (6%), but fell to 1% by aspiration wk 7 and increased to 10% by wk 10, whereas percentage of high quality ova in midlactation cows steadily declined to 2% by wk 10. Milk yield and energy balance affect patterns of folliculogenesis in lactating dairy cows.

Animals↗

PubMed1999

Effects of ultrasound-guided transvaginal follicular aspiration on oocyte recovery and hormonal profiles before and after GnRH treatment.

Endocrine changes and recovered oocytes were evaluated during 16 wk of ultrasound-guided transvaginal follicular aspiration (TVFA) and prior to and following administration of GnRH at the cessation of aspiration. Nonlactating previously aspirated (PAC, n = 4) and non-aspirated, (AC, n = 4) Holstein cows were subjected to 16 wk of twice-weekly aspiration. Four control cows (OAC) were aspirated 1 time only at the final TVFA session (wk 16). Jugular blood samples were collected from all cows during aspiration, before and after the final TVFA session, and during an 18-d period following cessation of aspiration. Ovarian activity was monitored in all cows after cessation of aspiration for 18 d. The PAC and AC cows averaged 3.4 +/- 1.2 (+/- SE) and 6.8 +/- 1.2 oocytes per session, respectively. Progesterone concentrations during TVFA did not differ between the PAC and AC (0.8 +/- 0.1 and 0.9 +/- 0.1 ng/mL, respectively). Progesterone concentration in OAC was 4.5 +/- 0.2 ng/mL before TVFA, while the PAC and AC averaged 0.5 +/- 0.2 and 0.3 +/- 0.2 ng/mL, respectively, at 16 wk. At Week 16 LH was 1.0 +/- 0.2 ng/mL and it increased to 7.5 +/- 0.1 ng/mL after GnRH treatment. The LH concentration before the final aspiration session was higher at peak amplitude in PAC than in AC groups and peak length was longer in OAC than in AC cows (P < 0.07). Between 18 and 24 h after the last aspiration there were more LH peaks and greater peak frequencies in PAC than in OAC cows (P < 0.07), and the interval between peaks was longer in PAC and AC cows (P < 0.10) than in OAC cows. Mean FSH concentrations were lower (P < 0.01) for OAC than for PAC and AC groups at 20 and 24 h after the last aspiration. Follicle numbers after GnRH varied most among treatment groups for follicles < 9 mm, with the PAC, AC and OAC averaging 5.1 +/- 1.0, 5.1 +/- 1.0, and 3.8 +/- 1.0 follicles/d, respectively. Progesterone concentrations increased to 1.1 +/- 0.3 ng/mL in PAC cows and 2.5 +/- 0.3 and 3.4 +/- 0.3 ng/mL in AC and OAC groups, respectively, during the 18-d period. These results suggest that long-term TVFA affects progesterone, LH and FSH profiles and ovarian dynamics in cows.

Animals↗

PubMed1999

Transgenic pigs as bioreactors: a comparison of gamma-carboxylation of glutamic acid in recombinant human protein C and factor IX by the mammary gland.

The mammary gland of transgenic livestock can be used as a bioreactor for producing complex therapeutic proteins. However, the capacity for making a given post-translational modification upon any given polypeptide is uncertain. For example, the efficiency of gamma-carboxylation of glutamic acid in the amino terminal regions of recombinant human protein C (rhPC) and recombinant human Factor IX (rhFIX) is different at similar expression levels. At an expression level of about 200 microg/ml in the milk of transgenic pigs, rhFIX is highly gamma-carboxylated as indicated by pro-coagulant activity and amino acid sequencing. However, only about 20-35% of rhPC has a native, gamma-carboxyglutamic acid-dependent conformation and anti-coagulant activity. Thus, this work provides an example of apparent differences in substrate specificity between two homologous proteins to the endogenous carboxylase of porcine mammary epithelium which leads to varying degrees of post-translational modification.

Amino Acid Sequence↗

PubMed1999

Technical note: machine milking of sows--lactational milk yield and litter weights.

This study was designed to 1) determine milk yield of sows that were machine milked; 2) assess the effects of pulsation rate, pulsation ratio, and pig removal on milk yield; and 3) assess litter weights. In Exp. 1, four sows were milked daily to 60 d postpartum. There were differences (P < .05) in milk yield among sows, the greatest being 1,898 mL/d. Daily milk yield peaked between 15 and 25 d postpartum. Litter weights were 18.0 +/- 1.0 kg at farrowing and 60.8 +/- 12.1 kg at d 60. In Exp. 2, four sows were milked daily for 28 d. Pulsation rate and ratio (150/min and 28:72, milk:rest, and 60/min and 50:50) were alternated on a daily basis and pigs were isolated for either 0 or 60 min before milking. The higher pulsation rate resulted in more milk per milking (202 +/- 13 vs 168 +/- 13 mL; P < .05). Pig removal resulted in 221 +/- 11 vs 148 +/- 14 mL milk (P < .01). Pig removal times and pulsation characteristics affect the amount of milk obtained, but milk removal from sows has a severe effect on litter weight. This system can be used to harvest sow's milk for pharmaceutical purposes, but supplementation of the pigs is necessary.

Animal Husbandry↗

PubMed1999

Influence of pig substitution on milk yield, litter weights, and milk composition of machine milked sows.

This study was conducted to 1) determine milk yield of sows that were machine milked up to four times daily; 2) determine the effect of pig substitution on milk yield; 3) assess litter weight changes for sows that are milked; and 4) determine milk composition. Eight sows were milked four times daily to d 51 postpartum. Sows either maintained their own litter or had a week-old replacement litter to replace 25-d-old pigs. Individual gland milk yields were obtained on random days throughout lactation, and different diameter and weighted teat cups were rotated so that all glands received all combinations. Composite milk samples were analyzed for fat, protein, and somatic cells. Milk yields peaked at about 19 d postpartum and declined to 45 d postpartum in sows with their own litter, whereas milk yields peaked earlier and had a more dramatic decline after fostering of a younger litter. Litter weights were 17.1 +/- 1.0 kg at farrowing with 13.6 +/- .6 pigs born alive. Final litter weights were 34.4 +/- 11.7 kg for sows with replacement litters and 74.4 +/- 13.5 kg for sows with their own litters, and numbers of pigs weaned were 6.5 +/- 1.3 and 9.7 +/- 1.5, respectively. Milk fat was influenced by route of oxytocin administration (6.53 +/- .12 for i.v. vs 7.21 +/- .19% for i.m. administration; P < .05). Milk fat percentage was highest on d 2 and declined to 13 d postpartum. Milk protein was influenced by time of day of milking (lowest at the fourth milking, 5.57 +/- .11%) and followed a pattern similar to that for milk fat. Milk protein was affected in a linear manner by milk yield, with highest protein associated with lowest milk yields. Somatic cells in milk were influenced by litter replacement (P < .05) and oxytocin administration (P < .01). There was a linear increase in somatic cells from about 8 x 10(6) cells/mL milk at d 2 to more than 12 x 10(6) cells/mL milk at d 51 postpartum. These results show that pig replacement affects the amount of milk obtained. Moreover, milk composition changes throughout lactation. However, milk removal from sows has a severe impact on litter weight gains, and in systems where sow's milk is needed for commercial purposes, pig supplementation is necessary.

Analysis of Variance↗

PubMed1999

Bovine follicular dynamics, oocyte recovery, and development of oocytes microinjected with a green fluorescent protein construct.

The present study was carried out to 1) evaluate the viability of in vitro fertilized zygotes after microinjection of DNA, 2) assess the influence of oocyte quality upon the development rate of embryos when injected with DNA, and 3) determine the integration frequency of green fluorescent protein DNA into microinjected embryos. Oocytes were aspirated from ovaries of nine nonlactating Holsteins and were categorized into grades A, B, C, and D. At 16 h after in vitro fertilization, approximately half of the pronuclear stage presumptive zygotes were classified as having 1 pronucleus or 2 pronuclei, and they were microinjected with DNA constructs. A potential predictor of DNA integration frequency at d 10 was assessment of the incidence of green fluorescing embryos. The proportion of cleaved embryos that developed to morulae or blastocysts was not different between groups with 1 pronucleus injected (45%), 1 pronucleus uninjected (64%), or 2 pronuclei injected (49%). However, the development of morulae or blastocysts was higher in the group with 2 pronuclei uninjected (69%). The overall developmental score of green fluorescent protein-positive embryos was higher for grade A oocytes (1.3 +/- 0.1) than for grade B (0.8 +/- 0.1), C (0.6 +/- 0.1), or D (0.3 +/- 0.1) oocytes. The results show that production of transgenic bovine blastocysts can occur from the microinjection of a presumptive zygote having only one visible pronucleus. Initial oocyte quality is an important factor in selection of oocytes suitable for microinjection of DNA and for preimplantation development to produce bovine transgenic embryos.

Animals↗

PubMed1999

Effects of energy balance of hormones, ovarian activity, and recovered oocytes in lactating Holstein cows using transvaginal follicular aspiration.

The effects of energy balance on hormonal secretion patterns and the structure of recovered oocytes were evaluated in 20 lactating Holstein cows during two trial periods. Cows were randomly assigned to one of two dietary treatments formulated so that dry matter consumption was 3.6% of body weight (high energy; 1.78 Mcal/kg) or 3.2% of body weight (low energy; 1.52 Mcal/kg). Ovum recovery procedures were conducted twice weekly between d 30 and 100 of lactation. Follicle size and number were recorded. Follicular fluid aspirated from the largest follicle and serum samples were collected for hormone assay. Milk yield averaged 41.6 +/- 0.3 kg/d (mean +/- SE) for high energy fed cows and 32.8 +/- 0.3 kg/d for low energy fed cows. Oocyte numbers increased linearly from d 30 to 100 postpartum. Cows fed high energy diets produced more good (+) oocytes than did cows fed low energy diets.

Animals↗

PubMed1998

Effectiveness of Menuzo's B2 medium with buffalo rat liver cells for development of in vitro matured/in vitro fertilized bovine oocytes.

PURPOSE: The effectiveness of two culture media on the development of bovine embryos in a buffalo rat liver (BRL) coculture system was investigated. METHODS: Bovine oocytes were matured and fertilized in vitro, then cocultured, 25 per well, for 7 days in 500 microliters of modified M199 or modified Menuzo's B2 medium over a BRL cell monolayer at 39 degrees C in an atmosphere of 5% CO2 in air. Medium 199 was modified by the addition of 10% of (v/v) fetal bovine serum (FBS), 9 mg/ml bovine serum albumin, 2 mM glycine, 1 mM alanine, and 0.1 mM nonessential amino acids (NEAA). Menuzo's B2 medium was modified by the addition of 10% (v/v) FBS, 1 mM alanine, and 0.1 mM NEAA. RESULTS: Modified Menuzo's B2 medium improved embryo development to the morula or blastocyst stage compared to modified M199 (121/353, 34.3%, versus 99/362, 27.3%, P < 0.05). CONCLUSIONS: These results suggest that Menuzo's B2 medium with modifications in a BRL coculture system can provide a significant benefit for culture of early bovine embryos over the traditional use of medium 199.

Animals↗