Progesterone supplementation to improve fertility of selected subgroups of lactating cows during the summer and fall.

One major cause of low fertility of cows in the summer is progesterone deficiency. We found that insertion of a controlled intravaginal drug-releasing (CIDR) device containing progesterone after artificial insemination (AI) increases pregnancy per AI (P/AI) in cows with uterine disease and low body condition score after calving. Here, we treated only these two subgroups, during the summer and autumn. Control (n = 191 AI) and treatment (n = 230 AI) cows were inseminated at estrus and the treated group received a CIDR device on day 5 post-AI, for 14 days. Overall analysis of data during the summer and autumn indicated no significant differences between treatment and control groups. Analysis of the summer data only indicated a significant effect of treatment: P/AI was higher in CIDR-treated vs. control groups (34.2% vs. 19.3%; p < .038). Results indicated a 15% increase in P/AI during the summer for CIDR-treated cows in subgroups that had responded positively to the progesterone treatment.

Administration of GnRH at Onset of Estrus, Determined by Automatic Activity Monitoring, to Improve Dairy Cow Fertility during the Summer and Autumn.

We examined gonadotropin-releasing hormone (GnRH) administration at onset of estrus (OE), determined by automatic activity monitoring (AAM), to improve fertility of dairy cows during the summer and autumn. The study was performed on two dairy farms in Israel. The OE was determined by AAM recorded every 2 h, and a single im dose of GnRH analogue was administered shortly after OE. Pregnancy was determined by transrectal palpation, 40 to 45 d after artificial insemination (AI). Conception risk was analyzed by the GLIMMIX procedure of SAS. Brief visual observation of behavioral estrus indicated that about three-quarters of the events (n = 40) of visually detected OE occurred within 6 h of AAM-detected OE. Accordingly, the GnRH analogue was administered within 5 h of AAM-detected OE, to overlap with the expected endogenous preovulatory LH surge. Overall, pregnancy per AI (P/AI) was monitored over the entire experimental period (summer and autumn) in 233 first, second or third AI (116 and 117 AI for treated and control groups, respectively). Least square means of P/AI for treated (45.8%) and control (39.4%) groups did not differ, but group-by-season interaction tended to differ (p = 0.07), indicating no effect of treatment in the summer and a marked effect of GnRH treatment (n = 58 AI) compared to controls (n = 59 AI) on P/AI in the autumn (56.6% vs. 28.5%, p < 0.03). During the autumn, GnRH-treated mature cows (second or more lactations), and postpartum cows exhibiting metabolic and uterine diseases, tended to have much larger P/AI than their control counterparts (p = 0.07-0.08). No effect of treatment was recorded in the autumn in first parity cows or in uninfected, healthy cows. In conclusion, administration of GnRH within 5 h of AAM-determined OE improved conception risk in cows during the autumn, particularly in those exhibiting uterine or metabolic diseases postpartum and in mature cows. Incorporation of the proposed GnRH treatment shortly after AAM-detected OE into a synchronization program is suggested, to improve fertility of positively responding subpopulations of cows.

Oocyte maturation in plasma or follicular fluid obtained from lipopolysaccharide-treated cows disrupts its developmental competence.

Mastitis has deleterious effects on ovarian function and reproductive performance. We studied the association between plasma or follicular fluid (FF) obtained from endotoxin-induced mastitic cows, and oocyte developmental competence. Lactating Holstein cows were synchronized using the Ovsynch protocol. On Day 6 of the synchronized cycle, an additional PGF2α dose was administered, and either Escherichia coli endotoxin (LPS, 10 µg; n = 3 cows) or saline (n = 3 cows) was administered to one udder quarter per cow, 36 h later. Milk samples were collected and rectal temperatures recorded. Cows treated with LPS showed a typical transient increase in body temperature (40.3 °C ±â€¯0.4), whereas cows treated with saline maintained normal body temperature (38.9 °C ±â€¯0.04). A higher (P < 0.05) somatic cell count was recorded for cows treated with LPS. Plasma samples were collected and FF was aspirated from the preovulatory follicles by transvaginal ultrasound probe, 6 h after LPS administration. Radioimmunoassay was performed on plasma samples to determine estradiol and cortisol concentrations. Either FF or plasma was further used as maturation medium. In the first experiment, oocytes were matured in TCM-199 (Control) or in FF aspirated from cows treated with saline (FF-Saline) or LPS (FF-LPS). Cleavage rate to the 2- to 4-cell stage embryo did not differ among groups. However, the proportion of developed blastocysts on Day 7 postfertilization in the FF-LPS group tended to be lower for that in FF-Saline and was lower (P < 0.05) than that in the Control groups (10.6 vs. 22.4 and 24.4%, respectively). In the second experiment, oocytes were matured in TCM-199 (Control), or in plasma obtained from cows treated with saline (Plasma-Saline) or LPS (Plasma-LPS). Similar to the FF findings, cleavage rate did not differ among groups; however, the proportion of developing blastocysts tended to be lower in the Plasma-LPS group than in the Plasma-Saline group and was lower (P < 0.05) from that in the Control group (11.0 vs. 25.5 and 34.7%, respectively). The proportion of apoptotic cells per blastocyst, determined by TUNEL assay, did not differ among the experimental groups. The findings shed light on the mechanism by which mastitis induces a disruption in oocyte developmental competence. Further studies are required to clarify whether the negative effect on oocyte developmental competence is a result of LPS, by itself, or due to elevation of secondary inflammatory agents.

Experimental model of toxin-induced subclinical mastitis and its effect on disruption of follicular function in cows.

This study establishes an experimental model for subclinical mastitis induced by Gram-positive (G+) exosecretions of Staphylococcus aureus origin or Gram-negative (G-) endotoxin of Escherichia coli origin to examine its effects on follicular growth and steroid concentrations in Holstein dairy cows. Cows were synchronized with the Ovsynch protocol followed by a series of follicular cycles that included GnRH and PGF2α doses administered every 8 days. Cows received small intramammary doses of either G+ (10 µg, n = 10) or G- (0.5 µg, n = 6) toxin, or saline (n = 6; uninfected control) every 48 hours for 20 days. Follicular fluids were aspirated from preovulatory follicles before (aspiration one: control), at the end of (aspiration two: immediate effect), and 16 days after the end of (aspiration three: carryover effect) toxin exposure. During the 3 weeks of subclinical mastitis induced by G+ or G-, no local inflammatory signs were detected in the mammary gland and no systemic symptoms were noted: body temperatures of the treated cows did not differ from controls; plasma cortisol and haptoglobin concentrations were not elevated and did not differ among groups. Somatic cell count was higher in the treated groups than in controls, and higher in the G- versus G+ group. For analysis of reproductive responses, cows were further classified as nonaffected or affected based on an more than 20% decline in follicular androstenedione concentration in aspiration two or three relative to the first, control aspiration. Most G- (5/6) and 40% of G+ (4/10) cows were defined as affected by induced mastitis. An immediate decrease in the number of medium-size follicles was recorded on Day 4 of the induced cycle, toward the end of the 20-day mastitis induction, in the affected G+ compared with uninfected control group (1.0 ± 0.5 vs. 3.0 ± 0.4 follicles; P < 0.05); the affected G- and nonaffected G+ subgroups exhibited a similar numerical decline in the number of follicles. A carryover (but not immediate) decrease to 51% and 62% in follicular estradiol concentrations in G- affected group and G+ affected group was detected relative to controls (P < 0.05). The nonaffected G+ subgroup did not differ from its control counterparts. Based on the current experimental model, subclinical IMI induced by G+ or G- toxin disrupts follicular functions, and it seems that the ovarian pool of early antral follicles is susceptible to subclinical mastitis.

Induction of heparanase in bovine granulosa cells by luteinizing hormone: possible role during the ovulatory process.

Follicular development, follicular rupture, and corpus luteum (CL) formation are accompanied by extensive tissue remodeling. We examined whether heparanase (HPSE), which cleaves heparan sulfate glycosaminoglycans, is induced during these processes. Prostaglandin F2alpha injection, which initiated luteolysis and the development of a preovulatory follicle, moderately increased HPSE mRNA in bovine granulosa cells (GCs). GnRH, used to induce gonadotropin surge, markedly augmented HPSE mRNA levels 12 h after its injection. The temporal pattern of HPSE gene expression in follicular-luteal transition was further examined in follicles collected before, and 4, 10, 20, 25, and 60 h after GnRH injection. HPSE mRNA increased transiently 10-20 h after GnRH injection to levels 10-fold higher than in untreated heifers. HPSE protein levels were similarly elevated 20 h after GnRH injection in GCs, but not in the theca layer. Cyclooxygenase-2 (PTGS2) mRNA peaked before ovulation when HPSE levels returned to baseline levels. HPSE mRNA abundance also remained low in the CLs. The antiprogesterone, RU-486, elevated HPSE levels in GC culture, suggesting that progesterone secreted by CLs may inhibit HPSE. HPSE immunostaining was more abundant in GCs than thecae. In cultured GCs, LH induced a transient increase in HPSE mRNA 3-6 h after its addition, but not at 24 h. However, PTGS2 mRNA was clearly induced at this time. These findings suggest that: 1) HPSE may play a role in ovulation but much less so during CL development, and 2) GC-derived HSPE may be a novel member of the LH-induced extracellular matrix-degrading enzyme family and may contribute to follicular rupture.

Follicular dynamics and concentrations of steroids and gonadotropins in lactating cows and nulliparous heifers.

Differences in follicular development and circulating hormone concentrations, between lactating cows and nulliparous heifers, that may relate to differences in fertility between the groups, were examined. Multiparous, cyclic, lactating Holstein cows (n=19) and cyclic heifers (n=20) were examined in the winter, during one estrous cycle. The examinations included ultrasound monitoring and daily blood sampling. Distributions of two-wave and three-wave cycles were similar in the two groups: 79 and 21% in cows, 70 and 30% in heifers, respectively. Cycle lengths were shorter by 2.6 days in heifers than in cows, and in two-wave than in three-wave cycles. The ovulatory follicle was smaller in heifers than in cows (13.0+/-0.3 mm versus 16.5+/-0.05 mm). The greater numbers of large follicles in cows than in heifers corresponded well to the higher concentrations of FSH in cows. The duration of dominance of the ovulatory follicle tended to be longer in cows than in heifers. Estradiol concentrations around estrus and the preovulatory LH surge were higher in heifers than in cows (20 versus 9 ng/ml). Progesterone concentrations were higher in heifers than in cows from Day 3 to Day 16 of the cycle. Circulating progesterone did not differ between two-wave and three-wave cycles. The results revealed differences in ovarian follicular dynamics, and in plasma concentrations of steroids and gonadotropins; these may account for the differences in fertility between nulliparous heifers and multiparous lactating cows.