Fertility of holstein dairy heifers after synchronization of ovulation and timed AI or AI after removed tail chalk

Nonlactating Holstein dairy heifers (n=352) 13 mo of age were managed using a 42-d artificial insemination (AI) breeding period in which they received AI after removed tail chalk evaluated once daily. At AI breeding period onset (d 0), heifers were randomly assigned to receive synchronization of ovulation (100 microg of GnRH, d 0; 25 mg of PGF2alpha, d 6; 100 microg of GnRH, d 8) and timed AI (TAI; d 8) and AI after removed tail chalk for the entire AI breeding period (GPG; n=175), or AI after removed tail chalk for the entire AI breeding period (TC; n=177). As expected, 17.7% (31/175) of GPG heifers received AI after removed tail chalk before scheduled TAI. Pregnancy rate per artificial insemination (PR/AI) at approximately 30 d after first AI tended to be greater for TC (46.5%) than for GPG (38.3%) heifers. No treatment x inseminator interaction was detected; however, overall PR/AI was low for heifers in both treatments due to variation among the 3 inseminators (24.8, 30.0, and 58.0%). Pregnancy loss from approximately 30 to approximately 75 d after first AI was 10% and did not differ between treatments. Based on survival analysis, days to first AI was greater for TC than for GPG heifers, whereas days to pregnancy across the 42-d AI breeding period did not differ between treatments. Overall, 81.2% of GPG heifers receiving TAI synchronized luteal regression and ovulated within 48 h after the second GnRH injection. We conclude that this synchronization protocol can yield acceptable fertility in dairy heifers if AI to estrus is conducted between treatment with GnRH and PGF2alpha and AI efficiency is optimized.     

Kisspeptin Neurons and Estrogen–Estrogen Receptor α Signaling: Unraveling the Mystery of Steroid Feedback System Regulating Mammalian Reproduction

Estrogen produced by ovarian follicles plays a key role in the central mechanisms controlling reproduction via regulation of gonadotropin-releasing hormone (GnRH) release by its negative and positive feedback actions in female mammals. It has been well accepted that estrogen receptor α (ERα) mediates both estrogen feedback actions, but precise targets had remained as a mystery for decades. Ever since the discovery of kisspeptin neurons as afferent ERα-expressing neurons to govern GnRH neurons, the mechanisms mediating estrogen feedback are gradually being unraveled. The present article overviews the role of kisspeptin neurons in the arcuate nucleus (ARC), which are considered to drive pulsatile GnRH/gonadotropin release and folliculogenesis, in mediating the estrogen negative feedback action, and the role of kisspeptin neurons located in the anteroventral periventricular nucleus-periventricular nucleus (AVPV-PeN), which are thought to drive GnRH/luteinizing hormone (LH) surge and consequent ovulation, in mediating the estrogen positive feedback action. This implication has been confirmed by the studies showing that estrogen-bound ERα down- and up-regulates kisspeptin gene (Kiss1) expression in the ARC and AVPV-PeN kisspeptin neurons, respectively. The article also provides the molecular and epigenetic mechanisms regulating Kiss1 expression in kisspeptin neurons by estrogen. Further, afferent ERα-expressing neurons that may regulate kisspeptin release are discussed.     

Reduced dry periods and varying prepartum diets alter postpartum ovulation and reproductive measures

There has been substantial recent interest in shortening dry periods; however, the effects of this management change on reproduction have not been adequately evaluated. Holstein cows (n = 58) were assigned in a randomized block design to 1 of 3 treatments: 1) traditional (T) dry period (∼56 d) in which cows were fed a low energy diet from 56 to 29 d prepartum followed by a moderate energy diet for 28 d; 2) shortened (S) dry period (∼28 d) in which cows were fed continuously a high energy diet; or 3) no planned (N) dry period in which cows were fed continuously a high energy diet. All cows received a high energy lactation diet after calving. Ovaries were evaluated by ultrasound and blood samples collected 3 times weekly beginning at d 6 or 7 postpartum until 7 d after second ovulation. Average days from calving until first detection of a 10-mm follicle were fewer in N (8.0 d) and S (8.9 d) than in T (10.5 d) cows. Time from calving to first ovulation was earlier in N (13.2 d) than in S (23.8 d) and T (31.9 d) cows. A greater percentage of follicles of the first follicular wave ovulated in N (89%; 16/18) than in T (42%; 8/19), with S (62%; 13/21) cows being intermediate. Double ovulation rate at the first ovulation was greater in T (61%) than N (16%), with S (35%) intermediate. No difference was detected in double ovulation rate at second ovulation (13/56). Number of cows with persistent corpus luteum (>30 d; 15/56) was not different among groups; however, short luteal phases were greater in N (28%; 5/18) than S (0%; 0/20) cows. Days to first artificial insemination were fewer in N (69.4 d) and S (68.0 d) than in T (75.0 d). First-service conception rate was greater in N (55%; 11/20) than in T (20%; 4/20), with S (26%; 6/23) cows being intermediate. Days open in pregnant cows were fewer in N (93.8 d) than in T (145.4 d), with S (121.2 d) cows being intermediate. Thus, shortening or eliminating the dry period leads to earlier postpartum ovulation and the results highlight the need for future large field studies to accurately evaluate the effect of dry period length on reproductive performance of lactating dairy cows.     

Resynchronizing estrus and ovulation after not-pregnant diagnosis and various ovarian states including cysts

We compared outcomes of 2 protocols used to resynchronize estrus and ovulation in dairy females after a not-pregnant diagnosis. Nulliparous heifers and lactating cows in which artificial insemination (AI) occurred 41 +/- 1 d earlier were presented every 2 to 3 wk for pregnancy diagnosis by using ultrasonography. Ovaries were scanned, follicles were mapped and sized, presence of corpus luteum was noted, and GnRH was injected (d 0). Females were assigned randomly to receive PGF(2alpha) 7 d later (d 7) and then either received estradiol cypionate (ECP) 24 h after PGF(2alpha) (d 8; Heatsynch; n = 230) or a second GnRH injection 48 h after PGF(2alpha) (d 9; Ovsynch; n = 224). Those detected in estrus since their not-pregnant diagnosis were inseminated, whereas the remainder received a timed AI (TAI) between 65 and 74 h after PGF(2alpha). Ovarian scans and blood collected before injections for progesterone analysis were used to classify 4 ovarian status groups: anestrus, follicular cysts, luteal cysts, and cycling, plus an unknown group of females in which no blood sampling or ovarian scans were made. Few females (5.1%) were inseminated between not-pregnant diagnosis and d 8. On d 10, more ECP- than GnRH-treated females were inseminated after detected estrus (24 vs. 6%). Overall, more Ovsynch than Heatsynch females received a TAI (82 vs. 62%). Conception rates tended to be greater for females inseminated after estrus (37%) than after TAI (29%), particularly for those treated with Heatsynch (41 vs. 27%) than with Ovsynch (33 vs. 31%). Those inseminated after estrus conceived 31 +/- 8 d sooner than those receiving the TAI. Conception rates for females having elevated progesterone 7 d after the not-pregnant diagnosis were greater than those having low progesterone in Heatsynch (42%; n = 133 vs. 25%; n = 55) and Ovsynch protocols (33%; n = 142 vs. 15%; n = 45), respectively. Conception rates were greater in nulliparous heifers than in lactating cows (43 vs. 28%) regardless of protocol used. Although overall pregnancy outcomes after a not-pregnant diagnosis were similar in response to either the Ovsynch and Heatsynch protocols, inseminations performed after detected estrus before the scheduled TAI reduced days to eventual conception and tended to increase conception rates, particularly after Heatsynch.     

Ovulation timing and conception risk after automated activity monitoring in lactating dairy cows

Using 1 market-available activity monitor, 3 experiments were conducted in dairy cows to determine timing of ovulation, compare within-herd conception risk of cows inseminated based on activity monitors versus timed artificial insemination (AI), and determine conception risk of cows inseminated at various intervals after achieving an activity threshold. In experiment 1, ovaries were scanned every 3 h by transrectal ultrasonography to determine the time of ovulation beginning 14 ± 0.5 h after the achieved activity threshold (n = 132) or first standing event (n = 59), or both (n = 59). Progesterone at the first ovarian scan (0.1 ± 0.01 ng/mL) and ovarian structures [1 or 2 preovulatory-sized follicles (16.5 ± 0.2 mm)] confirmed that 88.6% of cows identified by activity were in estrus. The remaining 15 cows (11.4%) with a corpus luteum and elevated progesterone concentration (5.3 ± 0.5 ng/mL) were classified as false positives. The average interval from first standing event to ovulation (n = 59) differed slightly from the interval after the achieved threshold (26.4 ± 0.7 vs. 24.6 ± 0.7 h, respectively). In 97 cows fitted with activity monitors, that interval was 25.7 ± 0.4 h. In experiment 2, the conception risk in 394 cows in 1 herd fitted with activity monitors was compared with that of 413 cows submitted to a timed AI program through 3 AI services. Days to first AI were reduced in cows fitted with activity monitors, and conception risk after activity threshold was less than that for timed AI at first service because of differing days in milk at first AI. Both median and mean days to pregnancy, however, were reduced in activity-group cows by 10 and 24 d, respectively, compared with timed AI cows. In experiment 3, 4,019 cows in 19 herds were inseminated after achieving the activity threshold. Conception risk was determined for cows inseminated at various intervals after the achieved activity threshold. A curvilinear conception risk curve peaked at 47.9% for primiparous cows inseminated between 13 and 16 h, whereas conception risk in multiparous cows was steady at 34% through 12 h and decreased thereafter. These experiments demonstrate that time of ovulation after activity threshold closely resembles the time of ovulation after first standing estrus. Time of insemination up to 12 h after the activity threshold produced similar conception risks for multiparous cows, whereas intervals shorter than 13 and greater than 16 h in primiparous cows seemed to compromise their conception risk. Although conception risk may not be improved at individual inseminations after achieving an activity threshold, the rate of achieving pregnancy is hastened. Activity monitors can accurately predict ovulation and time of AI.     

Alternative programs to presynchronize estrous cycles in dairy cattle before a timed artificial insemination program

The objective was to test potential presynchronization programs applied to cows before a timed artificial insemination (TAI) program to increase the percentage of cows ovulating in response to both GnRH injections of a TAI program and having a functional corpus luteum before the first GnRH injection of the TAI program. At calving, cows were blocked by lactation (1 vs. 2+) and assigned randomly to receive 1 of 5 presynchronization treatments. Two variants of the standard Presynch program were tested in which 2 injections of PGF_2α were administered 14 d apart with either 14 d (Pre14; n = 122), 12 d (Pre12; n = 123), or 10 d (Pre10; n = 151) intervening before a TAI program was initiated. Two other presynchronization programs consisted of administering a progesterone-releasing controlled internal drug release (CIDR) insert for 7 d plus PGF_2α administration at insert removal. Insert removal occurred either 10 d (CIDR10; n = 157) or 3 d (CIDR3; n = 117) before a TAI program was initiated. The TAI program was a standard Cosynch program [injection of GnRH 7 d before (GnRH-1) and 72 h after (GnRH-2) PGF_2α with TAI administered 72 h after PGF_2α). Cosynch served as the control (n = 157), and cows were assumed to be starting this program at random stages of the estrous cycle. From a subset of cows per treatment (ranging from 49 to 51 cows each), blood samples were collected from coccygeal vessels by using evacuated tubes at d −28, −14, 0 (onset of TAI program), 7, 9, 14, and 21. Ovarian scans were conducted on d 0, 7, 9, 14, and 21 by transrectal ultrasonography. Diameters of follicles and corpus luteum were measured at each exam, and ovulation was determined on d 7 (response to GnRH-1 on d 0) and d 14 (response to GnRH-2 on d 10). Ovulatory incidence after GnRH-1 (47.1 to 67.3%) and GnRH-2 (78 to 90.2%) varied but did not differ among treatments. Before GnRH-1, progesterone concentrations were less in the CIDR3 treatment than in all other treatments. Before GnRH-2, progesterone was greater in the CIDR3 treatment than in all other treatments. Luteal regression and synchronization rate (successful luteolysis and ovulation after GnRH-2) did not differ among treatments. Pregnancy rate per AI at 32 and 60 d post TAI was less in CIDR3 cows than in cows in all other treatments. None of the Presynch treatments improved key responses (ovulation, luteolysis, and synchronization rate) known to improve fertility compared with a standard Cosynch program without presynchronization.     

Naturally occurring mastitis effects on timing of ovulation, steroid and gonadotrophic hormone concentrations, and follicular and luteal growth in cows

The effects of naturally occurring subclinical chronic or clinical short-term mastitis on time of ovulation, plasma steroid and gonadotropin concentrations, and follicular and luteal dynamics were examined in 73 lactating Holstein cows. Cows were sorted by milk somatic cell count and bacteriological examination into an uninfected group (n = 22), a clinical mastitis group (n = 9; events occurring 20 ± 7 d before the study), and a subclinical chronic mastitis group (n = 42). In addition, uninfected and mastitic cows were further sorted by their estrus to ovulation (E-O) interval. About 30% of mastitic cows (mainly subclinical) manifested an extended E-O interval of 56 ± 9.2 h compared with 28 ± 0.8 h in uninfected cows and 29 ± 0.5 h in the other 70% of mastitic cows. In mastitic cows with extended E-O interval, the concentration of plasma estradiol at onset of estrus was lower than that of uninfected cows or mastitic cows that exhibited normal E-O intervals (3.1 ± 0.4, 5.8 ± 0.5, and 5.5 ± 0.5 pg/mL, respectively). The disruptive effect of mastitis on follicular estradiol probably does not involve alterations in gonadotropin secretion because any depressive effects of mastitis on pulsatile LH concentrations were not detected. Cortisol concentrations did not differ among groups. The preovulatory LH surge in mastitic cows with delayed ovulation varied among individuals, being lower, delayed, or with no surge noted compared with the normal LH surge exhibited by uninfected cows or mastitic cows with normal E-O interval (6.8 ± 0.7 ng/mL). The diameter of the second-wave dominant follicle was larger and the number of medium follicles was smaller in uninfected and subclinical cows with normal intervals compared with subclinical cows with extended intervals (13.4 ± 0.5 vs. 10.9 ± 0.9 mm, and 3.8 ± 0.2 vs. 6.7 ± 0.14 follicles, respectively). Mid-luteal progesterone concentrations were similar in uninfected and mastitic cows. These results indicate for the first time that around 30% of cows with subclinical chronic mastitis exhibit delayed ovulation that is associated with low plasma concentrations of estradiol and a low or delayed preovulatory LH surge.     

Assessment of an accelerometer system for detection of estrus and treatment with gonadotropin-releasing hormone at the time of insemination in lactating dairy cows

Two experiments were conducted to evaluate an accelerometer system (Heatime; SCR Engineers Ltd., Netanya, Israel) to manage reproduction in lactating dairy cows. In experiment 1, lactating Holstein cows (n = 112) were fitted with an accelerometer system and were treated with GnRH followed 7 d later by PGF_2α to synchronize estrus. A total of 89 cows that had a follicle >10 mm in diameter and a functional corpus luteum at the PGF_2α injection that regressed by 48 h after induction of luteolysis were included in the analysis. Overall, 71% of cows were detected in estrus by the accelerometer system and 95% of cows showing estrus ovulated within 7 d after induction of luteolysis. Of the cows not detected in estrus by the accelerometer system, 35% ovulated within 7 d after induction of luteolysis. Duration of estrus activity (mean ± SD) was 16.1 ± 4.7 h and was neither affected by parity nor milk production. Intervals (means ± SD) from induction of luteolysis, onset of activity, peak raw activity, and peak weighted activity to ovulation was 82.2 ± 9.5, 28.7 ± 8.1, 20.4 ± 7.8, and 16.4 ± 7.4 h, respectively, and the interval from AI to ovulation was 7.9 ± 8.7 h, but ranged from −12 to 26 h. In experiment 2, cows were assigned randomly to receive an intramuscular injection of GnRH at artificial insemination (AI) after detection of estrus by the accelerometer system or receive no treatment (control). Nine hundred seventy-nine AI services from 461 cows were analyzed. Treatment with GnRH at AI did not affect fertility at 35 or 65 d after AI, and no interaction was detected between treatment and season or treatment and AI number. Overall, two-thirds of the cows that were considered properly synchronized were inseminated based on the accelerometer system and ovulated after AI. The remaining cows either were not inseminated because they were not detected in estrus or would not have had a chance to conceive to AI because they failed to ovulate after estrus. Furthermore, mean time of AI in relation to ovulation determined by the accelerometer system was acceptable for most of the cows that displayed estrus; however, variability in the duration of estrus and timing of AI in relation to ovulation could lead to poor fertility in some cows. For lactating dairy cows detected in estrus by the accelerometer system, treatment with GnRH at the time of AI without reference to the onset of estrus did not increase fertility.     

Short communication: A positive relationship between the first ovulation postpartum and the increasing ratio of milk yield in the first part of lactation in dairy cows

The aim of the present study was to examine the relationship between characteristics of the lactation curve, on the basis of daily milk yield, and ovulation within 3 wk postpartum as an indicator of early return to luteal activity in dairy cows. Lactation records from 46 lactating Holstein cows between calving and 305 d postpartum were studied. Milk samples were collected twice weekly between d 7 and 100 for later determination of progesterone concentrations. Occurrence of an early first ovulation was determined by an increase in milk progesterone by 3 wk after calving. Milk yield was recorded daily until 305 d postpartum, and average yield was calculated weekly. The lactation curve was characterized by 8 indices on the basis of the weekly average of milk yield as follows: a) first-week milk yield; b) peak milk yield; c) actual 305-d milk yield; d) peak week; e) difference in milk yield between the first week and peak week; f) difference in milk yield between the peak week and last week (43rd week postpartum); g) ratio of increase in milk yield between wk 1 and the week of peak yield; and h) ratio of decline in milk yield between the week of peak yield and the last week. Indices g and h were calculated as linear. The number of cows having ovulated by 3 wk postpartum was 22 (47.8%). The resumption of ovarian cycles with normal luteal phases occurred earlier in ovular cows than in anovular cows (32.0 d vs. 57.1 d). Although total milk yield did not differ between ovular and anovular cows, the ratio of increase in milk yield from the first week to the peak week (index g) in ovular cows was smaller compared with that of anovular cows (1.71 vs. 2.54). In addition, the ratio of increase in milk yield from the first week to the third week postpartum was greater in anovular cows by 3 wk postpartum (ovular = 1.43 ± 0.23 vs. anovular = 2.32 ± 0.29). In conclusion, the present study demonstrates that a greater increasing ratio of milk yield during early lactation may delay resumption of ovarian cycles after parturition. Therefore, this study is the first to demonstrate statistically that a smaller increasing ratio of milk yield (index g) during early lactation may have a beneficial effect on the first ovulation by 3 wk postpartum.     

Peroxiredoxin 1 Controls Ovulation and Ovulated Cumulus–Oocyte Complex Activity through TLR4-Derived ERK1/2 Signaling in Mice

Peroxiredoxins (PRDXs) are expressed in the ovary and during ovulation. PRDX1 activity related to the immuno-like response during ovulation is unknown. We investigated the roles of Prdx1 on TLR4 and ERK1/2 signaling from the ovulated cumulus–oocyte complex (COC) using Prdx1-knockout (K/O) and wild-type (WT) mice. Ovulated COCs were collected 12 and 16 h after pregnant mare serum gonadotropin/hCG injection. PRDX1 protein expression and COC secretion factors (Il-6, Tnfaip6, and Ptgs2) increased 16 h after ovulated COCs of the WT mice were obtained. We treated the ovulated COCs in mice with LPS (0.5 μg/mL) or hyaluronidase (Hya) (10 units/mL) to induce TLR4 activity. Intracellular reactive oxygen species (ROS), cumulus cell apoptosis, PRDX1, TLR4/P38/ERK1/2 protein expression, and COC secretion factors’ mRNA levels increased in LPS- and Hya-treated COCs. The ERK inhibitor (U0126) and Prdx1 siRNA affected TLR4/ERK1/2 expression. The number and cumulus expansion of ovulated COCs by ROS were impaired in Prdx1 K/O mice but not in WT ones. Prdx1 gene deletion induced TLR4/P38/ERK1/2 expression and cumulus expansion genes. These results show the controlling roles of PRDX1 for TLR4/P38/ERK1/2 signaling activity in ovulated mice and the interlink of COCs with ovulation.