Effect of blood plasma collected after adrenocorticotropic hormone administration during the preovulatory period in the sow on oocyte in vitro maturation

Abstract

Reproduction may be affected by stressful events changing the female endocrine or metabolic profile. An altered environment during oocyte development could influence the delicate process of oocyte maturation. Here, the effect of simulated stress by media supplementation with blood plasma from sows after adrenocorticotropic hormone (ACTH) administration during the preovulatory period was assessed. Oocytes were matured for 46 h in the presence of plasma from ACTH-treated sows, or plasma from NaCl-treated control sows, or medium without plasma (BSA group). The plasma used had been collected at 36 h and 12 h (± 2 h) before ovulation (for the first 24 h + last 22 h of maturation, respectively).Subsequent fertilisation and embryo development were evaluated. Actin cytoskeleton and mitochondrial patterns were studied by confocal microscopy both in the oocytes and the resulting blastocysts. Nuclear maturation did not differ between treatments. Subtle differences were observed in the actin microfilaments in oocytes; however, mitochondrial patterns were associated with the treatment (P<0.001). These differences in mitochondrial patterns were not reflected by in vitro outcomes, which were similar in all groups. In conclusion, an altered hormonal environment provided by a brief exposure to plasma from ACTH-treated sows during in vitro oocyte maturation, could induce alterations in actin cytoskeleton and mitochondrial patterns in oocytes. However, these changes might not hamper the subsequent in vitro embryo development.[217 words]

Blood plasma collected after adrenocorticotropic hormone administration during the preovulatory period in the sow negatively affects in vitro fertilization by disturbing spermatozoa function

“…However, it is not possible to exclude completely an effect on the oocyte because the washing of the oocytes may have eliminated the potential detrimental effect of the altered environment, and the exposure to the treatments was performed for 1 hour in comparison with the 24 hours of coincubation of gamete in the first experiment. Nonetheless, the exposure of oocytes to blood plasma from ACTH-treated sows during in vitro oocyte maturation (for 46 hours) did not have an impact on the ability of the oocytes to be fertilized or progress to the blastocyst stage [14]. To determine if the altered milieu provided by the plasma collected from the simulated-stress group may have had an impact on sperm function, boar spermatozoa were exposed to the same conditions evaluated in experiments 1 and 2.…”

“…Cumulusoocyte complexes were collected by follicle puncture from 3 to 6 mm follicles as described [14,16]. Maturation medium consisted of TCM-199 (Tissue culture medium-199; M-2154) supplemented with 1 mM L-glutamine (G-8540), 1 mM sodium pyruvate (P-3662), 1 mg/mL BSA (A-3311), 100 mM 2b-mercapto-ethanol (M-7522), 50 mg/ mL gentamicin, 50 ng/mL epidermal growth factor (E-4127), 5 mL/mL insulin-transferrin-selenite (I-3146), 10 IU/ mL pregnant mare's serum gonadotropin, and 5 IU/mL hCG (Suigonan; Intervet Scandinavia, Skovlunde, Denmark).…”

“…Spermatozoa were washed by standard centrifugation. Briefly, 0.5 mL of semen was transferred to a 15 mL tube containing 4 mL of noncapacitating medium [14,16] and centrifuged (300Â g for 5 minutes). The supernatant was discarded and the pellet was washed again under the same conditions.…”

“…After 44 hours of maturation, oocytes were gently transferred to IVF medium and rinsed twice before being transferred to a 4-well culture dish containing a final volume of 500 mL IVF medium [14,16]. The IVF medium composition was 90 mM NaCl, 12 mM KCl, 0.5 mM NaH 2 PO 4 (S-5011), 25 mM NaHCO 3 , 0.5 mM MgSO 4 , supplemented with 2 mM sodium pyruvate, 8 mM CaCl 2 $2H 2 O (C-7902), 6 mM lactic acid (L-7900), 6 mg/mL BSA, 1.9 mM caffeine (C-0750), 50 mg/mL gentamicin, and 5 mg/mL phenol red.…”

“…However, it was not possible to address the potential influence of simulated stress mimicked by ACTH administration directly at the gamete level in the aforementioned in vivo studies. We have recently determined that an altered hormonal environment provided by a brief exposure to plasma from ACTH-treated sows during in vitro oocyte maturation could induce alterations in actin cytoskeleton and mitochondrial patterns in oocytes, but these changes might not hamper the subsequent in vitro embryo development [14]. However, the likely impact of an altered hormonal environment derived from ACTH administration during fertilization has not been analyzed yet.…”