Expression and localization of the mu-opioid receptor (MOR) in the equine cumulus-oocyte complex and its involvement in the seasonal regulation of oocyte meiotic competence.
Abstract: The micro-opioid receptor (MOR) was identified in equine oocytes, cumulus and granulosa cells. By RT-PCR, a 441bp fragment was observed. By immunoblotting, a 65 kDa band was detected in samples of winter anestrous whereas in cells recovered in breeding season, two bands, 65 and 50 kDa, were found. The 65 kDa band was significantly more intense in winter anestrous specimens. In samples recovered in the breeding season, this band significantly decreased with the raise of follicle size and was heavier in compact oocytes and cumulus cells. The protein was localized on the oolemma and within the cytoplasm of oocytes and cumulus cells. In vitro oocyte maturation rate (MR), analyzed by confocal microscopy for nuclear chromatin, microfilaments and microtubules, was reduced after the addition of 3 x 10(-8) M beta-endorphin in medium without additional hormones. Inhibitory effects of 10(-3) M Naloxone in oocytes collected in anestrous and spring transition were observed, both in presence and absence of hormones added to culture medium. Increased MRs were observed in oocytes collected in anestrous and cultured in presence of 10(-8) M Naloxone. The exposure to 10(-3) M Naloxone induced significant intracellular calcium increases in cumulus cells recovered all over the year. beta-Endorphin 3 x 10(-8) M induced significant calcium increases only in cumulus cells recovered in fall transition and anestrous. Naloxone 10(-8) M did not induce intracellular calcium modifications. We conclude that the MOR is differentially expressed in equine cumulus-oocyte complexes in the different seasons of the year and plays a role in the seasonal regulation of meiotic competence of equine oocytes.
Publication Date: 2008-01-24 PubMed ID: 18213645DOI: 10.1002/mrd.20869Google Scholar: Lookup
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- Comparative Study
- Journal Article
- Research Support
- Non-U.S. Gov't
Summary
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The research focuses on identifying the presence and influence of the mu-opioid receptor (MOR) in horse oocytes (egg cells), and how it affects the seasonal ability of these eggs to undergo meiosis, a type of cell division.
Introduction to MOR in Equine Oocytes
- The article reveals that the mu-opioid receptor (MOR) has been located in the oocytes (eggs), cumulus, and granulosa cells of horses.
- Through Reverse Transcription Polymerase Chain Reaction (RT-PCR), a 441 base pair segment was identified – a suggestive link to the presence of the MOR.
Identification Techniques
- Immunoblotting, a technique for detecting proteins, was used to further support the identification of the MOR.
- A 65 kiloDalton (kDa) band was observed in the samples taken during the winter anestrous phase (non-breeding season), implying increased expression of the 65kDa MOR protein during this period.
- In cells retrieved in the breeding season, two distinct bands (65 kDa and 50 kDa) were found, the 65kDa band significantly decreased with the growth of follicle size, indicating a possible seasonal expression modulation of MOR.
Localization of the Protein
- The MOR protein was specifically located on the oolemma (outer layer of an egg cell) and within the cytoplasm of the oocytes and cumulus cells, providing a possible regulatory role of MOR in equine oocyte maturation and release.
Impact on Maturation Rate
- The research also examined the effects of the presence of MOR on the maturation rate (MR) of the oocytes, using confocal microscopy a visual characterization tool.
- The MR reduced when beta-endorphin, a natural opioid that binds to MOR, was added to the media in which these oocytes were cultured without any additional hormones, suggesting MOR involvement in negatively regulating oocyte maturation.
- The inhibitory effects of Naloxone, an opioid antagonist that blocks MOR, were observed, which further confirms MOR’s role in equine oocyte maturation.
Role of Calcium Levels
- The study also noticed that the activation of MOR affected intracellular calcium levels within the cumulus cells, a process associated with oocyte maturation and ovulation.
- beta-Endorphin caused significant calcium increases only in cumulus cells collected in fall transition and anestrous, while Naloxone did not induce any intracellular calcium modifications.
Conclusion of the Study
- The study concludes that the MOR is differentially expressed in the equine cumulus-oocyte complex during different seasons of the year and plays a pivotal role in the seasonal regulation of meiotic competence of equine oocytes.
Cite This Article
APA
Dell'Aquila ME, Albrizio M, Guaricci AC, De Santis T, Maritato F, Tremoleda JL, Colenbrander B, Guerra L, Casavola V, Minoia P.
(2008).
Expression and localization of the mu-opioid receptor (MOR) in the equine cumulus-oocyte complex and its involvement in the seasonal regulation of oocyte meiotic competence.
Mol Reprod Dev, 75(8), 1229-1246.
https://doi.org/10.1002/mrd.20869 Publication
Researcher Affiliations
- Department of Animal Production, University of Bari, Valenzano-Bari, Italy. e.dellaquila@veterinaria.uniba.it
MeSH Terms
- Animals
- Blotting, Western
- Calcium / metabolism
- Cumulus Cells / drug effects
- Cumulus Cells / metabolism
- DNA Primers / genetics
- Female
- Fluorescent Antibody Technique
- Horses / metabolism
- Meiosis / genetics
- Meiosis / physiology
- Microscopy, Confocal
- Naloxone / pharmacology
- Oocytes / cytology
- Oocytes / drug effects
- Oocytes / metabolism
- Receptors, Opioid, mu / metabolism
- Reverse Transcriptase Polymerase Chain Reaction
- Seasons
Citations
This article has been cited 5 times.- Albrizio M, Guaricci AC, Milano S, Macrì F, Aiudi G. Mu opioid receptor in spermatozoa, eggs and larvae of gilthead sea bream (Sparus Aurata) and its involvement in stress related to aquaculture.. Fish Physiol Biochem 2014 Aug;40(4):997-1009.
- Ambruosi B, Uranio MF, Sardanelli AM, Pocar P, Martino NA, Paternoster MS, Amati F, Dell'Aquila ME. In vitro acute exposure to DEHP affects oocyte meiotic maturation, energy and oxidative stress parameters in a large animal model.. PLoS One 2011;6(11):e27452.
- Subirán N, Casis L, Irazusta J. Regulation of male fertility by the opioid system.. Mol Med 2011;17(7-8):846-53.
- Albrizio M, Lacalandra GM, Micera E, Guaricci AC, Nicassio M, Zarrilli A. Delta opioid receptor on equine sperm cells: subcellular localization and involvement in sperm motility analyzed by computer assisted sperm analyzer (CASA).. Reprod Biol Endocrinol 2010 Jun 25;8:78.
- Li ZH, Chu N, Shan LD, Gong S, Yin QZ, Jiang XH. Inducible expression of functional mu opioid receptors in murine dendritic cells.. J Neuroimmune Pharmacol 2009 Sep;4(3):359-67.
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