In vitro ageing of porcine oocytes: changes in phosphorylation of the mitogen-activated protein kinase (MAPK) and parthenogenetic activability.

The role of mitogen-activated protein kinase (MAPK) was investigated during ageing of porcine oocytes following in vitro maturation (IVM). Oocytes exhibiting an extruded first polar body after IVM for 46 h (79.3% metaphase II, M II) were used for the experiments. Nuclear maturation stages were not visibly altered after a further 12 h of ageing. Proportion of M II stages (42.9%) decreased significantly whereas fragmentation and degeneration of oocytes increased after an ageing time of 26 h. In vitro ageing for 12 and 26 h led to a significant reduction of MAPK phosphorylation (i.e. activation) compared to oocytes matured for 46 h. When MAPK was inhibited by U0126 in M II oocytes, 30.9% (12 h) and 39.7% (26 h) of oocytes, respectively, left metaphase II arrest and proceeded to early anaphase II. Pronuclear stages or fragmentation could be observed only sporadically (2.6-3.6%). After parthenogenetic activation of oocytes by ethanol/cycloheximide, cleavage stages were reached with rates of 51.9% (46 h IVM), 42.0% (12 h ageing) and 40.3% (26 h ageing), respectively. Furthermore, a significant higher proportion of long-term aged oocytes (26 h) showed pronuclear formation (8.6%) and fragmentation (7.9%) compared to non-aged oocytes (each 1.9%). It is concluded that both MAPK phosphorylation and cleavage rate after parthenogenetic activation decreased before alterations of nuclear stages could be detected during in vitro ageing of M II oocytes. A premature MAPK dephosphorylation of M II oocytes caused early anaphase II stages, but cleaved stages could not be achieved.