ERK-type MAP kinase activity is required for normal first meiotic (MI) metaphase spindle dynamics and first polar body formation at the MI/MII transition, and for MII arrest until egg activation. MEK and MAPK, however, remain active until meiosis is completed and pronuclei form, but whether MEK/MAPK activity affects MII spindle function during egg activation has been unknown. Polarized light microscopy revealed that the MII spindle rapidly (within approximately 15 min) lost birefringence upon treatment of the egg with U0126, indicating decreased organization at the molecular level upon MEK inhibition. In contrast, birefringence rapidly increased when MPF was inhibited with roscovitine, and this was similar to the increased birefringence previously shown after fertilization or parthenogenetic activation with Sr(2+). Confocal microscopy indicated that many spindles in U0126-activated eggs had failed to rotate or were dissociated from the egg cortex. Subsequently, abnormally-located midbodies were evident in U0126-induced parthenogenotes. Thus, MEK/MAPK activity is required to maintain the ordered structure of the MII spindle and for normal spindle dynamics during second polar body formation.