Our lab has previously demonstrated that AMP-activated protein kinase (AMPK) in mouse oocytes promotes germinal vesicle breakdown (GVB), and meiotic progression to MII. Active AMPK was localized to the germinal vesicle prior to GVB, condensed chromosomes after GVB, spindle poles at metaphase I and II, and the spindle midzone during anaphase. This staining pattern of active AMPK was dependent on microtubule spindle integrity. On the other hand, inhibiting AMPK activity during spontaneous maturation induced spontaneous activation of oocytes. In this study, we further investigated the effects of AMPK activation on the maturation and activation of oocytes. We used the genetically predisposed parthenogenetic activation strains, LT/SvEiJ mice and its related hybrid strain, LT/SvEiJ x C57B/6J (LTBL) to test the effect of AMPK. Oocytes were cultured in either control MEM/BSA or medium supplemented with the AMPK activator, AICAR (0.2 mM), for 17h. There was no sign of PB formation in the control LT/SvEiJ group; however, 29.8% formed PBs in the presence of AICAR. The PB rates for both control and AICAR-treated groups in LTBL oocytes were intermediate between LT/SvEiJ and B6SJL strains. When oocytes from LT/SvEiJ mice were cultured for 30h in either control medium or medium containing AICAR, 22% of the control group oocytes were activated as shown by formation of pronuclei and cleavage of oocytes, and AICAR reduced this number to 1.9%. AICAR had a similar effect on LTBL oocytes,reducing activation from 15.7% to 4%. To determine the critical window of AMPK activity required to convey this suppressive effect on oocyte activation, LT/SvEiJ oocytes were first cultured in either MEM or medium containing AICAR for 17h, then transferred to either MEM or AICAR-supplemented medium for another 13h before of activation was assessed. Oocytes had control levels of activation when they were first cultured in MEM and then transferred to AICAR. In contrast, oocytes showed low levels of activation when they were first cultured in the presence of AICAR even if AICAR was absent during the subsequent 13h. Experiments using B6SJL oocytes showed a similar effect. When superovulated B6SJL oocytes were retrieved from oviducts 16, 20 and 24h post-hCG, 0.5 mM but not 0.2 mM AICAR produced a subtle and transient suppression of activation during 6h of culture. These data indicated an early requirement of AMPK for suppressing premature activation. Immunofluorescent staining of LT/SvEiJ oocytes after 17h of in-vitro culture indicated that oocytes stayed arrested at MI stage. Interestingly, we observed abnormal active AMPK localization in those oocytes. Instead of staining at the both spindle poles, the localization was shifted from the poles in some oocytes and some were missing spindle pole localization. This suggested that the improper functioning of AMPK might also contribute to the defect of LT/SvEiJ oocytes. (poster)