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. Author manuscript; available in PMC: 2014 Apr 22.
Published in final edited form as: Reproduction. 2009 Aug 21;138(5):759–770. doi: 10.1530/REP-08-0535

Figure 3.

Figure 3

Effects of forskolin pulsing on oocyte maturation and cAMP and AMP concentrations in hypoxanthine (HX)-arrested oocytes. (A) Forskolin dose response. Hypoxanthine-arrested DO were pulsed 3 h with increasing concentrations of forskolin, washed free of forskolin and returned to HX-containing medium; alternatively, they were cultured continuously in HX alone or HX plus 100 μM forskolin. GVB was assessed after a total of 21-22 h. (B) Maturation kinetics. After 3 h pretreatment with HX plus 100 μM forskolin, DO were washed and cultured in HX alone. Meiotic resumption was determined at the indicated culture times after pulsing. Maturation kinetics were also determined for oocytes continuously cultured in HX alone or HX plus 100 μM forskolin (Fors). Time 0 h represents 3 h of culture. (C) cAMP assay. Fresh DO were collected (time 0) or DO were cultured in 4 mM HX-containing medium in the presence (Fors pulse) or absence (HX) of 100 μM forskolin. Oocytes in the HX group were collected after 3, 5 and 8 h of culture. Oocytes in the Fors pulse group were washed free of forskolin after 3 h and returned to HX-containing medium. Oocytes from this group were collected immediately after the forskolin pulse or 2 or 5 h after washing out the forskolin and subsequent culture in HX (3, 5 and 8 total h of culture, respectively). An additional group of oocytes was continuously cultured in HX plus forskolin for the entire 8-h period (Cont Fors). Only GV-stage oocytes were assayed for cAMP. Results shown here represent the mean ± SEM of 7 experiments. Groups with no common letter are significantly different. (D) AMP assay. Fresh oocytes were collected or DO were cultured in medium containing HX in the presence or absence of forskolin. Oocytes treated with HX alone were collected after 3 and 5 h of culture. Oocytes treated with HX plus forskolin were collected 3 h after forskolin pulsing and 2 h after pulsed oocytes were washed free of forskolin and returned to HX medium (5 h total culture time). Groups with no common letter are significantly different.