PARENT SESSION
PLATFORM SESSION 4. GAMETE BIOLOGY AND GAMETOGENESIS I

Sunday, August 1, 2004
3:00 PM–5:00 PM
Buchanan A104
Chairs: Leslie Heckert Co-Chair: Jason Swain

(26) THE RELATIONSHIP BETWEEN CUMULUS EXPANSION AND THE ALLOCATION OF OOCYTE -TUBULIN RESOURCES IN THE MOUSE OOCYTE.

Barrett, Susan^{1, 2}, Albertini, David¹, ¹ Department of Anatomy and Cellular Biology, Boston, MA² Program in Cellular, Molecular, and Developmental Biology, Boston, MA

ABSTRACT- How physiological changes elicited in the cumulus oocyte complex during ovulation impart developmental competence on the oocyte remains an unexplored area of study. These experiments were designed to test the role of the cumulus during in vivo (IVO) and in vitro maturation (IVM) in the mouse with respect to -tubulin distribution and function and during subsequent activation of M2 arrested oocytes. Oocytes were collected from superovulated mice (eCG, hCG) (IVO) or were in vitro-matured oocytes under conditions that support (supplemented IVM containing fetal bovine serum, D-glucosamine, and human recombinant FSH) or do not support (basal IVM lacking FBS, GlcNAc, and FSH) cumulus expansion. Samples were processed for quantitative analysis of -tubulin content using immunofluorescence and digital imaging. Nucleation competence of MTOCs was assessed by dual labeling with -tubulin and / tubulin specific probes. Under conditions that permit meiotic progression to M2, but not cumulus expansion (basal IVM), oocytes exhibit -tubulin throughout the spindle, a reduced MTOC number and an increased MTOC size and nucleation capacity even though -tubulin content was low on a per MTOC basis. In contrast, IVO and supplemented IVM oocytes show diminished -tubulin spindle content restricted to the poles and an increased MTOC number and staining intensity within compacted foci. Nucleation from MTOCs in supplemented IVM and IVO oocytes requires an activation stimulus and examination 3 hours post SrCl₂, showed synchronous progression to telophase of meiosis 2 and ongoing polar body extrusion. In contrast, within 2 hours after SrCl₂, treatment, basal IVM oocytes have completed meiosis and progressed into interphase as evidenced by the formation of pronuclei and cytoplasmic microtubule arrays. These results are consistent with the idea that under conditions that support cumulus expansion and oocyte maturation, signaling via the cumulus influences the retention of cortical -tubulin in the oocyte and regulates timely cycles of microtubule nucleation required for the completion of meiosis and egg activation.

KEY WORDS: gamma tubulin, cumulus, MTOC, signaling