Limited recovery of meiotic spindles in living human oocytes after cooling-rewarming observed using polarized light microscopy

BACKGROUND: Spindles are formed from microtubules and are exquisitely sensi tive to changes in temperature. An orientation-independent polarized light microscope, the Polscope, can be used to image spindles in living oocytes a llowing analysis of spindle kinetics in the living state. This study examin ed the effects of cooling on spindle disassembly in living human oocytes an d spindle recovery after rewarming. METHODS: Oocytes; were imaged continuou sly with the Polscope during cooling and rewarming. The quantity of microtu bules in the spindles was measured by its birefringence using the Polscope. RESULTS: Spindles had completely disassembled by 5 min after cooling and r ecovered by 20 min after rewarming to 37 degreesC if rewarming started soon after the oocyte's temperature dropped to room temperature. However, when oocytes were cooled and kept at 33, 28 or 25 degreesC for 10 min and then w armed, it was found that warming allowed 5/5, 2/5 and 0/5 oocytes of the sp indles to recover respectively. CONCLUSIONS: These results indicate that hu man meiotic spindles are exquisitely sensitive to alterations in temperatur e. The maintenance of temperature at 37 degreesC during in-vitro manipulati on is important for spindle integrity and,:therefore, is likely to be impor tant for normal fertilization and subsequent embryo development.