Influence of sperm immobilization on onset of Ca2+ oscillations after ICSI

Sperm immobilization prior to intracytoplasmic sperm injection (ICSI) is th ought to be necessary for efficient fertilization, A variety of methods of sperm immobilization (pipetting, squeezing and piezo application) are curre ntly employed in ICSI. The effect of differences in immobilization method o n the timing of initial Ca2+ oscillations of oocytes in ICSI was investigat ed. Motile spermatozoa were immobilized in eosin Y solution using pipetting , squeezing and piezo application, Complete staining of the sperm head was achieved after 220.7, 42.2 and 5.0 s respectively, Oscillations after ICSI were measured fluorometrically for each method. The onset of Ca2+ oscillati ons was observed at 4.8 to 80.4 min after ICSI, Ca2+ oscillations developed earlier with the piezo method (14.4 +/- 6.4 min) than other methods (pipet ting, 43.1 +/- 20.2 min, P < 0.01; squeezing, 18.4 +/- 3.8 min, P = NS), Th e piezo method produced the earliest staining of the sperm head and may hav e caused the most damage to the sperm membrane. A more rapid onset of Ca2oscillations was also observed with the piezo method. The method of sperm i mmobilization may be important for the rapid release of sperm factors that initiate oocyte activation. This study also showed that Ca2+ oscillations d evelop earlier in human oocytes treated by ICSI than indicated in previous reports.