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Novel methods of egg activation in human assisted reproductive technologies and animal somatic cell nuclear transfer are likely to alter the signalling process that occurs during normal fertilization. Intracytoplasmic sperm injection (ICSI) bypasses the normal processes of the acrosome reaction, sperm-egg fusion, and processing of the sperm plasma membrane, as well as alters some parameters of intracellular calcium ([Ca 2+ ] i ) dynamics (reported previously by Kurokawa and Fissore (2003)). Herein, we extend these studies to determine if ICSI alters the activity of the Ca 2+ -dependent protein, Ca 2+ /calmodulin-dependent kinase II (CaMKII), which is responsible for the completion of meiosis in vertebrate eggs. After ICSI or in vitro fertilization (IVF), individual mouse eggs were monitored for their relative changes in both [Ca 2+ ] i and CaMKII activity during the first [Ca 2+ ] i rise and a subsequent rise associated with second polar body extrusion. The duration of the first [Ca 2+ ] i rise was greater in ICSI than in IVF, but the amplitude of the rise was transiently higher for IVF than ICSI. However, a similar mean CaMKII activity was observed in both procedures. During polar body extrusion, the amplitude and duration of the Ca 2+ rises were increased by a small amount in ICSI compared with IVF, whereas the CaMKII activities were similar. Thus, compared with IVF, ICSI is not associated with decreased or delayed CaMKII activity in response to these Ca 2+ signals in the mouse.

Successful intracytoplasmic sperm injection (ICSI) is dependent upon the competence of the oocyte to respond to the injection of a spermatozoon in the presence of calcium. This study determined if oocytes that failed to become fertilized (absence of any pronuclei) failed to undergo cytoplasmic activation, as ascertained by an electrophoretic shift in the zona pellucida (zona) protein, huZP2. Of 48 zonae individually analysed, 58% did not have a detectable huZP2 shift. Three patterns were observed. In seven patients, none of the unfertilized oocyte huZP2 shifted (16 out of 48); in two, all zonae exhibited a huZP2 shift (five out of 48); in eight, there was a mixture of shifted and unshifted (27 out of 48) in each case. There was no clear relationship between the presence of the huZP2 shift and maternal age, pregnancy outcome, male-factor infertility, or fertilization rate. However, in six couples diagnosed as having no detectable male-factor infertility, 73% (11 out of 15) of the zonae had unshifted huZP2, suggesting that an oocyte defect is involved in some cases of failed fertilization after ICSI. One likely cause for the absence of the huZP2 shift is a failure of cytoplasmic activation. Since oocytes were injected at metaphase II, we hypothesize that some oocytes that failed to fertilize have completed nuclear (meiotic) without cytoplasmic maturation.