Assisted Oocyte ActivationUsing Ionomycin does not Lead to Subsequent Normal CalciumOscillation Pattern

Heytens E* *, Vanhoutte L , Lierman S , Cuvelier C , Gerris J , Dhont M , De Sutter P , Leybaert L ,


Introduction: At fertilization sperm triggers oscillations in the cytoplasmic calcium concentration ([Ca+2]i), that are necessary for oocyte activation and normal further embryo development. Artificially raising [Ca+2]i in oocytes injected with round-headed sperm using a Ca+2 ionophore is however sufficient to achieve normal fertilization and embryo development as evidenced by successful human pregnancies and births. However, it is not known yet if assisted oocyte activation can induce the calcium oscillations normally observed after fertilization. Materials and Methods: Mature mouse oocytes collected were loaded with fluo3-AM and injected with round-headed sperm from the consenting patient or sperm from a consenting proven fertile man. Thirty minutes after injection oocytes injected with roundheaded sperm were incubated with the Ca2+ ionophore ionomycin (10μM) for 10 minutes. Changes of fluorescent intensity were recorded immediately after treatment. Extrusion of the second polar body (PB2) and two-cell formation were assessed. Results: None of the oocytes injected with round-headed sperm were activated (0/12) and none of them displayed [Ca2+]i changes. After treatment with ionomycin more than half (42/59) of the round-headed sperm injected eggs were activated and extruded PB2 2 hours later. Ionomycin treatment was associated with a transient [Ca+2]i rise but was not followed by any oscillatory activity over the recording period. By contrast, in almost all of the 19/38 oocytes injected with fertile sperm that extruded PB2, [Ca+2]i oscillations were observed. Conclusion: A single transient rise in [Ca+2]i triggered by ionophore treatment appears to be sufficient to bring about activation of roundheaded sperm injected oocytes, but this does not lead to subsequent [Ca+2]i oscillations. Successful clinical application in the human, however, suggests that this does not seem to prevent the further embryonic development. These experiments thus bring up the question on the role of [Ca+2]i oscillations on embryo development after ICSI with round-headed sperm.