Intracellular Calcium Oscillations Signal Apoptosis Rather than Activation in In Vitro Aged Mouse Eggs1

Abstract

We have previously demonstrated that initiation of intracellular calcium ([Ca2+]i) oscillations in mouse eggs signals activation or apoptotic death depending on the age of the eggs in which the oscillations are induced. To extend these studies, mouse eggs were aged in vitro to 24, 32, and 40 h post-hCG and injected with sperm cytosolic factor (SF), adenophostin A, or sperm (intracytoplasmic sperm injection), and the times at which signs of apoptosis first appeared were examined. These treatments, which induced [Ca2+]i oscillations, caused fragmentation and other signs of programmed cell death in eggs as early as 32 h post-hCG. The susceptibility of aged eggs to apoptosis appeared to be due to cytoplasmic deficiencies, because fusion of recently ovulated eggs with aged, SF-injected eggs prevented fragmentation. Evaluation of mRNA and protein levels of the apoptotic regulatory proteins Bcl-2 and Bax showed a prominent decrease in the amounts of Bcl-2 mRNA and protein in aged eggs, whereas Bax mRNA levels did not appear to be changed. Lastly, the Ca2+ responses induced by the aforementioned Ca2+ agonists ceased in advance in aged eggs. Together, these results suggest that one or several critical cytosolic molecules involved in the regulation of Ca2+ homeostasis, and in maintaining the equilibrium between anti- and proapoptotic proteins, is either lost or inactivated during postovulatory egg aging, rendering the fertilizing Ca2+ signal into an apoptosis-inducing signal.

Oxidative stress and ageing of the post-ovulatory oocyte

“…As a result of this impaired Ca 2C homoeostasis, aged oocytes exhibit abnormal Ca 2C oscillations at fertilization that have significantly higher frequency and lower amplitude than those of fertilized fresh oocytes (Igarashi et al 1997, Takahashi et al 2003. These abnormal Ca 2C oscillations are purportedly associated with the onset of apoptosis following fertilization, rather than entry into a developmental pathway (Gordo et al 2000(Gordo et al , 2002.…”

“…Not surprisingly, the process of post-ovulatory ageing culminates in apoptosis (Fujino et al 1996) as a result of decreases in expression of the anti-apoptotic protein BCL2 (Gordo et al 2002, Ma et al 2005) and the activation of caspases (Takai et al 2007, Lord et al 2013.…”

The employment of strontium to activate mouse oocytes: effects on spermatid-injection outcome

“…Additionally, the pattern, frequency, and amplitude of early calcium oscillations impact on post-implantation development (Ozil & Huneau 2001). Prevention of calcium oscillations completing their full course during fertilization will retard pronucleus formation (Lawrence et al 1998) and calcium oscillations of abnormal frequencies will induce development arrest (Gordo et al 2002).…”

Molecular alterations during female reproductive aging: Can aged oocytes remind youth?

“…Overexpression of BCL-2 in mouse ovaries leads to decreased follicular apoptosis (Hsu et al 1996). A prominent decrease of BCL-2 was also observed in eggs aged in vitro (Gordo et al 2002). In addition, the upregulation of BIM in cumulus cells seems to accelerate oocyte aging .…”