Mouse eggs microinjected with physiological concentrations of inositol 1,4,5-trisphosphate (IP₃) do not emit the second polar body, form a pronucleus, or display a fertilization-associated set of changes in the pattern of protein synthesis. IP₃-injected eggs, however, display a conversion of the zona pellucida glycoprotein ZP2 to ZP2_f. The effect is concentration-dependent with an EC₅₀ (effective concentration, 50%) of 5–10 nM and also occurs in the presence of reduced levels of extracellular calcium. The egg-induced zona pellucida modification is not elicited by several other inositol phosphates that are not able to release calcium from intracellular stores in other systems. Analysis of individual eggs microinjected with IP₃ reveals a strong correlation between a reduced binding of sperm to the zona pellucida and the ZP2 to ZP2_f conversion. In addition, solubilized zonae pellucidae isolated from IP₃-injected eggs possess reduced levels of acrosome reaction-inducing activity. These egg-induced modifications of the zona pellucida—reduced sperm receptor and acrosome reaction-inducing activities and the ZP2 to ZP2_f conversion—elicited by microinjected-IP₃ are similar to those that occur following fertilization. Results of these experiments suggest that IP₃ generated in response to fertilization may play a role in the egg-induced modifications of the zona pellucida that result in the polyspermy block.