In early cleavage stage hamster embryos, the inability to regulate intracellular pH (pH_i) properly is associated with reduced developmental competence in vitro. The disruption of mitochondrial organization is also correlated with reduced development in vitro. To determine the relationship between pH_i and the disruption of cytoplasmic organization, we examined the effects of altering pH_i on hamster embryo development, mitochondrial distribution, and cytoskeletal organization. The weak base trimethylamine was used to increase pH_i and was found to reduce embryo development and disrupt the perinuclear organization of mitochondria. The weak acid 5,5-dimethyl-2,4-oxazolinedione was used to decrease pH_i and was also found to reduce development and disrupt the perinuclear organization of mitochondria. With either treatment, the microfilament organization was perturbed, but the microtubule cytoskeleton was not. However, the temporal progression of the disruption of mitochondrial distribution was more rapid in alkalinized embryos than acidified embryos, as revealed by two-photon imaging of living embryos. Additionally, the disruption of the microfilament network by the two treatments was not identical. The cytoplasmic disruptions observed were not due to acute toxicity of the compounds because embryos recovered developmentally when the treatment compounds were removed. These observations link ionic homeostasis, structural integrity and developmental competence in preimplantation hamster embryos.