Changes in Electrical Properties and Ooplasmic Activities of Na⁺, K⁺, H⁺, Ca⁺⁺, and Cl⁻ During Egg Development in the Locust

Abstract

This thesis addresses two hypotheses: 1) ooplasmic ion activity is regulated during water uptake by locust eggs, and, 2) activities of Ca⁺⁺ and H⁺ are maintained at levels appropriate for their use as signals for developmental processes, including activation, in insect eggs. Hypothesis 1 is based on the perturbing effect of large changes in intracellular Na and/or K activity on enzyme function in eukaryotic cells. Hypothesis 2 is based on the ionic hypothesis of activation developed from studies of eggs of marine invertebrates. Electrical potential difference across egg membranes (PD_egg), and ooplasmic activities of Na⁺, K⁺, Cl⁻, Ca⁺⁺ and H⁺ were measured with double-barrelled ion-selective microelectrodes. Locust eggs maintained a measurable potential difference across egg membranes throughout development. Input resistance (Rᵢₙ) decreased by approximately 5-fold in eggs after fertilization suggesting that the chorion is not the major barrier to ion movements into and out from the egg. Chilling and anoxia decreased the contribution of a metabolic component to PD_egg. The effects of hypercapnia on PD_egg and ooplasmic pH suggest that the metabolic component may be a H⁺-pump. Chloride diffusion contributes to PD_egg in eggs after fertilization; contributions of other ions were relatively small. Potential differences measured in internally perfused locust eggs after osmotic lysis of the serosal epithelium indicated that the chloride-dependent component of PD_egg is developed across the chorion and/or serosal cuticle. In vivo values of PD_egg are probably less negative than values measured in control saline because chloride content of ground water is low. Ooplasmic sodium and potassium activities remained at typically intracellular levels during water uptake, possibly due to release from internal stores. Chloride activities were typical of extracellular fluids and were not regulated during water uptake. Measurements of pH and pea in locust eggs are consistent with the increases in these parameters predicted by the ionic hypothesis of activation. calcium activity in the ooplasm of unfertilized locust eggs (pCa 6.4 -4.9) appeared to be at a level appropriate for the use of calcium as a signal or second messenger. Calcium activity increased 100fold within 1 day of fertilization, and 1000-fold by day 3. Calcium entry from external sources at fertilization and release from internal stores later in development may contribute to the progressive increase in ooplasmic Ca⁺⁺ activity. The ooplasm likely alkalinizes after oviposition as ambient pCo₂ declines. Available data suggest a metabolically-dependent proton pump may control ooplasmic pH in locust eggs, in contrast to the Na⁺/H⁺ exchanger implicated in alkalinization of marine invertebrate eggs.