The oocyte nucleus isolated in oil retains in vivo structure and functions.

We describe a technique for isolating the nucleus of the giant amphibian oocyte under paraffin oil. The method precludes the losses of small solutes and proteins that accompany isolation of nuclei into aqueous media. An individual oocyte is blotted, placed under oil, punctured near the animal pole and then squeezed to gently extrude the nucleus into the oil, thereby avoiding exposure to any aqueous environment. Light and electron microscopy of the oil-isolated nucleus demonstrate that its in vivo morphology is preserved. We also describe techniques that facilitate the study of nuclear functions under oil. Oil-isolated oocyte nuclei retain many in vivo functions for several hours, including size-selective envelope permeability, RNA synthesis and the ability to break down in response to cdc2/cyclin meiotic maturation promoting factor.

Gonadotropin stimulates oocyte translation by increasing magnesium activity through intracellular potassium-magnesium exchange.

We previously showed that gonadotropin increases the K+ activity in Xenopus oocytes and that this is a signal for increased translation. However, K+ need not act to control synthesis directly but may act through an unidentified downstream effector. Using microinjection to vary the salt content of oocytes and concomitantly measuring [3H]leucine incorporation, we found that small changes in Mg2+ greatly affect translation rates. (Ca2+ had little influence.) By measuring intracellular ion activities, we found that oocyte cations existed in a buffer-like (ion-exchange) equilibrium in which K+ and Mg2+ are the preponderant monovalent and divalent cations. Hence, increasing cellular K+ activity might increase translation by causing Mg2+ activity to rise. If so, the increased translation rates produced by hormone treatment or K+ injection would be prevented by EDTA, a Mg2+ chelating agent. This prediction was tested and confirmed. We conclude that, when gonadotropin increases K+ activity, the cell's internal ion-exchange equilibrium is altered thereby increasing Mg2+ activity and this up-regulates translation.