Xenopus system properties

Characterization of the properties of LGIC receptors expressed in Xenopus oocytes has heen facihtated hy the variety of studies that can be carried out on recombinant receptors in this system. 

CHO cells, using an inducible expression system [100], and in HEK cells [97]. While the recombinant channels show the expected pharmacology, some of the biophysical properties differ from native TTX-resistant currents, suggesting the existence of auxiliary subunits. In Xenopus oocytes and in mammalian cells, expression of rNayl.8 was increased substantially by co-expression with Navbl and Navb3, respectively [98,101]. 

Several studies employing oocytes of the clawed frog, Xenopus laevis, for the in vitro translation of sodium channel encoding mRNAs (53-55) suggest that this experimental system may be particularly useful toward this end. The biophysical properties of sodium channels expressed in oocytes following injection of rat brain mRNA were similar to those of sodium channels in their native membrane environment, and were specifically inhibited by the sodium channel blockers tetrodotoxin and saxitoxin (i5.). Sodium channels encoded by mRNAs from rat skeletal muscle and eel electroplax have also been expressed in Xenopus oocytes (56-57). To date the expression of insect sodium channels in the Xenopus oocyte has not been reported, but the utility of this system for the translation and expression of insect acetylcholine receptor mRNA has recently been demonstrated (58). Successful application of this methodology to the expression of insect mRNAs encoding functional sodium channels offers a novel method to test some of the hypotheses for the molecular basis of the kdr mechanism. 

When Urel is expressed in Xenopus oocytes, there is a large increase in urea uptake that is pH dependent. There is no increase at neutral pH but a large increase with acidification down to pH 4.0, the limit of survival of oocytes. The pH-transport curve can be overlaid on the pH-urease activity curve of intact bacteria. Half-maximal transport occurs at a medium pH of 5.9. The increased uptake is energy independent, temperature independent, and nonsaturable, properties indicating that Urel is a urea channel. This allows extremely fast transport of urea across the inner membrane of the bacteria. Complementation of Urel deletion mutants with wild-type Urel in H. pylori also restore urease activation at acidic pH, showing that Urel is a urea transporter in both the heterologous oocyte expression system and in the native organism. 

---