The Preparation And Establishment Of Beta Arrestins Transgenic Mice And The Preliminary Research Of Their Functions

In the field of signal transduction mediated by G protein-coupled receptors(GPCRs, 7TMRs), the classical theory is that β—arrestins uncouple the activated receptors from G proteins (desensitization), prevent further signaling and stimulate the endocytosis of receptors. It has been well established that β — arrestins not only terminate the GPCRs signals, but also mediate endocytosis and transportation of GPCRs. They also initiate and direct new signals, mediate the crosstalk between GPCRs and other signal passways. In order to discover the new functions of β—arrestins in vivo, we apply the transgene technology into my field. First, vectors where there are transgenes are constructed in vitro. Then, they are microinjected into pronucleus embryos. After that, the offsprings are screened by PCR and Southern Blot. Finally, we setup four kinds of transgenic mice, those are human β arrestin Ⅰ overexpression mice, human β arrestin Ⅱ overexpression mice, mouse β arrestin Ⅰ RNAi mice and mouse β arrestin Ⅱ RNAi mice. And the homozygous mice are screened by realtime PCR. So the transgenic mice system is established elementarily. Then the transgene expression is checked by RT-PCR and immunohistochemical technique. Opioid receptors belong to the large superfamilyof GPCRs. So overexpression of p—arrestins in vivo probably influences the drug addiction. We study the morphine tolerance in the transgenic mice by tail flick test. Because of the expression of transgenes in hippocampus, we try to discover whether it influences the spatial memory by open field behavior and Morris water maze test.