By P. Michael Conn (Eds.)
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K e t t m a n , G. M a r b a i x , D . Portetelle, and H . C h a n t r e n n e , Proc. Natl Acad. Sci. A. 74, 3230 (1977). 3. L . G e d a m u , G. H . D i x o n , and J. B . G u r d o n , Exp. Cell Res. Ill, 325 (1978). 4. A. C o l m a n , C. D . L a n e , R. Craig, A. Boulton, T. M o h u n , and J. M o r s e r , Eur. J. Biochem. 113, 339 (1981). 5. T. L u n d , R. B r a v o , H . R. J o h a n s e n , J. Z e u t h e n , and J. V u u s t , FEBS Lett. 208, 369 (1986). 6. K. S u m i k a w a , M . H o u g h t o n , J.
1 and 2). ), as well as the voltage-operated channels (Na+, K+, Ca2 +, CI-). This chapter describes the use of Xenopus oocytes for the study of brain neurotransmitter receptors and ion channels. Procedures used for the transplantation of neurontransmitter receptors and voltage-activated ion channels are shown diagrammatically in Fig. 1 and each stage is described in more detail below. Extraction and Purification of mRNA Extraction of Total RNA from the Brain Extraction of total RNA is the most important step in the transplantation, because the quality of the mRNA dictate's the type and number of neurotransmitter receptors and voltage-operated ion chan,nels which are expressed in the oocyte membrane.
Currents of tens of microamperes are sometimes required to charge rapidly the membrane capacitance when stepping the membrane potential to activate voltage-gated channels, and similarly large ligand-gated currents may be induced by potent m R N A preparations. In practice, the tip size of the pipettes is a matter of compromise; the electrical properties of the pipettes improve with increasing size, but more damage is done to the oocyte on impalement. Some donors yield oocytes which can withstand the use of large pipettes, while other oocytes may be very fragile.