Measurement of second messengers in signal transduction: cAMP and inositol phosphates.
cAMP acts as an intracellular mediator of hormone action and the importance of accurate quantitative determination of cAMP levels in cells and tissues is widely recognized. The most utilized procedures for the determination of adenylate cyclase activity in membranes are described here for measuring the conversion of [alpha-(32)P]ATP into [(32)P]cAMP after a two-step chromatographic separation. Also critical in signal transduction is phosphoinositide turnover, which is linked to receptor activation resulting from changes in cytosolic calcium concentrations. Phosphoinositide turnover can be measured as described in this unit by labeling phospholipid pools with [(3)H]-inositol and then analyzing for tritiated inositol phosphates.
Gusovsky
F
F
Eisai Research Institute, Andover, Massachusetts, USA.
D015056:1-Methyl-3-isobutylxanthine / Q000494:pharmacology; D000818:Animals; D002478:Cells, Cultured / Q000737:chemistry; D002852:Chromatography, Ion Exchange / Q000379:methods; D000242:Cyclic AMP / Q000032:analysis*; D006168:Guinea Pigs; D007295:Inositol Phosphates / Q000032:analysis*; D008297:Male; D010743:Phospholipids / Q000378:metabolism; D051381:Rats; D015290:Second Messenger Systems; D013574:Synaptosomes / Q000737:chemistry; D014316:Tritium / Q000032:analysis
D007295:Inositol Phosphates; D010743:Phospholipids; D014316:Tritium; D000242:Cyclic AMP; D015056:1-Methyl-3-isobutylxanthine