Cyclic AMP (cAMP) and Ca2+ are two ubiquitous second messenger in transduction pathways downstream of receptors for hormones, neurotransmitters and local signals. The availability of fluorescent Ca2+ dyes that are easily introduced into cells and tissues has facilitated analysis of the dynamics and spatial patterns for Ca2+ signaling pathways. A similar dissection of the role of cAMP has lagged because indicator dyes do not exist. Genetically encoded reporters for cAMP are available but they must be introduced by transient transfection in cell culture, which limits their utility. We report here that we have produced a strain of transgenic mice in which an enhanced cAMP repoter is integrated in the genome and can be expressed in any targeted tissue and with tetracycline induction. We have expressed the cAMP reporter in β-cell of pancreatic islets and conducted an analysis of intracellular cAMP levels in relation to glucose stimulation, Ca2+ levels, and membrane depolarization. Pancreatic function in transgenic mice was normal. In induced transgenic islets, glucose evoked an increase in cAMP in β-cells in a dose-dependent manner. The cAMP response in independent of (in fact, precedes) the Ca2+ indlux that results from glucose stimulation od islets. Glucose-evoked cAMP response are synchronous in cells throughout the islet and occur in 2 phases suggestive of the time course of insulin secretion. Insofar as cAMP in islets is known to ptentiate insulin secretion, the novel transgenic mouse model will for the first time permit detailed analyses of cAMP signals in β-cells within islets, i.e. in their native physiological context. Reporter expression in other tissues (such as the heart) where cAMP plays a critical regulatory role, will permit novel biomedical approaches.
ASJC Scopus subject areas
- Biochemistry, Genetics and Molecular Biology(all)
- Agricultural and Biological Sciences(all)