title: A method for mouse pancreatic islet isolation and intracellular cAMP determination.
abstract: Uncontrolled glycemia is a hallmark of diabetes mellitus and promotes morbidities like neuropathy, nephropathy, and retinopathy. With the increasing prevalence of diabetes, both immune-mediated type 1 and obesity-linked type 2, studies aimed at delineating diabetes pathophysiology and therapeutic mechanisms are of critical importance. The β-cells of the pancreatic islets of Langerhans are responsible for appropriately secreting insulin in response to elevated blood glucose concentrations. In addition to glucose and other nutrients, the β-cells are also stimulated by specific hormones, termed incretins, which are secreted from the gut in response to a meal and act on β-cell receptors that increase the production of intracellular cyclic adenosine monophosphate (cAMP). Decreased β-cell function, mass, and incretin responsiveness are well-understood to contribute to the pathophysiology of type 2 diabetes, and are also being increasingly linked with type 1 diabetes. The present mouse islet isolation and cAMP determination protocol can be a tool to help delineate mechanisms promoting disease progression and therapeutic interventions, particularly those that are mediated by the incretin receptors or related receptors that act through modulation of intracellular cAMP production. While only cAMP measurements will be described, the described islet isolation protocol creates a clean preparation that also allows for many other downstream applications, including glucose stimulated insulin secretion, [3(H)]-thymidine incorporation, protein abundance, and mRNA expression.
authors.affiliation: Department of Nutrional Sciences, University of Wisconsin-Madison.
authors.forename: Joshua C
authors.initials: JC
authors.lastname: Neuman
authors.affiliation: Department of Medicine, Division of Endocrinology, Diabetes, and Metabolism, University of Wisconsin-Madison.
authors.forename: Nathan A
authors.initials: NA
authors.lastname: Truchan
authors.affiliation: School of Pharmacy, University of Waterloo.
authors.forename: Jamie W
authors.initials: JW
authors.lastname: Joseph
authors.affiliation: Department of Medicine, Division of Endocrinology, Diabetes, and Metabolism, University of Wisconsin-Madison; mkimple@medicine.wisc.edu.
authors.forename: Michelle E
authors.initials: ME
authors.lastname: Kimple
mesh_terms: D000818:Animals; D002469:Cell Separation / Q000379:methods*; D000242:Cyclic AMP / Q000032:analysis* / Q000378:metabolism; D007124:Immunoenzyme Techniques / Q000379:methods*; D007515:Islets of Langerhans / Q000737:chemistry* / Q000166:cytology* / Q000378:metabolism; D051379:Mice
chemical_list: D000242:Cyclic AMP