title: Dynamics of glucagon secretion in mice and rats revealed using a validated sandwich ELISA for small sample volumes.
abstract: Glucagon is a metabolically important hormone, but many aspects of its physiology remain obscure, because glucagon secretion is difficult to measure in mice and rats due to methodological inadequacies. Here, we introduce and validate a low-volume, enzyme-linked immunosorbent glucagon assay according to current analytical guidelines, including tests of sensitivity, specificity, and accuracy, and compare it, using the Bland-Altman algorithm and size-exclusion chromatography, with three other widely cited assays. After demonstrating adequate performance of the assay, we measured glucagon secretion in response to intravenous glucose and arginine in anesthetized mice (isoflurane) and rats (Hypnorm/midazolam). Glucose caused a long-lasting suppression to very low values (1-2 pmol/l) within 2 min in both species. Arginine stimulated secretion 8- to 10-fold in both species, peaking at 1-2 min and returning to basal levels at 6 min (mice) and 12 min (rats). d-Mannitol (osmotic control) was without effect. Ketamine/xylazine anesthesia in mice strongly attenuated (P < 0.01) α-cell responses. Chromatography of pooled plasma samples confirmed the accuracy of the assay. In conclusion, dynamic analysis of glucagon secretion in rats and mice with the novel accurate sandwich enzyme-linked immunosorbent assay revealed extremely rapid and short-lived responses to arginine and rapid and profound suppression by glucose.
authors.affiliation: Faculty of Health and Medical Sciences, Department of Biomedical Sciences, University of Copenhagen, Copenhagen, Denmark; and Faculty of Health and Medical Sciences, Novo Nordisk Foundation Center for Basic Metabolic Research, University of Copenhagen, Copenhagen, Denmark.
authors.forename: Nicolai J
authors.initials: NJ
authors.lastname: Wewer Albrechtsen
authors.affiliation: Faculty of Health and Medical Sciences, Department of Biomedical Sciences, University of Copenhagen, Copenhagen, Denmark; and Faculty of Health and Medical Sciences, Novo Nordisk Foundation Center for Basic Metabolic Research, University of Copenhagen, Copenhagen, Denmark.
authors.forename: Rune E
authors.initials: RE
authors.lastname: Kuhre
authors.affiliation: Faculty of Health and Medical Sciences, Department of Biomedical Sciences, University of Copenhagen, Copenhagen, Denmark; and Faculty of Health and Medical Sciences, Novo Nordisk Foundation Center for Basic Metabolic Research, University of Copenhagen, Copenhagen, Denmark.
authors.forename: Johanne A
authors.initials: JA
authors.lastname: Windeløv
authors.affiliation: Faculty of Health and Medical Sciences, Department of Biomedical Sciences, University of Copenhagen, Copenhagen, Denmark; and Faculty of Health and Medical Sciences, Novo Nordisk Foundation Center for Basic Metabolic Research, University of Copenhagen, Copenhagen, Denmark.
authors.forename: Anne
authors.initials: A
authors.lastname: Ørgaard
authors.affiliation: Faculty of Health and Medical Sciences, Department of Biomedical Sciences, University of Copenhagen, Copenhagen, Denmark; and Faculty of Health and Medical Sciences, Novo Nordisk Foundation Center for Basic Metabolic Research, University of Copenhagen, Copenhagen, Denmark.
authors.forename: Carolyn F
authors.initials: CF
authors.lastname: Deacon
authors.affiliation: Faculty of Health and Medical Sciences, Department of Biomedical Sciences, University of Copenhagen, Copenhagen, Denmark; and.
authors.forename: Hannelouise
authors.initials: H
authors.lastname: Kissow
authors.affiliation: Faculty of Health and Medical Sciences, Department of Biomedical Sciences, University of Copenhagen, Copenhagen, Denmark; and Faculty of Health and Medical Sciences, Novo Nordisk Foundation Center for Basic Metabolic Research, University of Copenhagen, Copenhagen, Denmark.
authors.forename: Bolette
authors.initials: B
authors.lastname: Hartmann
authors.affiliation: Faculty of Health and Medical Sciences, Department of Biomedical Sciences, University of Copenhagen, Copenhagen, Denmark; and Faculty of Health and Medical Sciences, Novo Nordisk Foundation Center for Basic Metabolic Research, University of Copenhagen, Copenhagen, Denmark jjholst@sund.ku.dk.
authors.forename: Jens J
authors.initials: JJ
authors.lastname: Holst
mesh_terms: D000778:Anesthetics, Dissociative / Q000494:pharmacology; D000818:Animals; D001120:Arginine / Q000494:pharmacology; D002850:Chromatography, Gel; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D005934:Glucagon / Q000032:analysis / Q000187:drug effects / Q000378:metabolism*; D050416:Glucagon-Secreting Cells / Q000187:drug effects / Q000378:metabolism*; D005947:Glucose / Q000494:pharmacology; D006993:Hypnotics and Sedatives / Q000494:pharmacology; D007649:Ketamine / Q000494:pharmacology; D008297:Male; D008353:Mannitol / Q000494:pharmacology; D051379:Mice; D008810:Mice, Inbred C57BL; D051381:Rats; D017208:Rats, Wistar; D013549:Sweetening Agents / Q000494:pharmacology; D014991:Xylazine / Q000494:pharmacology
chemical_list: D000778:Anesthetics, Dissociative; D006993:Hypnotics and Sedatives; D013549:Sweetening Agents; D014991:Xylazine; D008353:Mannitol; D007649:Ketamine; D005934:Glucagon; D001120:Arginine; D005947:Glucose
keywords: enzyme-linked immunosorbent assay; immunoassay; islet biology; α-cell