PT  - JOURNAL ARTICLE
AU  - Ayala, Julio E.
AU  - Bracy, Deanna P.
AU  - McGuinness, Owen P.
AU  - Wasserman, David H.
TI  - Considerations in the Design of Hyperinsulinemic-Euglycemic Clamps in the Conscious Mouse
AID  - 10.2337/diabetes.55.02.06.db05-0686
DP  - 2006 Feb 01
TA  - Diabetes
PG  - 390--397
VI  - 55
IP  - 2
4099  - http://diabetes.diabetesjournals.org/content/55/2/390.short
4100  - http://diabetes.diabetesjournals.org/content/55/2/390.full
SO  - Diabetes2006 Feb 01; 55
AB  - Despite increased use of the hyperinsulinemic-euglycemic clamp to study insulin action in mice, the effects of experimental parameters on the results obtained have not been addressed. In our studies, we determined the influences of sampling sites, fasting duration, and insulin delivery on results obtained from clamps in conscious mice. Carotid artery and jugular vein catheters were implanted in C57BL/6J mice (n = 6–10/group) fed a normal diet for sampling and infusions. After a 5-day recovery period, mice underwent a 120-min clamp (2.5-mU · kg−1 · min−1 insulin infusion; ∼120–130 mg/dl glucose) while receiving [3-3H]glucose to determine glucose appearance (endoRa) and disappearance (Rd). Sampling large volumes (∼100 μl) from the cut tail resulted in elevated catecholamines and basal glucose compared with artery sampling. Catecholamines were not elevated when taking small samples (∼ 5 μl) from the cut tail. Overnight (18-h) fasting resulted in greater loss of total body, lean, and fat masses and hepatic glycogen but resulted in enhanced insulin sensitivity compared with 5-h fasting. Compared with a 16-mU/kg insulin prime, a 300-mU/kg prime resulted in hepatic insulin resistance and slower acquisition of steady-state glucose infusion rates (GIR) after a 5-h fast. The steady-state GIR was expedited after the 300-mU/kg prime in 18-h–fasted mice. The GIR and Rd rose with increasing insulin infusions (0.8, 2.5, 4, and 20 mU · kg−1 · min−1), but endoRa was fully suppressed with doses higher than 0.8 mU · kg−1 · min−1. Thus, common variations in experimental factors yield different results and should be considered in designing and interpreting clamps.