Down-regulation of glucagon receptors on the surface of broiler adipocytes

A radioreceptor assay that specifically measures the affinity and concentration of glucagon receptors on the surface of broiler adipocytes was used to determine whether down-regulation of glucagon receptors is involved in the mechanism whereby glucagon induces desensitization of its ability to acutely stimulate lipolysis. Adipocytes from abdominal fat of 42- to 49-d-old female broilers were preincubated in vitro with 0 to 8 nM glucagon for 0 to 24 h before removal of glucagon by washing and assessment of lipolysis during a subsequent 1 h incubation at 37 C and binding of ([125I]iodo-Tyr10)glucagon during an ensuing 6 h incubation at 12 C. In Experiment 1, pretreatment of adipocytes with 0 to 8 nM glucagon for 24 h reduced lipolysis 70% with a preincubation dose of glucagon (ED50) of 0.4 +/- 0.04 nM, whereas glucagon binding was decreased 75% with an ED50 of 0.7 +/- 0.1 nM. In Experiment 2, pretreatment of adipocytes with 5.4 nM glucagon for 0 to 2 h decreased lipolysis 60% with a time of preincubation with glucagon (T1/2) of < 1 min, whereas glucagon binding was reduced 40% with a T1/2 of 2 +/- 0.6 min. In Experiment 3, detailed analysis of lipolysis and glucagon binding to adipocytes pretreated with 5.4 nM glucagon for 5 min, 2 h, or 24 h revealed that sensitivity of adipocytes to acute stimulation of lipolysis by glucagon and the number of glucagon receptors on the adipocyte surface were decreased by glucagon pretreatment. The assay concentration of glucagon (EC50) for glucagon stimulation of lipolysis, a measure of adipocyte sensitivity, increased 0.8-fold at 5 min, 1.6-fold at 2 h, and 4.9-fold at 24 h of preincubation. The number of low-affinity glucagon receptors decreased 12% at 5 min, 20% at 2 h, and 50% at 24 h of preincubation with glucagon. Throughout this study, decreased glucagon binding was closely associated with reduced lipolysis, implicating down-regulation of cell-surface glucagon receptors in the mechanism whereby glucagon induces desensitization of its ability to acutely stimulate lipolysis in broiler adipocytes.