Role of beta3-adrenergic receptors in the action of a tumour lipid mobilizing factor

Abstract

Induction of lipolysis in murine white adipocytes, and stimulation of adenylate cyclase in adipocyte plasma membranes, by a tumour-produced lipid mobilizing factor, was attenuated by low concentrations (10(-7)--10(-5)M) of the specific beta3-adrenoceptor antagonist SR59230A. Lipid mobilizing factor (250 nM) produced comparable increases in intracellular cyclic AMP in CHOK1 cells transfected with the human beta3-adrenoceptor to that obtained with isoprenaline (1 nM). In both cases cyclic AMP production was attenuated by SR59230A confirming that the effect is mediated through a beta3-adrenoceptor. A non-linear regression analysis of binding of lipid mobilizing factor to the beta3-adrenoceptor showed a high affinity binding site with a Kd value 78 +/- 45 nM and a B(max) value (282 +/- 1 fmole mg protein(-1)) comparable with that of other beta3-adrenoceptor agonists. These results suggest that lipid mobilizing factor induces lipolysis through binding to a beta3-adrenoceptor.

Figure 1

(A) Effect of the β3-AR antagonist SR59230A on lipolysis in murine white adipoctes; induced by LMF. Adipocytes were preincubated with the indicated concentration of SR59230A for 30 min prior to the addition of LMF (465 nM). (B) Effect of SR59230A on the stimulation of adenylate cyclase in isolated murine adipocyte plasma membranes by LMF. Membranes were preincubated with the indicated concentrations of SR59230A for 30 min prior to the addition of LMF (2.35 μM) and adenylate cyclase was determined as described in Materials and Methods in the presence of 0.1 μM GTP. The results are expressed as means ±s.d. and the data is representative of three separate experiments. Differences from incubation in the absence of SR59230A is indicated c, P<0.005 and d, P<0.001 as determined by Student's t-test.

Figure 2

Effect of LMF (A) and isoprenaline (B) on cyclic AMP levels in CHOK1β3 cells in the absence (○) or presence of 10 μM propranolol (•) or 10 μM SR59230A (▴) Differences from controls are indicated as b, P<0.01 and d, P<0.001 as determined by ANOVA.