Br. J. clin. Pharmac. (1990), 30, 139S-141S
Effects of insulin on human ,3-adrenoceptors G. SAGER Department of Pharmacology, Institute of Medical Biology, University of Troms0, P.O. Box 977, N-9001 Troms0, Norway
The effect of insulin on j-adrenoceptor stimulation and binding in mononuclear leucocytes from healthy subjects was studied. After 1 min exposure to insulin, the sensitivity and maximum response to (-)-isoprenaline and the number of ,-adrenoceptors increased. After 35 min exposure to insulin, the sensitivity and maximal response to (-)-isoprenaline stimulation were lower than the pretreatment values. The ,-adrenoceptor density was similar to the control value after 35 min exposure to insulin. The present observations may account for some of the cardiovascular effects of insulin.
Keywords insulin P-adrenoceptors cyclic AMP
Introduction
Methods
Several observations suggest that insulin plays an important regulatory role in the cardiovascular system. It is well known that the administration of insulin is associated with cardiovascular responses which include increases in heart rate (Page et al., 1976) and in peripheral blood flow (Creager et al., 1985). Insulin increases the myocardial contractility in healthy individuals and in patients with chronic congestive heart failure (Majid et al., 1972). The resultant cardiac effect of insulin is an increase of cardiac output (Arner et al., 1963). The cardiovascular effects of insulin can in part be ascribed to the elevation of plasma catecholamines in hypoglycaemia (Christensen et al., 1975), but also other adrenergic mechanisms exist (Creager et al., 1985). The aim of the present study was to characterize the effects of insulin on the 1adrenergic receptor/effector system in vitro. Mononuclear leucocytes were employed because they are frequently used to study Padrenoceptor function in man and mirror at least changes of P2-adrenoceptors in human lung (Liggett et al., 1988) or myocardium (Michel et al. 1988).
Cells Blood was obtained from healthy young subjects and mononuclear leukocytes were isolated by density gradient centrifugation (B0yum, 1968) using Lymphoprepg (Nycomed A/S, Oslo, Norway).
Cyclic AMP Intact cells were incubated at 370 C with theophylline (4 mM) for 35 min and insulin (4 ,uiu ml-1) for 1 or 35 min before 3 min exposure to (-)-isoprenaline (lnM - lmM). The concentrations of cyclic AMP were determined by r.i.a. (Skomedal et al., 1980) with antibodies raised in our own laboratory.
Radioligand binding Intact cells were incubated with insulin (4 ,uiu
ml-1) for 1 or 35 min at 370 C before immediate cooling. The cells were then washed twice in icecold buffer and equilibrated with [3H]-CGP 12177 (0.08 - 2.85 nM) at 40 C for 18 h. Non-
Correspondence: Dr G. Sager, Department of Pharmacology, Institute of Medical Biology, University of Troms0, P.O. Box 977, N-9001 Troms0, Norway
139S
140S
G. Sager Table 1 The effect of insulin exposure (4 ,uiu ml- for 1 or 35 min) at 37 Con maximal (-)isoprenaline-induced cAMP levels (pmol/106 cell above basal level), EC50-values (nM) for (-)-isoprenaline stimulation and densities of ,B-adrenoceptors (binding sites/cell). The results are presented as mean value ± s.e. mean Insulin Exposure time Maximum response (n = 9)
EC50-value (n = 9) Receptor density (n = 7)
0 44.4 ± 4.2 260 ± 50 1420 ± 215
4,uiu ml-' 1 min
63.3 ± 4.9*** 170 ± 40** 2160 ± 240***
4,uiu ml-' 35 min
28.6 ± 4.5*** 600 ± 100*** 1480 ± 245NS
Significance vs control (paired t-test, two-tailed). NS: non-significant, **: P < 0.025, ***: P
Effects of insulin on human ,3-adrenoceptors G. SAGER Department of Pharmacology, Institute of Medical Biology, University of Troms0, P.O. Box 977, N-9001 Troms0, Norway
The effect of insulin on j-adrenoceptor stimulation and binding in mononuclear leucocytes from healthy subjects was studied. After 1 min exposure to insulin, the sensitivity and maximum response to (-)-isoprenaline and the number of ,-adrenoceptors increased. After 35 min exposure to insulin, the sensitivity and maximal response to (-)-isoprenaline stimulation were lower than the pretreatment values. The ,-adrenoceptor density was similar to the control value after 35 min exposure to insulin. The present observations may account for some of the cardiovascular effects of insulin.
Keywords insulin P-adrenoceptors cyclic AMP
Introduction
Methods
Several observations suggest that insulin plays an important regulatory role in the cardiovascular system. It is well known that the administration of insulin is associated with cardiovascular responses which include increases in heart rate (Page et al., 1976) and in peripheral blood flow (Creager et al., 1985). Insulin increases the myocardial contractility in healthy individuals and in patients with chronic congestive heart failure (Majid et al., 1972). The resultant cardiac effect of insulin is an increase of cardiac output (Arner et al., 1963). The cardiovascular effects of insulin can in part be ascribed to the elevation of plasma catecholamines in hypoglycaemia (Christensen et al., 1975), but also other adrenergic mechanisms exist (Creager et al., 1985). The aim of the present study was to characterize the effects of insulin on the 1adrenergic receptor/effector system in vitro. Mononuclear leucocytes were employed because they are frequently used to study Padrenoceptor function in man and mirror at least changes of P2-adrenoceptors in human lung (Liggett et al., 1988) or myocardium (Michel et al. 1988).
Cells Blood was obtained from healthy young subjects and mononuclear leukocytes were isolated by density gradient centrifugation (B0yum, 1968) using Lymphoprepg (Nycomed A/S, Oslo, Norway).
Cyclic AMP Intact cells were incubated at 370 C with theophylline (4 mM) for 35 min and insulin (4 ,uiu ml-1) for 1 or 35 min before 3 min exposure to (-)-isoprenaline (lnM - lmM). The concentrations of cyclic AMP were determined by r.i.a. (Skomedal et al., 1980) with antibodies raised in our own laboratory.
Radioligand binding Intact cells were incubated with insulin (4 ,uiu
ml-1) for 1 or 35 min at 370 C before immediate cooling. The cells were then washed twice in icecold buffer and equilibrated with [3H]-CGP 12177 (0.08 - 2.85 nM) at 40 C for 18 h. Non-
Correspondence: Dr G. Sager, Department of Pharmacology, Institute of Medical Biology, University of Troms0, P.O. Box 977, N-9001 Troms0, Norway
139S
140S
G. Sager Table 1 The effect of insulin exposure (4 ,uiu ml- for 1 or 35 min) at 37 Con maximal (-)isoprenaline-induced cAMP levels (pmol/106 cell above basal level), EC50-values (nM) for (-)-isoprenaline stimulation and densities of ,B-adrenoceptors (binding sites/cell). The results are presented as mean value ± s.e. mean Insulin Exposure time Maximum response (n = 9)
EC50-value (n = 9) Receptor density (n = 7)
0 44.4 ± 4.2 260 ± 50 1420 ± 215
4,uiu ml-' 1 min
63.3 ± 4.9*** 170 ± 40** 2160 ± 240***
4,uiu ml-' 35 min
28.6 ± 4.5*** 600 ± 100*** 1480 ± 245NS
Significance vs control (paired t-test, two-tailed). NS: non-significant, **: P < 0.025, ***: P
