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Regulation of plasma insulin level by α2-adrenergic receptors
European Journal of Pharmacology, 65 (1980) 421--424
421
© Elsevier/North-HoUand Biomedical Press
Short communication REGULATION OF PLASMA INSULIN LEVEL BY a2-ADRENERGIC RECEPTORS TERUO NAKADATE, TOSHIO NAKAKI, TAKAMURA MURAKI and RYUICHI KATO *
Department of Pharmacology, School of Medicine, Keio University, Shinanomachi 35, Shinjuku-ku, Tokyo 160, Japan Received 10 June 1980, accepted 12 June 1980
T. NAKADATE, T. NAKAKI, T. MURAKI and R. KATO, Regulation of plasma insulin level by ~2-adrenergic receptors, European J. Pharmacol. 65 (1980) 421--424. Phentolamine, yohimbine or dihydroergotamine markedly increased plasma immunoreactive insulin (IRI)and inhibited epinephrine-induced hyperglycemia in fasted mice. On the other hand, phenoxybenzamine or prazosin only slightly increased plasma IRI and enhanced epinephrine-induced hyperglycemia. These results indicate that there is a distinct difference in the effects of ~-adrenergic blockers on the plasma IRI and glucose levels, and that ~-adrenergic receptors responsible for the plasma IRI level resemble ~2adrenergic receptors more closely. Insulin
~2-Adrenergic receptor
Plasma glucose
1. Introduction The release of immunoreactive insulin (IRI) from mammalian pancreatic islets is inhibited by ~-adrenergic stimulation and is increased by 13-adrenergic stimulation (Smith and Porte, 1976). Recently, it was suggested that a-adrenergic receptors could be subdivided into al and ~2-adrenergic receptors (Langer, 1977). The a l-adrenergic receptors are located at the postsynaptic site of nerves, While the a2adrenergic receptors are located at the presynaptic site. We showed recently that the increase in plasma glucose induced by epinephrine or phenylephrine was blocked by phentolamine and yohimbine, but not by phenoxybenzamine and prazosin in fed mice (Nakadate et al., 1980; Nakadate et al., submitted for publication). From these results we suggested the presence in mice of ~2-adrenergic receptors which regulate plasma glucose levels. Hyper-
* To whom correspondence should be addressed.
~-Adrenergic blockers
glycemia induced by adrenergic agonists involves mechanisms such as: (a) stimulation of hepatic glycogenolysis, (b) inhibition of pancreatic insulin release and (c) decrease of peripheral glucose uptake (Hornbrook, 1970). In the present study we examined whether the discrepancy in effects of ~-adrenergic blockers on plasma glucose levels may be related with the activities of the agents in regulating plasma IRI levels.
2. Materials and methods
Six-week-old male mice of ddY strain were kept in a room at constant temperature (2223°C). For mimimizing the influence of hepatic glycogenolysis on plasma glucose levels, the mice were fasted for 18 h. The mice were treated with ~-adrenergic blockers 30 min before the administration of glucose (1.0 g/kg, i.p.) and epinephrine (300/~g/kg, s.c.). In all experiments, the animals were decapitated at 12:00-14:00 and the blood collected immediately. The plasma IRI was
422
T. NAKADATE ET AL.
d e t e r m i n e d using a d o u b l e - a n t i b o d y radioi m m u n o a s s a y kit f r o m D a i n a b o t R I I n s t i t u t e Co., T o k y o , J a p a n , a n d t h e p l a s m a glucose was e s t i m a t e d b y m e a n s o f t h e glucose o x i d a s e p r o c e d u r e using a k i t f r o m B o e h r i n g e r M a n n h e i m Co., West G e r m a n y . D o s e s w e r e calcul a t e d o n t h e basis o f t h e salt o f e a c h drug. Results w e r e e v a l u a t e d b y using S t u d e n t ' s t-test. T h e f o l l o w i n g drugs w e r e used: L - e p i n e p h rine b i t a r t r a t e , p h e n o x y b e n z a m i n e h y d r o c h l o r i d e a n d y o h i m b i n e h y d r o c h l o r i d e (Nakarai C h e m i c a l s Co., K y o t o , J a p a n ) ; p h e n t o l a m i n e h y d r o c h l o r i d e ( C I B A G e i g y Co., T a k a razuka, Japan): dihydroergotamine tartrate (Sigma C h e m i c a l s Co., St. Louis, U.S.A.); p r a z o s i n h y d r o c h l o r i d e ( T a i t o Pfizer Co., Tokyo, Japan).
3. Results The time-course of the effects of phentolamine and phenoxybenzamine on the plasma I R I a n d glucose levels in fasted m i c e w e r e e x a m i n e d (fig. 1). P r e t r e a t m e n t w i t h p h e n t o l a m i n e (5 X 10 -s m o l / k g , i.p.) increased t h e plasma IRI and inhibited the hyperglycemia i n d u c e d b y a d m i n i s t r a t i o n o f glucose (1.0 g/ kg, i.p.) a n d e p i n e p h r i n e ( 3 0 0 p g / k g , s.c.) w h i c h h a d b e e n given at 0 m i n . O n t h e o t h e r hand, the pretreatment with phenoxybenzRasma glucose (rng/dl)
o Cont Glu
1oo
2~
I
GluoE • PBZ • PRA
i IIII
I III
I.X-
I
t~
*PILE
Plasma glucose 300 mg/dl)
•
YOH
200
0 L
Plasma IRI ( pU/rnl) 200 400
60O i
Cont GI.u
100
GIu • E
0
,
*PBZ *PRA • PHE *YOH *DHE
Ptasma IRI 600J ( yu~i ) 400* PHE
2000-
-30
0 15 30
60
120min
Fig. 1. Time course of the effect of phenoxybenzamine (e) or phentolamine (o) on the plasma glucose and IRI levels in 18 h fasted mice (control, glucose + epinephrine, &). Phentolamine (5 x 10 -s mol/kg = 16 mg/kg) or phenoxybenzamine (5 X 10 -s mol/kg = 17 mg/kg) was given i.p. 30 min before the administration (0 min)of glucose (1.0 g/kg, i.p.) and epinephrine (300 pg]kg, s.c.). Blood was collected 0, 15, 30, 60 and 120 min after glucose and epinephrine injections. The vertical bars represent standard errors. Each result represents an average from 5 mice. Abbreviations: PBZ, phenoxybenzamine; PHE, phentolamine.
Fig. 2. Effects of various ~-adrenergic blockers on plasma IRI and glucose levels in 18 h fasted mice. Phenoxybenzamine (5 × 10 -s mol/kg = 17 mg/kg), prazosin (5 × 10 -s mol/kg = 21 mg/kg), phentolamine (5 X 10-s mol/kg = 16 mg/kg), yohimbine (5 X 10 -s mol/kg = 19.5 mg/kg) or dihydroergotamine (5 x 10 -s mol/kg = 33 mg/kg) was given i.p. 30rain before the administration of glucose (1.0 g]kg, i.p.) and epinephrine (300pg/kg, s.c.). The determinations of plasma IRI and glucose were carried out 15 min after the administrations ofgiucose and epinephrine. Each result represents mean with standard error from 8 mice. Abbreviations: Cont, control; Glu, glucose; E, epinephrine; PBZ, phenoxybenzamine; PRA, prazosin; PHE, phentolamine; YOH, yohombine; DHE, dihydroergotamine. * P < 0.05 vs. G l u + E , * * P < 0.01 vs. Glu + E, * * * P < 0.01 vs. Glu + E, Glu + E + PBZ and Glu + E + PRA.
PLASMA INSULIN AND a2-ADRENOCEPTORS
amine (5 × 10 -s mol/kg, i.p.) caused only a slight increase in the plasma IRI and enhanced the hyperglycemia induced by glucose and epinephrine. These results indicate that phentolamine is dissimilar to phenoxybenzamine in exerting effects on the ~-adrenergic receptors regulating plasma IRI levels and on the hyperglycemia induced by glucose and epinephrine. To support this view, the effects of various a-adrenergic blockers on the plasma glucose and IRI levels were then examined (fig. 2). The pretreatment with 5 × 10 -s mol/kg of phenoxybenzamine or prazosin only slightly increased plasma IRI levels, whereas the same dose of phentolamine, yohimbine or dihydroergotamine markedly increased plasma IRI. Moreover, 10 -6 and 10 -7 mol/kg of phentolamine and yohimbine also effectively increased plasma IRI. On the other hand, phentolamine, yohimbine and dihydroergotamine clearly inhibited the hyperglycemia induced by glucose and epinephrine, but phenoxybenzamine and prazosin were ineffective.
4. Discussion ~-Adrenergic blockers, including phentolamine, yohimbine and dihydroergotamine blocked the hyperglycemia induced by glucose and epinephrine, whereas other ~-adrenergic blockers, including phenoxybenzamine and prazosin were ineffective. The former increased the plasma IRI markedly and the latter produced only slight increases in the plasma IRI. These results indicate that the effects of a-adrenergic blockers on the epinephrineinduced hyperglycemia are related with the activity which modulates plasma IRI levels and that there is a distinct difference in their ability to block the ~-adrenergic receptors responsible for plasma IRI levels. Under physiological conditions, the basal plasma IRI levels remain low in vivo irrespective of catecholamine administration, because of the ballance of ~- and ~-adrenergic stimulation induced by endogenous catecholamines
423
(Woods and Porte, 1974). In the presence of phentolamine, the plasma IRI increases markedly because the a-adrenergic receptors are blocked and the/3-adrenergic receptors are stimulated by endogenous or exogenous catecholamines (Woods and Porte, 1974; Lundquist, 1972). It was reported that prazosin and phenoxybenzamine preferentially block postsynaptic a-adrenergic receptors and that yohimbine and phentolamine are more potent in blocking presynaptic than postsynaptic ~-adrenergic receptors (Doxey et al., 1977). Although ~-adrenergic receptors were first subdivided on the basis of their location on either the pre- or the postsynaptic membrane (Langer, 1977), more recently it was reported that a2adrenergic receptors are located in extra presynaptic sites (Berthelsen and Pettinger, 1977). Our results suggest that the ~-adrenergic receptors regulating plasma IRI levels differ from classical ~-adrenergic receptors and resemble ~2-adrenergic receptors more closely. Our in vitro studies on pancreatic islets isolated from rats further support the present postulate (Nakaki et al., 1980). Recently Chung et al. (1979) reported that ~2adrenergic receptors regulated human platelet aggregation.
References Berthelsen, S. and W.A. Pettinger, 1977, A functional basis for classification of ~-adrenergic receptors, Life Sci. 21,595. Chung, Y.H., D.R. Knapp and P.V. Halushka, 1979, The effect of alpha adrenergic agents on human platelet aggregation, J. Pharmacol. Exp. Therap. 208, 366. Doxey, J.C., C.F.C. Smith and J.M. Walker, 1977, Selectivity blocking agents for pre- and postsynaptic ~-adrenoceptors, Br. J. Pharmacol. 60, 91. Hornbrook, K.R., 1970, Adrenergic receptors for metabolic responses in the liver, Federation Proc. 29, 1381. Langer, S.Z., 1977, Presynaptic receptors and their role in the regulation of transmitter release, Br. J. Pharmacol. 60,481.
424 Lundquist, I., 1972, Interaction of amines and aminergic blocking agents with blood glucose regulation. II. 0~-Adrenergic blockade, European J. Pharmacol. 18, 225. Nakadate, T., T. Muraki and R. Kato, 1980, Effects of 0~-and ~-adrenergic blockers on chlorpromazineinduced elevation of plasma glucose and cyclic AMP in fed mice, Jap. J. Pharmacol. 30, 199. Nakaki, T., T. Nakadate and R. Kato, 1980, ~2-
T. NAKADATE ET AL. Adrenoceptors modulating insulin release from pancreatic islets, Naunyn-Schmiedeb. Arch. Pharmacol. (in press). Smith, P.H. and D. Porte, Jr., 1976, Neuropharmacology of the islets, Ann. Rev. Pharmacol. Toxicol. 16, 269. Woods, S.C. and D. Porte, Jr., 1974, Neural control of the endocrine pancreas, Physiol. Rev. 54, 596.
421
© Elsevier/North-HoUand Biomedical Press
Short communication REGULATION OF PLASMA INSULIN LEVEL BY a2-ADRENERGIC RECEPTORS TERUO NAKADATE, TOSHIO NAKAKI, TAKAMURA MURAKI and RYUICHI KATO *
Department of Pharmacology, School of Medicine, Keio University, Shinanomachi 35, Shinjuku-ku, Tokyo 160, Japan Received 10 June 1980, accepted 12 June 1980
T. NAKADATE, T. NAKAKI, T. MURAKI and R. KATO, Regulation of plasma insulin level by ~2-adrenergic receptors, European J. Pharmacol. 65 (1980) 421--424. Phentolamine, yohimbine or dihydroergotamine markedly increased plasma immunoreactive insulin (IRI)and inhibited epinephrine-induced hyperglycemia in fasted mice. On the other hand, phenoxybenzamine or prazosin only slightly increased plasma IRI and enhanced epinephrine-induced hyperglycemia. These results indicate that there is a distinct difference in the effects of ~-adrenergic blockers on the plasma IRI and glucose levels, and that ~-adrenergic receptors responsible for the plasma IRI level resemble ~2adrenergic receptors more closely. Insulin
~2-Adrenergic receptor
Plasma glucose
1. Introduction The release of immunoreactive insulin (IRI) from mammalian pancreatic islets is inhibited by ~-adrenergic stimulation and is increased by 13-adrenergic stimulation (Smith and Porte, 1976). Recently, it was suggested that a-adrenergic receptors could be subdivided into al and ~2-adrenergic receptors (Langer, 1977). The a l-adrenergic receptors are located at the postsynaptic site of nerves, While the a2adrenergic receptors are located at the presynaptic site. We showed recently that the increase in plasma glucose induced by epinephrine or phenylephrine was blocked by phentolamine and yohimbine, but not by phenoxybenzamine and prazosin in fed mice (Nakadate et al., 1980; Nakadate et al., submitted for publication). From these results we suggested the presence in mice of ~2-adrenergic receptors which regulate plasma glucose levels. Hyper-
* To whom correspondence should be addressed.
~-Adrenergic blockers
glycemia induced by adrenergic agonists involves mechanisms such as: (a) stimulation of hepatic glycogenolysis, (b) inhibition of pancreatic insulin release and (c) decrease of peripheral glucose uptake (Hornbrook, 1970). In the present study we examined whether the discrepancy in effects of ~-adrenergic blockers on plasma glucose levels may be related with the activities of the agents in regulating plasma IRI levels.
2. Materials and methods
Six-week-old male mice of ddY strain were kept in a room at constant temperature (2223°C). For mimimizing the influence of hepatic glycogenolysis on plasma glucose levels, the mice were fasted for 18 h. The mice were treated with ~-adrenergic blockers 30 min before the administration of glucose (1.0 g/kg, i.p.) and epinephrine (300/~g/kg, s.c.). In all experiments, the animals were decapitated at 12:00-14:00 and the blood collected immediately. The plasma IRI was
422
T. NAKADATE ET AL.
d e t e r m i n e d using a d o u b l e - a n t i b o d y radioi m m u n o a s s a y kit f r o m D a i n a b o t R I I n s t i t u t e Co., T o k y o , J a p a n , a n d t h e p l a s m a glucose was e s t i m a t e d b y m e a n s o f t h e glucose o x i d a s e p r o c e d u r e using a k i t f r o m B o e h r i n g e r M a n n h e i m Co., West G e r m a n y . D o s e s w e r e calcul a t e d o n t h e basis o f t h e salt o f e a c h drug. Results w e r e e v a l u a t e d b y using S t u d e n t ' s t-test. T h e f o l l o w i n g drugs w e r e used: L - e p i n e p h rine b i t a r t r a t e , p h e n o x y b e n z a m i n e h y d r o c h l o r i d e a n d y o h i m b i n e h y d r o c h l o r i d e (Nakarai C h e m i c a l s Co., K y o t o , J a p a n ) ; p h e n t o l a m i n e h y d r o c h l o r i d e ( C I B A G e i g y Co., T a k a razuka, Japan): dihydroergotamine tartrate (Sigma C h e m i c a l s Co., St. Louis, U.S.A.); p r a z o s i n h y d r o c h l o r i d e ( T a i t o Pfizer Co., Tokyo, Japan).
3. Results The time-course of the effects of phentolamine and phenoxybenzamine on the plasma I R I a n d glucose levels in fasted m i c e w e r e e x a m i n e d (fig. 1). P r e t r e a t m e n t w i t h p h e n t o l a m i n e (5 X 10 -s m o l / k g , i.p.) increased t h e plasma IRI and inhibited the hyperglycemia i n d u c e d b y a d m i n i s t r a t i o n o f glucose (1.0 g/ kg, i.p.) a n d e p i n e p h r i n e ( 3 0 0 p g / k g , s.c.) w h i c h h a d b e e n given at 0 m i n . O n t h e o t h e r hand, the pretreatment with phenoxybenzRasma glucose (rng/dl)
o Cont Glu
1oo
2~
I
GluoE • PBZ • PRA
i IIII
I III
I.X-
I
t~
*PILE
Plasma glucose 300 mg/dl)
•
YOH
200
0 L
Plasma IRI ( pU/rnl) 200 400
60O i
Cont GI.u
100
GIu • E
0
,
*PBZ *PRA • PHE *YOH *DHE
Ptasma IRI 600J ( yu~i ) 400* PHE
2000-
-30
0 15 30
60
120min
Fig. 1. Time course of the effect of phenoxybenzamine (e) or phentolamine (o) on the plasma glucose and IRI levels in 18 h fasted mice (control, glucose + epinephrine, &). Phentolamine (5 x 10 -s mol/kg = 16 mg/kg) or phenoxybenzamine (5 X 10 -s mol/kg = 17 mg/kg) was given i.p. 30 min before the administration (0 min)of glucose (1.0 g/kg, i.p.) and epinephrine (300 pg]kg, s.c.). Blood was collected 0, 15, 30, 60 and 120 min after glucose and epinephrine injections. The vertical bars represent standard errors. Each result represents an average from 5 mice. Abbreviations: PBZ, phenoxybenzamine; PHE, phentolamine.
Fig. 2. Effects of various ~-adrenergic blockers on plasma IRI and glucose levels in 18 h fasted mice. Phenoxybenzamine (5 × 10 -s mol/kg = 17 mg/kg), prazosin (5 × 10 -s mol/kg = 21 mg/kg), phentolamine (5 X 10-s mol/kg = 16 mg/kg), yohimbine (5 X 10 -s mol/kg = 19.5 mg/kg) or dihydroergotamine (5 x 10 -s mol/kg = 33 mg/kg) was given i.p. 30rain before the administration of glucose (1.0 g]kg, i.p.) and epinephrine (300pg/kg, s.c.). The determinations of plasma IRI and glucose were carried out 15 min after the administrations ofgiucose and epinephrine. Each result represents mean with standard error from 8 mice. Abbreviations: Cont, control; Glu, glucose; E, epinephrine; PBZ, phenoxybenzamine; PRA, prazosin; PHE, phentolamine; YOH, yohombine; DHE, dihydroergotamine. * P < 0.05 vs. G l u + E , * * P < 0.01 vs. Glu + E, * * * P < 0.01 vs. Glu + E, Glu + E + PBZ and Glu + E + PRA.
PLASMA INSULIN AND a2-ADRENOCEPTORS
amine (5 × 10 -s mol/kg, i.p.) caused only a slight increase in the plasma IRI and enhanced the hyperglycemia induced by glucose and epinephrine. These results indicate that phentolamine is dissimilar to phenoxybenzamine in exerting effects on the ~-adrenergic receptors regulating plasma IRI levels and on the hyperglycemia induced by glucose and epinephrine. To support this view, the effects of various a-adrenergic blockers on the plasma glucose and IRI levels were then examined (fig. 2). The pretreatment with 5 × 10 -s mol/kg of phenoxybenzamine or prazosin only slightly increased plasma IRI levels, whereas the same dose of phentolamine, yohimbine or dihydroergotamine markedly increased plasma IRI. Moreover, 10 -6 and 10 -7 mol/kg of phentolamine and yohimbine also effectively increased plasma IRI. On the other hand, phentolamine, yohimbine and dihydroergotamine clearly inhibited the hyperglycemia induced by glucose and epinephrine, but phenoxybenzamine and prazosin were ineffective.
4. Discussion ~-Adrenergic blockers, including phentolamine, yohimbine and dihydroergotamine blocked the hyperglycemia induced by glucose and epinephrine, whereas other ~-adrenergic blockers, including phenoxybenzamine and prazosin were ineffective. The former increased the plasma IRI markedly and the latter produced only slight increases in the plasma IRI. These results indicate that the effects of a-adrenergic blockers on the epinephrineinduced hyperglycemia are related with the activity which modulates plasma IRI levels and that there is a distinct difference in their ability to block the ~-adrenergic receptors responsible for plasma IRI levels. Under physiological conditions, the basal plasma IRI levels remain low in vivo irrespective of catecholamine administration, because of the ballance of ~- and ~-adrenergic stimulation induced by endogenous catecholamines
423
(Woods and Porte, 1974). In the presence of phentolamine, the plasma IRI increases markedly because the a-adrenergic receptors are blocked and the/3-adrenergic receptors are stimulated by endogenous or exogenous catecholamines (Woods and Porte, 1974; Lundquist, 1972). It was reported that prazosin and phenoxybenzamine preferentially block postsynaptic a-adrenergic receptors and that yohimbine and phentolamine are more potent in blocking presynaptic than postsynaptic ~-adrenergic receptors (Doxey et al., 1977). Although ~-adrenergic receptors were first subdivided on the basis of their location on either the pre- or the postsynaptic membrane (Langer, 1977), more recently it was reported that a2adrenergic receptors are located in extra presynaptic sites (Berthelsen and Pettinger, 1977). Our results suggest that the ~-adrenergic receptors regulating plasma IRI levels differ from classical ~-adrenergic receptors and resemble ~2-adrenergic receptors more closely. Our in vitro studies on pancreatic islets isolated from rats further support the present postulate (Nakaki et al., 1980). Recently Chung et al. (1979) reported that ~2adrenergic receptors regulated human platelet aggregation.
References Berthelsen, S. and W.A. Pettinger, 1977, A functional basis for classification of ~-adrenergic receptors, Life Sci. 21,595. Chung, Y.H., D.R. Knapp and P.V. Halushka, 1979, The effect of alpha adrenergic agents on human platelet aggregation, J. Pharmacol. Exp. Therap. 208, 366. Doxey, J.C., C.F.C. Smith and J.M. Walker, 1977, Selectivity blocking agents for pre- and postsynaptic ~-adrenoceptors, Br. J. Pharmacol. 60, 91. Hornbrook, K.R., 1970, Adrenergic receptors for metabolic responses in the liver, Federation Proc. 29, 1381. Langer, S.Z., 1977, Presynaptic receptors and their role in the regulation of transmitter release, Br. J. Pharmacol. 60,481.
424 Lundquist, I., 1972, Interaction of amines and aminergic blocking agents with blood glucose regulation. II. 0~-Adrenergic blockade, European J. Pharmacol. 18, 225. Nakadate, T., T. Muraki and R. Kato, 1980, Effects of 0~-and ~-adrenergic blockers on chlorpromazineinduced elevation of plasma glucose and cyclic AMP in fed mice, Jap. J. Pharmacol. 30, 199. Nakaki, T., T. Nakadate and R. Kato, 1980, ~2-
T. NAKADATE ET AL. Adrenoceptors modulating insulin release from pancreatic islets, Naunyn-Schmiedeb. Arch. Pharmacol. (in press). Smith, P.H. and D. Porte, Jr., 1976, Neuropharmacology of the islets, Ann. Rev. Pharmacol. Toxicol. 16, 269. Woods, S.C. and D. Porte, Jr., 1974, Neural control of the endocrine pancreas, Physiol. Rev. 54, 596.