AJH
1990;
3:22S-24S
Massimo Mannelli, Cinzia Pupilli, Rossana Lanzillotti, Lucia lanni, Francesco Alberto Ricci, and Mario Serio
Amenta,
To evaluate the presence of dopamine-2(DA2) bind ing sites in pheochromocytoma tissue, we per formed binding studies and light microscopy auto radiography on sections of 7 different tumors. 3-H-Spiroperidol, a DA2 ligand, bound specifically to tumor sections with a (mean ± SD) K value of 1.93 ± 0.62 n m o l / L . Binding site density ( B ) was 29.16 ± 2.33 fmol/mg tissue. Light microscopy auto radiography showed a nonhomogenous localization of silver grains within chromaffin cells. T h e spe cific binding was about 50% of total. To investigate whether DA2 binding sites found on pheochromocytoma cells might modulate cate cholamine (CA) release, we studied the effects of
oral bromocriptine (2.5 mg) on circulating CA of 5 patients with pheochromocytoma. In these patients bromocriptine caused a decrease in blood pressure (P < .05) but no significant change in plasma CA. Our study shows the presence of DA2 binding sites on tumoral chromaffin tissue. As bromocrip tine, a DA2 receptor agonist, was not able to modify tumor CA secretion, the functional role, if any, played by these binding sites on tumor secretion is still to be clarified. Am J Hypertens 1990;3:22S-24S
he presence of dopamine (DA) receptors on ad renal chromaffin cells has only recently been demonstrated. In 1985 Artalejo et al showed that DA and apomorphine inhibit nicotineevoked catecholamine (CA) release in perfused cat adre nal glands. They also showed that this inhibitory effect was completely blocked by haloperidol, a DA2 antago nist, and that this latter, when given alone, was able to enhance nicotine-evoked CA release. The same results were obtained in bovine adrenal medulla where DA receptors were also characterized as DA2. ' While in these animal species the presence and the modulatory role played by DA2 receptors on adrenal CA release has been clearly demonstrated, little is
T
known concerning DA receptors and the human adre nal medulla. In a recent paper we demonstrated the presence of DA modulation of human adrenal secretion showing that domperidone (DMP), a DA2 antagonist, is able to increase epinephrine (E) secretion induced by physical exercise. On the other hand, the hypertensive effect elicited by antidopaminergic agents such as metoclopramide or L-sulpiride in patients with pheochro mocytoma is well known. To elucidate the mechanism of the DA-antagonistinduced CA release from pheochromocytomas, we eval uated the presence of DA binding sites on tumor tissue as well as the effect of bromocriptine (BC), a DA2 ago nist, on plasma CA levels of patients with pheochromo cytoma.
From the Endocrinology Service, Endocrinology Unit, University o f F l o r e n c e , I t a l y ( M M , C P , R L , L I , M S ) , a n d D i p a r t i m e n t o di S c i e n z e N e u r o l o g i c h e , U n i v e r s i t a la S a p i e n z a , R o m a , I t a l y ( F A , A R ) . Address correspondence a n d reprint requests to M a s s i m o Mannelli, M D , Endocrinology Service, Endocrinology Unit, University of Flor ence, Viale M o r g a g n i 8 5 , 5 0 1 3 4 Florence, Italy.
MATERIALS AND M E T H O D S
d
m a x
1
2
© 2 9 9 0 by the American
Journal
of Hypertension,
Inc.
3
Dopamine receptors, pheochromocy toma, hypertension. KEY WORDS:
4
5
6
Clinical Studies Five patients (3 women, 2 men), all affected by a sporadic adrenal pheochromocytoma, were studied starting in the morning between 08:00 and
0895-7061/'90/'$3.00
Downloaded from https://academic.oup.com/ajh/article-abstract/3/6_Pt_2/22S/216557 by [email protected] user on 11 December 2019
Dopamine Modulation of Pathological Human Chromaffin Tissue
AJH-JUNE
1990-VOL
3, NO. 6, PART
2
D O P A M I N E IN C H R O M A F F I N
7
In Vitro Studies Pheochromocytoma specimens were obtained at surgery from seven different patients. As soon as removed, tumors were cut, frozen in liquid ni trogen and stored at — 80 °C until processed. Sections of pheochromocytoma were incubated for 60 min at room temperature in 170 nmol/L Tris-HCl buffer containing NaCl (120 nmol/L), KC1 (5 nmol/L), CaCl (2 nmol/L), MgCl (1 nmol/L) and ascorbic acid (0.001%) (final pH 7.4), in the presence of increasing concentrations (0.1 to 5 nmol/L) of 3-H-spiroperidol. The possible binding of the radioligand to 5-HT-2 or adrenergic receptors was prevented by adding to the incubation medium the 5-HT-2 receptor antagonist, ketanserin (40 nmol/L), and the adrenergic receptor antag onist, labetalol (1 /rniol/L). Nonspecific binding was calculated by adding 1 //mol/L (+)-butaclamol to the incubation medium. At the end of incubation, the slides were washed in ice cold incubation buffer ( 2 X 5 min), quickly rinsed in distilled water, wiped onto Whatman GF/B glass fiber filters and counted in a Beckman (Palo Alto, CA) liquid scintillation spectrometer. For the autoradiographic demonstration of DA-2 receptor sites, sections of pheochromocytomas were incubated for 60 min at room temperature in the abovementioned buffer containing 2.5 nmol/L 3-Hspiroperidol plus ketanserin (40 nmol/L) and labetalol (1 μηιοί/L) in the presence or in the absence of 1 //mol/L (+)-butaclamol to define nonspecific binding. At the end of incubation, sections were washed in ice-cold buffer ( 2 X 5 min), quickly rinsed in distilled water and air-dried. Sections were then processed for the autora diographic demonstration of reversible and diffusible radioligands according to the technique proposed by Young and Kuhar, by apposing Ilford L4 nuclear emul sion (diluted 1:1 in distilled water) coated coverslips to the tissue sections. After 8 to 12 weeks of exposure autoradiographs were developed with Kodak (Rochester, NY) D19, fixed with Agefix, washed in distilled water and stained with toluidine blue. The tissue and overly 2
2
8
ing silver grains in the emulsion layer were viewed and photographed with a bright- and dark-field equipped Zeiss II photomicroscope. We used the following chemicals: 3-H-spiroperidol (specific activity 75 Ci/mmol) from Amersham Radio chemical Center (England); (+)-butaclamol from Re search Biochemicals Inc. (USA); ketanserin from Janssen (Belgium); labetalol from Glaxo (UK). All data from in vitro studies were expressed as mean ± SD. The receptor concentration ( B ^ ) and the equilibrium dissociation constant (K ) values were cal culated by linear regression analysis of Scatchard plots of saturation binding data. d
RESULTS Patients with Pheochromocytoma Administration of BC caused a fall in mean arterial pressure (MAP) which was significant after 5 h (Figure 1). At that time, when compared to basal values, no consistent fall in plasma CA was observed in any of the patients. In Vitro Studies Sections bound 3-H-spiroperidol in a manner consisting with the labeling of DA2 receptor binding sites. In fact, the binding was time and concen tration dependent, reversible and, at least in the range of the concentrations we used, belonging to a single class of high affinity binding sites. Evaluation of Scatchard isotherms showed a mean K value of 1.93 ± 0.62 nmol/L and a receptor density value ( B ) of 29.16 ± 2.33 fmol/mg tissue. Light microscopy autoradiography showed a not ho mogenous localization of silver grains within chromaf fin cells as well as within pheochromocytoma areas not containing chromaffin cells. The specific binding was about 5 0 % of total. d
max
DISCUSSION The results of our in vitro studies demonstrate the pres ence of DA2 binding sites on pheochromocytoma cells. As we were not able to obtain normal human adrenal tissue for comparison, it is not possible to state whether tumor tissue possesses DA2 binding sites with charac teristics and concentrations which are different from normal chromaffin tissue. In an attempt to elucidate whether these binding sites could be involved in the hypertensive crises induced in pheochromocytoma by DA2 antagonists, we tried to activate these receptors through in vivo BC administra tion. As in normal volunteers and pheochromocy toma patients given clonidine, ' our patients showed a fall in MAP after BC. As this fall in MAP was not accom panied by a decrease in tumor CA secretion, as evi denced by unchanged plasma CA levels, we cannot draw conclusions on the role, if any, played by endoge nous DA and DA2 receptors in the determination of 5,6
9-11
12 13
Downloaded from https://academic.oup.com/ajh/article-abstract/3/6_Pt_2/22S/216557 by [email protected] user on 11 December 2019
09:00. The patients fasted and were supine with an antecubital vein cannula ted and kept patent with saline. None of the patients was receiving therapy at the time of the study. Plasma samples for CA measurement were drawn before and 1, 3, and 5 h after oral administration of 2.5 mg BC. Plasma samples (5 mL) were drawn in vacutainers containing 100 //L of a solution with EGTA (90 mg/mL) and reduced glutathione (60 mg/mL). Plasma CA was measured in duplicate by a radioenzymatic assay previously described, using reagents purchased from Amersham (Amersham, Bucks, En gland). Heart rate (HR) and blood pressure (BP) were recorded at regular intervals. Statistical analysis was performed using analysis of variance and Student's t test for paired data.
23S
24S
AJH-JUNE
MANNELLI ET AL
mmHg
1990-VOL
3, NO. 6, PART
2
tissue are necessary to understand the mechanism through which DA antagonists induce CA discharge in pheochromocytoma.
MAP
14θ1-
REFERENCES
1
3
5
0
1
3
5 HR
FIGURE 1. Plasma norepinephrine (NE) and epinephrine (E) in 5 patients with pheochromocytoma before and after oral adminis tration of 2.5 mg bromocriptine (BC). Each single patient is identi fied by a number. Jn the upper right panel mean arterial pressure (MAP) (mean ± SEM) after placebo (PL) or BC is reported. *P< .05.
tumor CA discharge in pheochromocytoma. In fact, the lack of decrease in plasma CA after BC might be due to an insufficient dose (a dose-dependent study is not ethi cally feasible) or to an abnormal intracellular transduc tion of the signal by the tumoral DA2 binding sites or to an opposite action induced on tumoral cells by in vivo BC administration as observed with normal adrenal medulla. In fact, BC, inhibiting NE release from sympa thetic nerve terminals, causes a decrease in MAP which, in vivo, induces a compensatory reflex stimulation of the adrenal gland. Such cholinergic discharge might stimulate tumor cells to release CA and, therefore, mask possible inhibitory modulation.
Artalejo AR, Garcia AG, Montiel C, et al: A dopaminer gic receptor modulates catecholamine release from the cat adrenal gland. J Physiol (Lond) 1985;362:359-368.
2.
Gonzalez MC, Artalejo AR, Montiel C, et al: Character ization of a dopaminergic receptor that modulates adrenomedullary catecholamine release. J Neurochem 1986;47:382-388.
3.
Quik M, Bergeron L, Mount H, et al: Dopamine D2 re ceptor binding in adrenal medulla: characterization using (3-H)-spiperone. Biochem Pharmacol 1987; 36:3707-3713.
4.
Mannelli M, Pupilli C, Fabbri G, et al: Endogenous dopa mine (DA) and DA2 receptors: a mechanism limiting excessive sympathetic-adrenal discharge in humans. J Clin Endocrinol Metab 1988;66:626-631.
5.
Plouin PF, Menard J, Corvol P: Hypertensive crisis in patient with pheochromocytoma given metoclopramide. Lancet 1976;2:1357-1358.
6.
Marafelia P, Perrot L, Berthezene F, et al: Valeur semeiologique du test au sulpiride et au glucagon dans le depistage du pheochromocytome. Rev Franc Endocrinol Clin 1986;27:197-200.
7.
Neri-Serneri GG, Masotti G, Gensini GF, et al: Prostacy clin and thromboxane A2 formation in response to adrenergic stimulation in humans: a mechanism for local control of vascular response to sympathetic activation? Cardiovasc Res 1981;15:287-295.
8.
Young WS, Kuhar MJ: A new method for receptor autora diography: 3H-opioid receptors in rat brain. Brain Res 1979;179:255-270.
9.
Ziegler MG, Lake CR, Williams AC, et al: Bromocriptine inhibits norepinephrine release. Clin Pharmacol Ther 1979;25:137-142.
10.
Carey RM, Van Loon GR, Baines AD, et al: Suppression of basal and stimulated noradrenergic activities by the dopamine agonist bromocriptine in man. J Clin Endo crinol Metab 1983;56:595-602.
11.
Mannelli M, Delitala G, De Feo ML, et al: Effects of different dopaminergic antagonists on bromocriptineinduced inhibition of norepinephrine release. J Clin En docrinol Metab 1984;59:74-78.
12.
Bravo EL, Tarazi RC, Fouad FM, et al: Clonidinesuppression test. A useful aid in the diagnosis of pheochromocytoma. Ν Engl J Med 1981;305:623-626.
13.
Mannelli M, DeFeo ML, Maggi M, et al: Usefulness of basal catecholamine plasma levels and clonidine sup pression test in the diagnosis of pheochromocytoma. J Endocrinol Invest 1987;10:377-382.
14.
Robertson D, Garland AJ, Robertson RM, et al: Compara tive assessment of stimuli that release neural and ad renomedullary catecholamines in man. Circulation 1979;59:637-643.
14
We believe that different in vitro studies on tumoral
Downloaded from https://academic.oup.com/ajh/article-abstract/3/6_Pt_2/22S/216557 by [email protected] user on 11 December 2019
0
1.
1990;
3:22S-24S
Massimo Mannelli, Cinzia Pupilli, Rossana Lanzillotti, Lucia lanni, Francesco Alberto Ricci, and Mario Serio
Amenta,
To evaluate the presence of dopamine-2(DA2) bind ing sites in pheochromocytoma tissue, we per formed binding studies and light microscopy auto radiography on sections of 7 different tumors. 3-H-Spiroperidol, a DA2 ligand, bound specifically to tumor sections with a (mean ± SD) K value of 1.93 ± 0.62 n m o l / L . Binding site density ( B ) was 29.16 ± 2.33 fmol/mg tissue. Light microscopy auto radiography showed a nonhomogenous localization of silver grains within chromaffin cells. T h e spe cific binding was about 50% of total. To investigate whether DA2 binding sites found on pheochromocytoma cells might modulate cate cholamine (CA) release, we studied the effects of
oral bromocriptine (2.5 mg) on circulating CA of 5 patients with pheochromocytoma. In these patients bromocriptine caused a decrease in blood pressure (P < .05) but no significant change in plasma CA. Our study shows the presence of DA2 binding sites on tumoral chromaffin tissue. As bromocrip tine, a DA2 receptor agonist, was not able to modify tumor CA secretion, the functional role, if any, played by these binding sites on tumor secretion is still to be clarified. Am J Hypertens 1990;3:22S-24S
he presence of dopamine (DA) receptors on ad renal chromaffin cells has only recently been demonstrated. In 1985 Artalejo et al showed that DA and apomorphine inhibit nicotineevoked catecholamine (CA) release in perfused cat adre nal glands. They also showed that this inhibitory effect was completely blocked by haloperidol, a DA2 antago nist, and that this latter, when given alone, was able to enhance nicotine-evoked CA release. The same results were obtained in bovine adrenal medulla where DA receptors were also characterized as DA2. ' While in these animal species the presence and the modulatory role played by DA2 receptors on adrenal CA release has been clearly demonstrated, little is
T
known concerning DA receptors and the human adre nal medulla. In a recent paper we demonstrated the presence of DA modulation of human adrenal secretion showing that domperidone (DMP), a DA2 antagonist, is able to increase epinephrine (E) secretion induced by physical exercise. On the other hand, the hypertensive effect elicited by antidopaminergic agents such as metoclopramide or L-sulpiride in patients with pheochro mocytoma is well known. To elucidate the mechanism of the DA-antagonistinduced CA release from pheochromocytomas, we eval uated the presence of DA binding sites on tumor tissue as well as the effect of bromocriptine (BC), a DA2 ago nist, on plasma CA levels of patients with pheochromo cytoma.
From the Endocrinology Service, Endocrinology Unit, University o f F l o r e n c e , I t a l y ( M M , C P , R L , L I , M S ) , a n d D i p a r t i m e n t o di S c i e n z e N e u r o l o g i c h e , U n i v e r s i t a la S a p i e n z a , R o m a , I t a l y ( F A , A R ) . Address correspondence a n d reprint requests to M a s s i m o Mannelli, M D , Endocrinology Service, Endocrinology Unit, University of Flor ence, Viale M o r g a g n i 8 5 , 5 0 1 3 4 Florence, Italy.
MATERIALS AND M E T H O D S
d
m a x
1
2
© 2 9 9 0 by the American
Journal
of Hypertension,
Inc.
3
Dopamine receptors, pheochromocy toma, hypertension. KEY WORDS:
4
5
6
Clinical Studies Five patients (3 women, 2 men), all affected by a sporadic adrenal pheochromocytoma, were studied starting in the morning between 08:00 and
0895-7061/'90/'$3.00
Downloaded from https://academic.oup.com/ajh/article-abstract/3/6_Pt_2/22S/216557 by [email protected] user on 11 December 2019
Dopamine Modulation of Pathological Human Chromaffin Tissue
AJH-JUNE
1990-VOL
3, NO. 6, PART
2
D O P A M I N E IN C H R O M A F F I N
7
In Vitro Studies Pheochromocytoma specimens were obtained at surgery from seven different patients. As soon as removed, tumors were cut, frozen in liquid ni trogen and stored at — 80 °C until processed. Sections of pheochromocytoma were incubated for 60 min at room temperature in 170 nmol/L Tris-HCl buffer containing NaCl (120 nmol/L), KC1 (5 nmol/L), CaCl (2 nmol/L), MgCl (1 nmol/L) and ascorbic acid (0.001%) (final pH 7.4), in the presence of increasing concentrations (0.1 to 5 nmol/L) of 3-H-spiroperidol. The possible binding of the radioligand to 5-HT-2 or adrenergic receptors was prevented by adding to the incubation medium the 5-HT-2 receptor antagonist, ketanserin (40 nmol/L), and the adrenergic receptor antag onist, labetalol (1 /rniol/L). Nonspecific binding was calculated by adding 1 //mol/L (+)-butaclamol to the incubation medium. At the end of incubation, the slides were washed in ice cold incubation buffer ( 2 X 5 min), quickly rinsed in distilled water, wiped onto Whatman GF/B glass fiber filters and counted in a Beckman (Palo Alto, CA) liquid scintillation spectrometer. For the autoradiographic demonstration of DA-2 receptor sites, sections of pheochromocytomas were incubated for 60 min at room temperature in the abovementioned buffer containing 2.5 nmol/L 3-Hspiroperidol plus ketanserin (40 nmol/L) and labetalol (1 μηιοί/L) in the presence or in the absence of 1 //mol/L (+)-butaclamol to define nonspecific binding. At the end of incubation, sections were washed in ice-cold buffer ( 2 X 5 min), quickly rinsed in distilled water and air-dried. Sections were then processed for the autora diographic demonstration of reversible and diffusible radioligands according to the technique proposed by Young and Kuhar, by apposing Ilford L4 nuclear emul sion (diluted 1:1 in distilled water) coated coverslips to the tissue sections. After 8 to 12 weeks of exposure autoradiographs were developed with Kodak (Rochester, NY) D19, fixed with Agefix, washed in distilled water and stained with toluidine blue. The tissue and overly 2
2
8
ing silver grains in the emulsion layer were viewed and photographed with a bright- and dark-field equipped Zeiss II photomicroscope. We used the following chemicals: 3-H-spiroperidol (specific activity 75 Ci/mmol) from Amersham Radio chemical Center (England); (+)-butaclamol from Re search Biochemicals Inc. (USA); ketanserin from Janssen (Belgium); labetalol from Glaxo (UK). All data from in vitro studies were expressed as mean ± SD. The receptor concentration ( B ^ ) and the equilibrium dissociation constant (K ) values were cal culated by linear regression analysis of Scatchard plots of saturation binding data. d
RESULTS Patients with Pheochromocytoma Administration of BC caused a fall in mean arterial pressure (MAP) which was significant after 5 h (Figure 1). At that time, when compared to basal values, no consistent fall in plasma CA was observed in any of the patients. In Vitro Studies Sections bound 3-H-spiroperidol in a manner consisting with the labeling of DA2 receptor binding sites. In fact, the binding was time and concen tration dependent, reversible and, at least in the range of the concentrations we used, belonging to a single class of high affinity binding sites. Evaluation of Scatchard isotherms showed a mean K value of 1.93 ± 0.62 nmol/L and a receptor density value ( B ) of 29.16 ± 2.33 fmol/mg tissue. Light microscopy autoradiography showed a not ho mogenous localization of silver grains within chromaf fin cells as well as within pheochromocytoma areas not containing chromaffin cells. The specific binding was about 5 0 % of total. d
max
DISCUSSION The results of our in vitro studies demonstrate the pres ence of DA2 binding sites on pheochromocytoma cells. As we were not able to obtain normal human adrenal tissue for comparison, it is not possible to state whether tumor tissue possesses DA2 binding sites with charac teristics and concentrations which are different from normal chromaffin tissue. In an attempt to elucidate whether these binding sites could be involved in the hypertensive crises induced in pheochromocytoma by DA2 antagonists, we tried to activate these receptors through in vivo BC administra tion. As in normal volunteers and pheochromocy toma patients given clonidine, ' our patients showed a fall in MAP after BC. As this fall in MAP was not accom panied by a decrease in tumor CA secretion, as evi denced by unchanged plasma CA levels, we cannot draw conclusions on the role, if any, played by endoge nous DA and DA2 receptors in the determination of 5,6
9-11
12 13
Downloaded from https://academic.oup.com/ajh/article-abstract/3/6_Pt_2/22S/216557 by [email protected] user on 11 December 2019
09:00. The patients fasted and were supine with an antecubital vein cannula ted and kept patent with saline. None of the patients was receiving therapy at the time of the study. Plasma samples for CA measurement were drawn before and 1, 3, and 5 h after oral administration of 2.5 mg BC. Plasma samples (5 mL) were drawn in vacutainers containing 100 //L of a solution with EGTA (90 mg/mL) and reduced glutathione (60 mg/mL). Plasma CA was measured in duplicate by a radioenzymatic assay previously described, using reagents purchased from Amersham (Amersham, Bucks, En gland). Heart rate (HR) and blood pressure (BP) were recorded at regular intervals. Statistical analysis was performed using analysis of variance and Student's t test for paired data.
23S
24S
AJH-JUNE
MANNELLI ET AL
mmHg
1990-VOL
3, NO. 6, PART
2
tissue are necessary to understand the mechanism through which DA antagonists induce CA discharge in pheochromocytoma.
MAP
14θ1-
REFERENCES
1
3
5
0
1
3
5 HR
FIGURE 1. Plasma norepinephrine (NE) and epinephrine (E) in 5 patients with pheochromocytoma before and after oral adminis tration of 2.5 mg bromocriptine (BC). Each single patient is identi fied by a number. Jn the upper right panel mean arterial pressure (MAP) (mean ± SEM) after placebo (PL) or BC is reported. *P< .05.
tumor CA discharge in pheochromocytoma. In fact, the lack of decrease in plasma CA after BC might be due to an insufficient dose (a dose-dependent study is not ethi cally feasible) or to an abnormal intracellular transduc tion of the signal by the tumoral DA2 binding sites or to an opposite action induced on tumoral cells by in vivo BC administration as observed with normal adrenal medulla. In fact, BC, inhibiting NE release from sympa thetic nerve terminals, causes a decrease in MAP which, in vivo, induces a compensatory reflex stimulation of the adrenal gland. Such cholinergic discharge might stimulate tumor cells to release CA and, therefore, mask possible inhibitory modulation.
Artalejo AR, Garcia AG, Montiel C, et al: A dopaminer gic receptor modulates catecholamine release from the cat adrenal gland. J Physiol (Lond) 1985;362:359-368.
2.
Gonzalez MC, Artalejo AR, Montiel C, et al: Character ization of a dopaminergic receptor that modulates adrenomedullary catecholamine release. J Neurochem 1986;47:382-388.
3.
Quik M, Bergeron L, Mount H, et al: Dopamine D2 re ceptor binding in adrenal medulla: characterization using (3-H)-spiperone. Biochem Pharmacol 1987; 36:3707-3713.
4.
Mannelli M, Pupilli C, Fabbri G, et al: Endogenous dopa mine (DA) and DA2 receptors: a mechanism limiting excessive sympathetic-adrenal discharge in humans. J Clin Endocrinol Metab 1988;66:626-631.
5.
Plouin PF, Menard J, Corvol P: Hypertensive crisis in patient with pheochromocytoma given metoclopramide. Lancet 1976;2:1357-1358.
6.
Marafelia P, Perrot L, Berthezene F, et al: Valeur semeiologique du test au sulpiride et au glucagon dans le depistage du pheochromocytome. Rev Franc Endocrinol Clin 1986;27:197-200.
7.
Neri-Serneri GG, Masotti G, Gensini GF, et al: Prostacy clin and thromboxane A2 formation in response to adrenergic stimulation in humans: a mechanism for local control of vascular response to sympathetic activation? Cardiovasc Res 1981;15:287-295.
8.
Young WS, Kuhar MJ: A new method for receptor autora diography: 3H-opioid receptors in rat brain. Brain Res 1979;179:255-270.
9.
Ziegler MG, Lake CR, Williams AC, et al: Bromocriptine inhibits norepinephrine release. Clin Pharmacol Ther 1979;25:137-142.
10.
Carey RM, Van Loon GR, Baines AD, et al: Suppression of basal and stimulated noradrenergic activities by the dopamine agonist bromocriptine in man. J Clin Endo crinol Metab 1983;56:595-602.
11.
Mannelli M, Delitala G, De Feo ML, et al: Effects of different dopaminergic antagonists on bromocriptineinduced inhibition of norepinephrine release. J Clin En docrinol Metab 1984;59:74-78.
12.
Bravo EL, Tarazi RC, Fouad FM, et al: Clonidinesuppression test. A useful aid in the diagnosis of pheochromocytoma. Ν Engl J Med 1981;305:623-626.
13.
Mannelli M, DeFeo ML, Maggi M, et al: Usefulness of basal catecholamine plasma levels and clonidine sup pression test in the diagnosis of pheochromocytoma. J Endocrinol Invest 1987;10:377-382.
14.
Robertson D, Garland AJ, Robertson RM, et al: Compara tive assessment of stimuli that release neural and ad renomedullary catecholamines in man. Circulation 1979;59:637-643.
14
We believe that different in vitro studies on tumoral
Downloaded from https://academic.oup.com/ajh/article-abstract/3/6_Pt_2/22S/216557 by [email protected] user on 11 December 2019
0
1.
