Use of plasma catecholamines in diagnosing pheochromocytoma and neuroblastoma

The diagnosis in two cases of neural crest tumours was confirmed by the measurement of concentration of plasma catecholamines. A histamine provocative test monitored by values of plasma catecholamines provided support for a diagnosis of pheochromocytoma in a patient with normal blood pressure and elevated values of urinary amines. In the second case the presence of a ganglioneuroblastoma secreting abnormal amounts of catecholamines was detected by plasma epinephrine and norepinephrine measurements when values of urinary free catecholamines were normal.     

Simultaneous elevation of plasma insulin and catecholamines during endotoxicosis in the conscious and anesthetized rat

Plasma levels of glucose, insulin and catecholamines were assessed during the early phase of sub-lethal endotoxicosis in fasted male rats which were either conscious or continuously anesthetized with sodium pentobarbital. Exogenous glucose challenge was administered during endotoxicosis to probe insulin release at a time when plasma catecholamines were elevated. An endogenous hyperglycemia occurred following endotoxin but was moderated by continuous pentobarbital anesthesia. Plasma insulin was elevated in the conscious but not anesthetized rats during endogenous hyperglycemia following endotoxin. Hyperglycemia with exogenous glucose elevated plasma insulin levels in both conscious and anesthetized groups and occurred in the presence of elevated levels of norepinephrine, epinephrine and dopamine. Simultaneous elevation of plasma catecholamine and insulin levels during endotoxicosis suggests that glucose utilization may be promoted at the same time that glucose is mobilized through adrenergic mechanisms. These events may contribute to the rapid depletion of carbohydrate stores leading to the hypoglycemia of the agonal stage of endotoxic shock.     

Enzymatic deconjugation of catecholamines in human and rat plasma and red blood cell lysate

We have developed a method for enzymatic hydrolysis of both sulfated and glucuronidated catecholamines in plasma and red blood cell lysate. Hydrolysis occurs in the course of the radioenzymatic assay for catecholamines. In human plasma, catecholamines are conjugated almost entirely with sulfate while, in rat plasma, glucuronides are the main conjugates of epinephrine and dopamine but not norepinephrine. Rat plasma contains less percent conjugated catecholamine than human plasma. Human red blood cell lysate contains less conjugated catecholamine than plasma, whereas free E in lysate exceeds that of plasma and free NE has same level both in plasma and lysate. This method is useful in detecting total (free + sulfated + glucuronidated) catecholamines and the nature of conjugated catecholamines.     

Catecholamine concentrations in plasma and organs of the fetal guinea pig during normoxemia, hypoxemia, and asphyxia

To examine the responses of the sympatho-adrenal system to reduced oxygen supply we studied plasma and tissue concentrations of catecholamines during normoxemia, hypoxemia, and asphyxia in 22 fetal guinea pigs near term. Fetal blood was obtained by cardiopuncture in utero under ketamine/xylazine-anesthesia. Catecholamines were determined in plasma and tissue of 15 organs and 14 brain parts by HPLC-ECD. During normoxemia (SO2 54 +/- 4 (SE) %, pH 7.36 +/- 0.02, n = 5) plasma catecholamine levels were low (norepinephrine 447 +/- 53, epinephrine 42 +/- 12, dopamine 44 +/- 6 pg/ml). During hypoxemia (SO2 27 +/- 3%, pH 7.32 +/- 0.01, n = 6) and asphyxia (SO2 24 +/- 2%, pH 7.23 +/- 0.02, n = 11) tissue catecholamine concentrations changed with changing blood gases and with increasing plasma catecholamines. Norepinephrine concentrations increased in both skin and lung and decreased in liver, pancreas, and scalp; those of epinephrine increased in the heart, lung liver, and scalp and decreased in the adrenal. There were only minor changes in brain catecholamine concentrations except for a 50% reduction in dopamine in the caudate nucleus. Concentrations of dopamine catabolite 3,4-dihydroxyphenylacetic acid decreased in many brain parts, suggesting that cerebral catecholamine metabolism was affected by hypoxemia and asphyxia. We conclude that the sympatho-adrenal system of fetal guinea pigs near term is mature and that its stimulation by reduced fetal oxygen supply leads to changes in both plasma and tissue catecholamine concentrations.     

Catecholamines in head and body blood of eels and rats

1. When compared with other vertebrates, the circulating titers of norepinephrine and epinephrine of the yellow eel are very low. 2. The ratio of the catecholamine titers in the eel differs from that reported for other vertebrates. 3. Following decapitation, the titers of the catecholamines are higher in head blood than in body blood of both unanesthetized and anesthetized eels. In decapitated rats, only the dopamine titer is higher in head blood. 4. As in the lamprey, agitation stress causes a drop of circulating catecholamines. However, other forms of stress cause the expected increase. 5. It appears that many data on catecholamines in both brain and circulation of vertebrates in general have been influenced by stress effects.     

Free and conjugated plasma catecholamines, DOPA and 3-O-methyldopa in humans and in various animal species

The aim of the present study was to determine the extent to which plasma catecholamines are conjugated in different animals compared to man and how widespread is the presence of dihydroxyphenylalanine (DOPA) and 3-methoxy-4-hydroxyphenylalanine (3-OMD) in plasma among the different animal species. Free and conjugated norepinephrine, epinephrine, and dopamine were measured in plasma in humans and in several animal species (dog, rat, Gunn rat, cat, rabbit, guinea pig, African green monkey, young pig, calf, and one American black bear) using HPLC with electrochemical detection. The same technique was used to measure free and conjugated DOPA and 3-OMD in plasma of man, dog, rat, Gunn rat, calf, and American black bear. Human plasma contains the highest concentration of total (free and conjugated) catecholamines (46.1 pmole/ml), while low concentrations (below 15 pmole/ml) were observed in unstressed rats, calves, cats, and young pigs. In man, 95.3% of total plasma catecholamines were conjugated. The extent to which plasma catecholamines were conjugated varied greatly between animal species. The conjugated fraction expressed as percentages of the total catecholamines is lowest in the young pig (4.7%) and highest in the bear (100%). Conjugated dopamine was present in the plasma of all species, varying between 3% of the total catecholamine pool in young pig to 90% in dog. Conjugated norepinephrine was also present in plasma of all species except in unstressed rats with access to food. Conjugated epinephrine was detected only in cat and rat. Free DOPA and 3-OMD were present in plasma of all tested species with especially high levels of 3-OMD being present in dog. Conjugated DOPA and 3-OMD were not consistently found in any species. Our results indicate that man, dog, bear, and African green monkey are particularly good catecholamine conjugators and that young pig, guinea pig, rabbit, and calf are poor conjugators.     

Effects of asphyxia on arterial blood pressure, formation of nitric oxide in medulla and blood parameters in the cat

The rostral ventrolateral medulla (RVLM) plays an important role in the integration of cardiovascular functions. We examined the effect of asphyxia on cardiovascular responses, on sympathetic vertebral nerve activity (VNA) and nitric oxide (NO) formation in the RVLM, on hemodynamics, and on plasma concentrations of catecholamines, blood gas partial pressures and carbohydrate metabolites. Using 16 anesthetized cats we found that the systemic arterial pressure (SAP), VNA, NO formation and the release of plasma catecholamine components of norepinephrine and epinephrine were increased during asphyxia. The onset of NO production was significantly earlier than that of SAP and VNA. The venous partial pressure of O2 decreased, while the partial pressure of CO2 increased. Furthermore, metabolism of glucose and lactate increased, as did the blood concentrations of white and red blood cells, hemoglobin and platelets. Thus, asphyxia increased SAP, VNA and NO formation. It increased the plasma catecholamines, blood gases, carbohydrate metabolites and blood cells.     

III. Determination of plasma catecholamines and free 3, 4-dihydroxyphenylacetic acid in continuously collected human plasma by high performance liquid chromatography with electrochemical detection

We have presented a sensitive and relatively simple and inexpensive method for continuous sampling and determination of plasma catecholamines and a major dopamine metabolite, DOPAC. This method provides the basis for determination of the short-term magnitude of catecholamine response as well as the time course of such a response following several physical or psychological interventions. Resting levels of plasma catecholamines--norepinephrine 292 pg/ml, epinephrine 81 pg/ml and dopamine 29 pg/ml--are comparable to those obtained by other methods. Dopamine and free DOPAC were unaffected by physical or psychological interventions while norepinephrine was considerably increased by isometric handgrip, knee bends, and cold pressor and epinephrine increased during knee bends, mental arithmetic, cold pressor, and blood pressure measurement.     

Cardiovascular regulation in TGR(mREN2)27 rats: 24h variation in plasma catecholamines, angiotensin peptides, and telemetric heart rate variability

Dysfunction of the sympathetic nervous system might play an important role in disturbed 24h blood pressure regulation in transgenic hypertensive TGR (mREN2)27 (TGR) rats. Our study was performed to determine possible differences in activity of the sympathetic nervous system in TGR rats in comparison to their normotensive Sprague-Dawley (SPRD) controls; we measured plasma catecholamine and angiotensin concentrations throughout 24h under synchronized light-dark 12h:12H (LD 12:12) conditions. In the TGR rat strain, rhythms of plasma catecholamines were blunted, and the concentrations were significantly decreased. In addition, TGR rats showed increased plasma angiotensin I and II concentrations without any significant rhythm. An impaired autonomic regulation was confirmed by monitoring heart rate variability in TGR rats. Data showed that the TGR rat strain is characterized by a reduction in plasma catecholamines and an increase in angiotensin peptides. At present, it is not clear whether the reduction in catecholamines represents a decrease in sympathetic tone mediated by baroreflex activation or an increased catecholamine turnover induced by elevated angio-tensin II. However, the blunted, but normally phased, rhythms in plasma catecholamines in TGR rats make it unlikely that the sympathetic nervous system is mainly responsible for the inverse circadian blood pressure rhythm in the transgenic strain. (Chronobiology International, 18(3), 461-474, 2001)     

Plasma catecholamines: Arterial-venous differences and the influence of body temperature

Using sensitive radio-enzymatic assays, levels of plasma total catecholamines and norepinephrine in rats change dramatically with changes in body temperature. The decrease in plasma catecholamines induced by warming the animal is reflected in an apparent arterio-venous difference when arterial blood is obtained at room temperature and tail sampling is aided by heat induced vasodilation. Combined blockade of extraneuronal and neuronal uptake reduces this arterio-venous difference. Blood samples obtained from the decapitated trunk of the rat contain similar levels of plasma catecholamines as those obtained from indwelling carotid catheters. Blood levels of dopamine-betahydroxylase were similar whether obtained by venous sampling during heat-induced vasodilation, decapitation or indwelling arterial cannula.     

CARDIOVASCULAR REGULATION IN TGR(mREN2)27 RATS: 24h VARIATION IN PLASMA CATECHOLAMINES,ANGIOTENSIN PEPTIDES,AND TELEMETRIC HEART RATE VARIABILITY

Dysfunction of the sympathetic nervous system might play an important role in disturbed 24h blood pressure regulation in transgenic hypertensive TGR (mREN2)27 (TGR) rats. Our study was performed to determine possible differences in activity of the sympathetic nervous system in TGR rats in comparison to their normotensive Sprague-Dawley (SPRD) controls; we measured plasma catecholamine and angiotensin concentrations throughout 24h under synchronized light-dark 12h:12H (LD 12:12) conditions. In the TGR rat strain, rhythms of plasma catecholamines were blunted, and the concentrations were significantly decreased. In addition, TGR rats showed increased plasma angiotensin I and II concentrations without any significant rhythm. An impaired autonomic regulation was confirmed by monitoring heart rate variability in TGR rats. Data showed that the TGR rat strain is characterized by a reduction in plasma catecholamines and an increase in angiotensin peptides. At present, it is not clear whether the reduction in catecholamines represents a decrease in sympathetic tone mediated by baroreflex activation or an increased catecholamine turnover induced by elevated angio-tensin II. However, the blunted, but normally phased, rhythms in plasma catecholamines in TGR rats make it unlikely that the sympathetic nervous system is mainly responsible for the inverse circadian blood pressure rhythm in the transgenic strain. (Chronobiology International, 18(3), 461–474, 2001)     

Modulation of catecholamine release by endogenous adenosine in the rat adrenal medulla

Adenosine was shown to inhibit norepinephrine (NE) release from sympathetic nerve endings. The purpose of this study was to examine whether endogenous adenosine restrains NE and epinephrine release from the adrenal medulla. The effects of an adenosine receptor antagonist, 1,3-dipropyl-8-(p-sulfophenyl) xanthine (DPSPX), on epinephrine and NE release induced by intravenous administration of insulin in conscious rats were examined. Plasma catecholamines were measured by HPLC with an electrochemical detector. DPSPX significantly increased plasma catecholamine in both control rats and rats treated with insulin. The effect of DPSPX on plasma catecholamine was significantly greater in rats treated with insulin. Additional experiments were performed in adrenalectomized rats to investigate the contribution of the adrenal medulla to the effect of DPSPX on plasma catecholamine. The effect of DPSPX and insulin on epinephrine in adrenalectomized rats was significantly reduced compared with that of the controls. Finally, we tested whether endogenous adenosine restrains catecholamine secretion partially through inhibiting the renin-angiotensin system. The effect of DPSPX on plasma catecholamine in rats pretreated with captopril (an angiotensin-converting enzyme inhibitor) was reduced. These results demonstrate that under basal physiological conditions, endogenous adenosine tonically inhibits catecholamine secretion from the adrenal medulla, and this effect is augmented when the sympathetic system is stimulated. The effect of endogenous adenosine on catecholamine secretion from the adrenal medulla is achieved partially through the inhibitory effect of adenosine on the renin-angiotensin system.     

Somatostatin mediates the effect of growth hormone surges on splanchnic biogenic amines

Experiments were conducted in trained, conscious dogs fitted with an indwelling portal catheter. Radioenzymatic methods were employed for the quantitative measurement of plasma-free serotonin and catecholamines. An injection of ovine growth hormone (GH, 100 micrograms/kg) or an equimolar amount of somatostatin (somatotropin release inhibitory factor, SRIF, 7.5 micrograms/kg) into a saphenous vein led, within the first 15 min, to a transient but significant increase in plasma serotonin and a decrease in the concentrations of dopamine, norepinephrine, and epinephrine. The changes were frequently in excess of 40% of baseline values, and were found only in the portal and not in the peripheral circulation. When the animals were pretreated with an antiserum specifically directed against SRIF, GH surges no longer caused alterations in the portal levels of biogenic amines. Thus, the effects of spike concentrations of GH on plasma serotonin and catecholamines are apparently mediated by SRIF, a novel and unexpected function for a hormone that is known as an inhibitor of GH secretion.     

Effect of pertussis toxin on the adrenergic regulation of plasma renin activity

Basal plasma renin activity (PRA) was not modified by pertussis toxin administration. On the contrary, the modulation of PRA by adrenergic amines was markedly affected by the toxin. Administration of epinephrine did not modified PRA in the controls but markedly increased it in toxin-treated rats. This effect of epinephrine was reproduced in control rats when yohimbine was given before the catecholamine. Clonidine decreased PRA to a much more significant extent in control rats than in animals treated with the toxin. Isoproterenol stimulated PRA to a greater level in toxin-treated rats. Our data indicates that pertussis toxin blocks the alpha 2-adrenergic modulation of renin release and magnifies the ability of beta adrenergic activation to stimulate PRA.     

Ameliorating effects of amlodipine on plasma and myocardial catecholamines in BIO 53.58 Syrian hamsters, a model of dilated cardiomyopathy

Our previous study has shown that the concentrations of norepinephrine, epinephrine and dopamine in the plasma of BIO 53.58 hamsters (a model of dilated cardiomyopathy: DCM) at 18 weeks of age (severe cardiomyopathic stage) were twice those of age-matched F1B control and conversely the myocardial norepinephrine level was decreased. The present study was undertaken to examine the effect of amlodipine on catecholamine concentration, myocardial receptors and histopathological changes in BIO 53.58 hamsters. Oral administration of amlodipine (10 mg/kg/day) for 7 weeks in 11 week-old-BIO 53.58 hamsters brought about marked decreases in the concentrations of norepinephrine, epinephrine and dopamine in the plasma, compared with those in vehicle-treated BIO 53.58 hamsters. This was accompanied by a concomitant increase in the concentration of myocardial catecholamine concentration. In other words, the concentrations of catecholamines in plasma and myocardium of amlodipine administered BIO 53.58 hamsters approximated to the control level in age-matched F1B. In addition, amlodipine administration caused a significant reduction of calcium deposition with a tendency toward a decrease in the myocardial necrosis, and it had little effect on the affinity and number of specific binding for (+)-[3H]PN 200-110, (-)-[125I]iodocyanopindolol (CYP) and [3H]prazosin in the myocardium. In conclusion, the present study shows that administration of amlodipine in BIO 53.58 hamsters may exhibit ameliorating effect on plasma and myocardial catecholamines with a significant reduction of calcium deposition. These data may offer further support for the use of amlodipine in patients with DCM.     

Effects of exercise on plasma catecholamine levels in the toad, Bufo paracnemis: role of the adrenals and neural control

Resting plasma epinephrine (E) and norepinephrine (N) concentrations for intact toads (Bufo paracnemis) were 5.57+/-1.0 and 0.88+/-0.38 ng/ml, respectively. Exercise induced a significant increase in heart rate, blood pressure and plasma epinephrine (about 4.3 times), whereas norepinephrine remained unchanged. The resting [E]/[N] ratio was 6.3 and increased to 32.9 during exercise. Adrenal denervation did not alter the basal plasma catecholamine or norepinephrine levels after exercise, but prevented the increase in epinephrine during exercise, suggesting that in the intact toad this increase is due to adrenal secretion whereas resting norepinephrine may be liberated by extra-adrenal chromaffin tissues. This also suggests that the adrenal glands can release selectively the two catecholamines. The increases in heart rate and blood pressure in denervated toads were not significantly different from those of intact animals, suggesting that during exercise the sympathetic nerves play the main role in inducing cardiovascular responses. Spinal transection induced a significant increase in basal norepinephrine levels, which remained elevated after exercise. Since spinal toads are unable to perform spontaneous movements it is possible that this increase may be caused by this stressful condition. The increases in heart rate and blood pressure observed in spinal toads during exercise may be due to direct mechanical effects of venous return on the heart.     

Plasma catecholamines: effects of footshock level and hormonal modulators of memory storage

Plasma levels of norepinephrine (NE) and epinephrine (EPI) were measured in male Sprague-Dawley rats before and at several times after training injections of agents known to enhance or to impair later retention performance for a one-trial inhibitory (passive) avoidance task. Two days before testing, each animal was surgically prepared with a chronic tail artery catheter that allows for repeated blood sampling in unhandled rats. Exposure to a single, intense training footshock (3.0 mA, 2.0 sec duration) resulted in an immediate but transient increase in plasma levels of EPI and to a lesser extent NE. Plasma levels of both catecholamines did not differ between unshocked controls and animals that received a weak training footshock (0.6 mA, 0.5 sec duration). An injection of EPI at a dose that enhances retention performance (0.1 mg/kg, sc) resulted in increments in plasma EPI levels of 0.8-1.9 ng/ml from 5 to 40 min after injection. An injection of EPI (0.5 mg/kg, sc) at a dose that produces retrograde amnesia resulted in increments in plasma EPI ranging from 3.7 to 4.5 ng/ml during the 40 min after injection. Plasma NE levels were not significantly altered following an EPI injection. A single injection of adrenocorticotropin (ACTH, 0.3 or 3.0 IU per rat) did not alter the plasma catecholamine responses to training with a weak footshock. Similarly, the synthetic ACTH analog, Organon 2766 (125 or 250 mg/Kg) did not affect plasma catecholamines in untrained (unshocked) rats.These results demonstrate that significant increments in plasma levels of NE and EPI occur shortly after inhibitory avoidance training. Furthermore, an injection of EPI that enhances retention of an inhibitory avoidance task mimics the magnitude, though not the temporal characteristics, of the endogenous adrenal medullary response to a training footshock. Other hormonal treatments (ACTH and Organon 2766) which enhance memory storage do not affect plasma levels of NE and EPI.     

Maternal plasma catecholamines in the rhesus macaque during late gestation: effect of photoperiod and timed melatonin infusion.

This study tested the hypothesis that changes in photoperiod alter plasma catecholamine concentrations in the rhesus monkey during late gestation. Twelve chronically catheterized pregnant rhesus macaques were acclimated to a 12-h photoperiod (lights-on, 0700-1900 h). Under the control L:D cycle, blood samples were collected at 3-h intervals over 24 h for catecholamine analysis. Plasma concentrations (mean +/- SEM, pg/ml) ranged from 678 +/- 90 to 928 +/- 142 for norepinephrine; 230 +/- 22 to 631 +/- 141 for epinephrine; and 282 +/- 70 to 1090 +/- 362 for dopamine. A diurnal rhythm was observed in epinephrine with peak concentrations during lights-on (0900-1800 h; p less than 0.05, compared to lights-off). After the first sampling protocol, the animals were divided equally between two groups: phase shift, in which lights-on was shifted 11 h (2000-0800 h) and constant light, with lights on continuously. After the phase shift, a parallel shift in the plasma epinephrine rhythm was noted, with peak levels observed between 2200 and 0700 h (p less than 0.05). Constant light abolished the rhythm in epinephrine, with an overall reduction in mean basal levels of all three catecholamines. Daily melatonin infusions (0.2 micrograms/kg/h, 1900-0630 h) under constant light failed to restore the epinephrine rhythm or to return basal catecholamine concentrations to control photoperiod levels. These data suggest that photoperiod entrains the rhythm in epinephrine secretion, but the rhythm is ablated under constant conditions. Further, melatonin does not appear to play a role in the regulation of catecholamine secretion in the pregnant rhesus macaque.     

Estimation of catecholamines in human plasma by ion-exchange chromatography coupled with fluorimetry

Estimation of catecholamines in human plasma was made by ion-exchange chromatography coupled with fluorimetry.Catecholamines in deproteinized plasma were adsorbed onto Amberlite CG-50 (pH 6.5, buffered with 0.4 M phosphate buffer) and selectively eluted by 0.66 M boric acid. The catecholamine fraction was separated further on a column of Amberlite IRC-50 which was coupled with a device for the automated performance of the trihydroxyindole method (epinephrine and norepinephrine) or the 4-aminobenzoic acid—oxidation method (dopamine). One sample could be analysed within 25 min with either method. The lower detection limits were 0.02 ng for epinephrine and dopamine, and 0.04 ng for norepinephrine.Plasma catecholamine contents of healthy adults at rest were epinephrine 0.07 ± 0.01 ng/ml (n = 19), norepinephrine 0.27 ± 0.03 ng/ml (n = 19) and dopamine 0.22 ± 0.03 ng/ml (n = 26).The procedure of adsorption and elution of the plasma catecholamines by ion-exchange resin was simple, the simplicity contributing to constant recovery. The catecholamine fraction could be analysed without evaporation of the eluate. The analytical column could be used for the analysis of more than 1000 samples before excessive back-pressure developed. Our method of continuous measurement of plasma catecholamine fulfils clinical requirements.     

Analysis of plasma catecholamines by high-performance liquid chromatography with fluorescence detection: simple sample preparation for pre-column fluorescence derivatization

Analysis of plasma catecholamines (norepinephrine, epinephrine and dopamine) by high-performance liquid chromatography using 1,2-diphenylethylenediamine as a fluorescent reagent is described. We have developed an automatic catecholamine analyser, based on pre-column fluorescence derivatization and column switching. The analysis time for one assay was 15 min. The correlation coefficients of the linear regression equations were greater than 0.9996 in the range 10–10 000 pg/ml. The detection limit, at a signal-to-noise ratio of 3, was 2 pg/ml for dopamine. A new method of sample preparation for the pre-column fluorescence derivatization of plasma catecholamines was used. In order to protect the catecholamines from decomposition, an ion-pair complex between boric acid and the diol group in the catecholamine was formed at a weakly alkaline pH. The stabilities of plasma catecholamines were evaluated at several temperatures. After complex formation, the catecholamines were very stable at 17°C for 8 h, and the coefficients of variation for norepinephrine, epinephrine and dopamine were 1.2, 4.2 and 9.3%, respectively.