Interactions of nimodipine and cocaine on endogenous catecholamines in the squirrel monkey

The effects of nimodipine on the cocaine-induced alterations in blood pressure, heart rate, and plasma catecholamines were studied in the squirrel monkey. Cocaine in intravenously administered doses of 0.5, 1, and 2 mg/kg produced significant increases in blood pressure and significant decreases in heart rate. These cardiovascular changes were associated with transient episodes of arrhythmias and with significant increases in plasma concentrations of dopamine, epinephrine, and norepinephrine. Nimodipine, 1 micrograms/kg/min for 5 min administered intravenously 5 min after cocaine, corrects the cardiovascular and plasma catecholamine concentration changes induced by this alkaloid. The same dose of nimodipine administered 5 min before cocaine prevents elevations of blood pressure. Plasma catecholamine increments are also prevented except for the highest dose of cocaine. Cardiovascular changes induced by cocaine administration in the squirrel monkey are temporally associated with significant increments in plasma catecholamines. Administration of nimodipine prevents or minimizes these endocrine and physiologic changes.     

Changes in plasma free and sulfoconjugated catecholamines before and after acute physical exercise: experimental and clinical studies.

To elucidate whether sulfoconjugated catecholamines in plasma, especially dopamine, serve as a source of free catecholamines, we examined the change in afterload on the deconjugating activity of catecholamines in isolated Langendorff perfused rat hearts. Dopamine-sulfate was administered under ordinary or high-work-load conditions. Free dopamine in the effluent was increased by the high-work-load of the hearts, whereas conjugated dopamine showed an apparent decrease. These results indicate the possibility that deconjugation of sulfoconjugated catecholamines is accelerated by a high-work-load. To obtain further evidence in humans, we also examined the changes in the plasma levels of free and sulfoconjugated catecholamines in healthy volunteers before and after marathon running. Free dopamine increased 1.99-fold from the baseline value after exercise, whereas conjugated dopamine decreased by 12%. Similarly, the plasma levels of free noradrenaline and adrenaline increased after exercise to 2.45- and 1.51-fold their respective baseline values, while conjugated noradrenaline and adrenaline both decreased. These clinical results, as well as those of the experimental studies, suggest that the increase in plasma free catecholamines after exercise is due not only to increased release from the sympathoadrenal system but also to accelerated conversion from sulfoconjugated catecholamines in the plasma.     

Central Regulation of Adrenal Tyrosine Hydroxylase: Effect of Induction on Catecholamine Levels in the Adrenal Medulla and Plasma

The effect of induction of adrenal tyrosine hydroxylase (TH) by various centrally acting drugs on catecholamine levels in adrenal and plasma was investigated in rats. All the drugs tested, namely oxotremorine, Piribedil, B-HT 920, and HA-966, produced significant increases in adrenal dopamine content and plasma epinephrine level. Denervation of the adrenal abolished the increase in adrenal dopamine as it did the induction of tyrosine hydroxylase. The results suggest that the induced increase of adrenal TH activity, as mediated by certain drugs, results in an elevation of the plasma epinephrine level and that the adrenal dopamine content is a better indicator of the catecholamine-synthesizing capacity of the adrenal medulla than are the other catecholamines.     

A sensitive method for the determination of plasma catecholamines using liquid chromatography with electrochemical detection

Simple and sensitive methods for the determination of plasma catecholamines are of great interest since the level of catecholamines in plasma reflects the activity of the sympatho-adrenal system. In the present work a previously described procedure based on high pressure liquid chromatography with electrochemical detection has been adapted for assay of plasma catecholamines. This method permits simultaneous detection of noradrenaline, adrenaline and dopamine in concentrations down to 0.1 nmol/1 in less than one ml plasma.     

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 induced growth of gram negative bacteria.

The addition of various catecholamines to cultures of gram negative bacteria resulted in dramatic increases in growth. The ability of norepinephrine, epinephrine, dopamine and dopa to enhance the growth of Escherichia coli, Yersinia enterocolitica and Pseudomonas aeruginosa was observed to be dependent on the bacterium employed with each strain showing marked preference for one or more of the catecholamines. Catecholamine induced increases in growth were confirmed by one or more of the following methods: uptake of tritiated thymidine into newly synthesized DNA, changes in optical density or pour plate analysis. None of the catecholamine metabolites resulting from either oxidative deamination or catechol-O-methylation were able to effect similar increases in bacterial growth as compared to either norepinephrine, epinephrine or dopamine. Norepinephrine was consistently observed to effect the greatest increase in bacterial growth for all strains tested.     

Release of dopamine, noradrenaline and dopamine B-hydroxylase from mouse neuroblastoma

The murine C1300 neuroblastoma tumor was found to secrete dopamine, noradrenaline and dopamine B-hydroxylase into the circulation of tumor-bearing A/J mice. The plasma levels of dopamine, noradrenaline and dopamine B-hydroxylase increased with the size of the tumor, and the increase in noradrenaline paralleled the increase in dopamine B-hydroxylase (r = 0.86). The vesicular storage of dopamine and noradrenaline in the tumor was evidenced by a decrease of the tissue content of dopamine and noradrenaline 24 hours after the administration of reserpine (5 micrograms/g) respectively to 17.6% and 7.8% of control values. A similar observation could be made for the levels of dopamine and noradrenaline in the plasma of reserpinized C1300 mice. The total activity of dopamine B-hydroxylase in the tumor and in plasma was unaffected by the reserpine treatment. Chronic administration of 6-hydroxydopamine (100 micrograms/g for 8 days) had no effect on the tissue contents of dopamine, noradrenaline or dopamine B-hydroxylase. The release of catecholamines and dopamine B-hydroxylase from the C1300 neuroblastoma was studied in vitro on superfused tumor slices. Stimulation of these slices with 56 mM KC1 or with 5.10(-5) M tyramine failed to induce the release of endogenous dopamine, noradrenaline or dopamine B-hydroxylase above the basal outflow levels. These results are suggestive for a non-exocytotic release of catecholamines and dopamine B-hydroxylase from the neuroblastoma tumor.     

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.     

High-pressure liquid chromatographic analysis of hemolymph plasma catecholamines in immune-reactive Aedes aegypti

Tyrosine and catecholamines have been implicated as substrates for the encapsulation reactions involved in the immune response of mosquitoes to microfilariae (mff). Identification and quantitation of tyrosine and catecholamines present in Aedes aegypti hemolymph plasma were accomplished by ion-pair high-pressure liquid chromatography with electrochemical detection at either +650 or +850 mV vs Ag/AgCl. Tyrosine, dopamine, and N-beta-alanyldopamine were detected in the hemolymph plasma of naive A. aegypti. Although no differences in these compounds were observed in hemolymph plasma from A. aegypti inoculated with Dirofilaria immitis mff, the chromatogram showed a single major peak (PI) (65 microM, expressed as dopamine equivalents) that was not present in naive hemolymph plasma. Saline-inoculated controls contained only 5% of the PI in immune reactive hemolymph plasma. A high concentration of PI (127 +/- 39 microM) was also detected after treatment of hemolymph plasma with mild alkaline conditions (pH 9.0), indicating that it is normally present as an electrochemically inert form in naive mosquitoes. High concentrations of PI were also detected in the naive hemolymph plasma from three other mosquito species, but no PI was found in A. trivittatus under any conditions. PI did not cochromatograph with any of the catecholamines commonly thought to be involved in immune responses of dipterans against metazoan parasites, suggesting that it may be a unique substrate for these reactions. The biological relevance of PI was evidenced by its appearance in the hemolymph plasma of two strains of D. immitis-inoculated A. aegypti.     

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.     

Tyrosine hydroxylase activity is regulated by two distinct dopamine-binding sites

Tyrosine hydroxylase (TH), the rate-limiting enzyme in the biosynthesis of the catecholamines dopamine, noradrenaline and adrenaline, is regulated acutely by feedback inhibition by the catecholamines and relief of this inhibition by phosphorylation of serine 40 (Ser40). Phosphorylation of serine 40 abolishes the binding of dopamine to a high affinity ( K _D < 4 nM) site on TH, thereby increasing the activity of the enzyme. We have found that TH also contains a second low affinity ( K _D = 90 nM) dopamine-binding site, which is present in both the non-phosphorylated and the Ser40-phosphorylated forms of the enzyme. Binding of dopamine to the high-affinity site decreases V _max and increases the K _m for the cofactor tetrahydrobiopterin, while binding of dopamine to the low-affinity site regulates TH activity by increasing the K _m for tetrahydrobiopterin. Kinetic analysis indicates that both sites are present in each of the four human TH isoforms. Dissociation of dopamine from the low-affinity site increases TH activity 12-fold for the non-phosphorylated enzyme and 9-fold for the Ser40-phosphorylated enzyme. The low-affinity dopamine-binding site has the potential to be the primary mechanism responsible for the regulation of catecholamine synthesis under most conditions.     

Plasma free and sulfoconjugated catecholamine responses to varying exercise intensity

Plasma free catecholamines rise during exercise, but sulfoconjugated catecholamines reportedly fall. This study examined the relationship between exercise intensity and circulating levels of sulfoconjugated norepinephrine, epinephrine, and dopamine. Seven exercise-trained men biked at approximately 30, 60, and 90% of their individual maximal oxygen consumption (VO2max) for 8 min. The 90% VO2max period resulted in significantly increased plasma free norepinephrine (rest, 219 +/- 85; exercise, 2,738 +/- 1,149 pg/ml; P less than or equal to 0.01) and epinephrine (rest, 49 +/- 49; exercise, 555 +/- 516 pg/ml; P less than or equal to 0.05). These changes were accompanied by consistent increases in sulfoconjugated norepinephrine at both the 60% (rest, 852 +/- 292; exercise, 1,431 +/- 639; P less than or equal to 0.05) and 90% (rest, 859 +/- 311; exercise, 2,223 +/- 1,015; P less than or equal to 0.05) VO2max periods. Plasma sulfoconjugated epinephrine and dopamine displayed erratic changes at the three exercise intensities. These findings suggest that sulfoconjugated norepinephrine rises during high-intensity exercise.     

Effects of Handling or Immobilization on Plasma Levels of 3,4-Dihydroxyphenylalanine, Catecholamines, and Metabolites in Rats

In conscious animals, handling and immobilization increase plasma levels of the catecholamines norepinephrine (NE) and epinephrine (EPI). This study examined plasma concentrations of endogenous compounds related to catecholamine synthesis and metabolism during and after exposure to these stressors in conscious rats. Plasma levels of 3,4-dihydroxyphenylalanine (DOPA), NE, EPI, and dopamine (DA), the deaminated catechol metabolites 3,4-dihydroxyphenylglycol (DHPG), and 3,4-dihydroxyphenylacetic acid (DOPAC), and their O-methylated derivatives methoxyhydroxyphenylglycol (MHPG) and homovanillic acid (HVA) were measured using liquid chromatography with electrochemical detection at 1, 3, 5, 20, 60, and 120 min of immobilization. By 1 min of immobilization, plasma NE and EPI levels had already reached peak values, and plasma levels of DOPA, DHPG, DOPAC, and MHPG were increased significantly from baseline, whereas plasma DA and HVA levels were unchanged. During the remainder of the immobilization period, the increased levels of DOPA, NE, and EPI were maintained, whereas levels of the metabolites progressively increased. In animals immobilized briefly (5 min), elevated concentrations of the metabolites persisted after release from the restraint, whereas DOPA and catecholamine levels returned to baseline. Gentle handling for 1 min also significantly increased plasma levels of DOPA, NE, EPI, and the NE metabolites DHPG and MHPG, without increasing levels of DA or HVA. The results show that in conscious rats, immobilization or even gentle handling rapidly increases plasma levels of catecholamines, the catecholamine precursor DOPA, and metabolites of NE and DA, indicating rapid increases in the synthesis, release, reuptake, and metabolism of catecholamines.     

Effect of dietary proteins and carbohydrates on urinary and sympathoadrenal catecholamines

We previously observed that administration of tyrosine to rats or humans elevated urinary dopamine, norepinephrine and epinephrine levels. The present studies examine the effects on these urinary catecholamines of varying the ratio of protein to carbohydrate in the diets.Rats consumed diets containing 0, 18 or 40% protein (76, 58 and 36% carbohydrate respectively) for 8 days. The stress of consuming the protein-free food was associated with a 16% weight reduction, and with significantly lower serum, heart and brain tyrosine levels than those noted in rats eating the 18 or 40% protein diets. Absence of protein from the diet also decreased urinary levels of dopamine and DOPA but increased urinary norepinephrine and epinephrine, probably by increasing sympathoadrenal discharge; it also increased the excretion of DOPA in animals pretreated with carbidopa, a DOPA decarboxylase inhibitor. Carbidopa administration decreased urinary dopamine, norepinephrine and epinephrine as expected; however, among carbidopa-treated rats urinary norepinephrine and epinephrine concentrations were highest for animals consuming the protein-free diet, again suggesting enhanced release of stored catecholamines from sympathoadrenal cells. The changes in urinary catecholamines observed in animals eating the protein-free diet were similar to those seen in rats fasted for 5 days: dopamine levels fell sharply while norepinephrine and epinephrine increased.These data indicate that the effects of varying dietary protein and carbohydrate contents on dopamine secretion from peripheral structures differ from its effects on structures secreting the other two catecholamines. Protein consumption increases dopamine synthesis and release probably by making more of its precursor, tyrosine, available to peripheral dopamine-producing cells; it decreases urinary norepinephrine and epinephrine compared with that seen in protein-deprived animals, probably by diminishing the firing of sympathetic neurons and adrenal chromaffin cells.     

Plasma catecholamines in rats during rewarming from induced hypothermia

The present study sought to quantitate the levels of plasma catecholamines [norepinephrine (NE), epinephrine (E), and dopamine (DA)] during induction and rewarming from hypothermia. Male rats (317 +/- 8 g) were made hypothermic by exposure to 0.9% halothane at -10 to -15 degrees C while blood pressure (carotid artery), heart rate, and colonic temperature (Tc) were monitored. Anesthesia was discontinued when Tc reached 28 degrees C. Tc continued to fall but was held at 20-20.5 degrees C for 30 min. Rewarming was then initiated by raising ambient temperature to 22 degrees C. Arterial blood samples were taken 1) before cooling, 2) just before rewarming, 3) when Tc reached 22 degrees C during rewarming, and 4) when Tc reached 27 degrees C during rewarming. Plasma was assayed radioenzymatically for catecholamines using both phenylethanolamine-N-methyltransferase and catechol-O-methyltransferase procedures, and hypothermic induction resulted in significant increases in NE, E, and DA above control levels (P less than 0.01). With rewarming to Tc = 22 degrees C, all catecholamines increased above the level observed during hypothermia (P less than 0.01), and NE and DA increased still further (P less than 0.01) when Tc reached 27 degrees C. The levels of plasma catecholamines observed during hypothermia and during the rewarming phase indicate a role of the sympathoadrenal medullary system in the metabolic adjustments associated with hypothermia and recovery. During rewarming, the levels of E and NE attained exceed those at which both substances may be expected to act as circulating hormones.     

Activation of angiotensin II receptors in brain potentiates the stimulating effect of endogenous opioid neurons on central sympathetic outflow

Endogenous opioid peptides appear to have neurotransmitter or neuromodulator functions in brain mediating a wide variety of effects. We have reported that intracisternal administration of synthetic human beta-endorphin increases plasma concentration of catecholamines, apparently by acting at unknown brain sites to increase sympathetic outflow to the adrenal medulla and sympathetic nerves. In the present study we examined the possibility that angiotensin II, acting in brain, modulates endorphin-induced catecholamine secretion. Simultaneous intracisternal administration of angiotensin II 1.0 nmol together with synthetic human beta-endorphin 1.45 nmol potentiated the plasma epinephrine, norepinephrine and dopamine responses to intracisternal beta-endorphin. In contrast, simultaneous intracisternal administration of the angiotensin II antagonist, [Sar1, Val5, Ala8]-angiotensin II (saralasin), 1.1 nmol together with beta-endorphin, blunted the plasma epinephrine, norepinephrine and dopamine responses to beta-endorphin. These data are consistent with the hypothesis that activation of angiotensin II receptors in brain potentiates the endorphin-induced stimulation of central sympathetic outflow. It remains to be demonstrated whether angiotensin II acting in brain to modulate activity of opioid neurons is synthesized in brain or is derived peripherally.     

Calcium dependent regulation of catecholamine and serotonin metabolism in human neuroblastoma cells

Three human neuroblastoma cell lines were shown to have markedly different contents of catecholamines and serotonin. Two of the cell lines (CHP-134 and IMR-5) have higher levels of dopamine and its metabolites, while CHP-404 cells have higher levels of serotonin and its metabolites. Each cell line responded to the addition of D,L-2-amino-5-phosphonovalerate, an agent which increases plasma membrane permeability to Ca2+ (Pastuszko and Wilson, 1988; with striking changes in the metabolism of the neurotransmitters. These changes were dependent on the extracellular calcium concentration and include activation of dopamine synthesis (tyrosine hydroxylase), increased levels of dihydroxyphenylacetic acid and increased formation of N-methylated dopamine derivatives. Catabolism of serotonin to 5-hydroxyindole acetic acid was inhibited while that to 5-hydroxytryptophol was stimulated. These data clearly identify several important sites for regulation of neurotransmitter metabolism by calcium. The mechanisms, direct or indirect, by which the enzyme activities are modulated by calcium remain to be established.     

Catecholamines in the sub-esophageal ganglia and the hemolymph of the giant African snail (Achatina fulica Férussac) determined by gas chromatography]

The content of three catecholamines (dopamine, L-norepinephrine and L-epinephrine) in the subesophageal ganglia and the hemolymph of Achatina fulica Férussac was measured by a gas chromatograph with electron capture detector. The dopamine content of the ganglia was 4.3 +/- 0.9 mug/g. L-Norepinephrine and L-epinephrine in the ganglia were not detected. The three catecholamines in the hemolymph were also not detected.     

Simultaneous single isotope radioenzymatic assay of plasma norepinephrine,epinephrine and dopamine

Modification of the original single isotope radioenzymatic assay of Passon and Peuler (1) permits the direct and simultaneous analysis of norepinephrine, epinephrine and dopamine in plasma samples of 50 μl or less. Plasma or cerebrospinal fluid without prior extraction of catecholamines or deproteinization is added directly into a mixture of 100 μl. This catechol-O-methyl-transferase-catalyzed assay is sensitive to 1 pg (20 pg/ml of plasma) for norepinephrine and epinephrine and 6 pg (120 pg/ml) for dopamine. A rapid thin layer chromatographic separation of the three ³H-methylcatecholamines contributes to the excellent specificity of the differential assay of the three catecholamines. The differential analysis of 15–20 plasma samples can be completed easily within one day. A total assay which omits the chromatographic step and, thus, measures norepinephrine plus epinephrine at the same sensitivity can be completed in 20 samples in one-half a working day.     

Footshock-induced changes in brain catecholamines and indoleamines are not mediated by CRF or ACTH

Stressful treatments have long been associated with increased activity of brain catecholaminergic and serotonergic neurons. An intracerebroventricular (icv) injection of the corticotropin-releasing factor (CRF) also activates brain catecholaminergic neurons. Because brain CRF-containing neurons appear to be activated during stress, it is possible that CRF mediates the catecholaminergic activation. This hypothesis has been tested by assessing the responses in brain catecholamines and indoleamines to footshock in mice pretreated icv with a CRF receptor antagonist, and in mice lacking the gene for CRF (CRFko mice). Consistent with earlier results, icv administration of CRF increased catabolites of dopamine and norepinephrine, but failed to alter tryptophan concentrations or serotonin catabolism. A brief period of footshock increased plasma corticosterone and the concentrations of tryptophan and the catabolites of dopamine, norepinephrine and serotonin in several brain regions. Mice injected icv with 25 microg alpha-helical CRF(9-41) prior to footshock had neurochemical responses that were indistinguishable from controls injected with vehicle, while the increase in plasma corticosterone was slightly attenuated in some experiments. CRFko mice exhibited neurochemical responses to footshock that were indistinguishable from wild-type mice. However, whereas wild-type mice showed the expected increase in plasma corticosterone, there was no such increase in CRFko mice. Similarly, hypophysectomized mice also showed normal neurochemical responses to footshock, but no increase in plasma corticosterone. Hypophysectomy itself elevated brain tryptophan and catecholamine and serotonin metabolism. Treatment with ACTH icv or peripherally failed to induce any changes in cerebral catecholamines and indoleamines. These results suggest that CRF and its receptors, and ACTH and other pituitary hormones, are not involved in the catecholamine and serotonin responses to a brief period of footshock.