Levodopa, placebo and rotigotine change biomarker levels for oxidative stress.

INTRODUCTION: Homocysteine increase and glutathione derivative cysteinyl-glycine fall are indirect biomarkers for oxidative stress, for instance due to dopamine D1 receptor stimulation. OBJECTIVES: To investigate the influence of the D1 receptor agonists levodopa and rotigotine compared with placebo on homocysteine and cysteinyl-glycine in plasma of patients with Parkinson's disease. METHODS: Patients received 100 mg levodopa, 4 mg rotigotine or placebo. Cysteinyl-glycine and homocysteine were measured every 30 min over three hours. RESULTS: Homocysteine rose during levodopa- and placebo administration. Rotigotine had no effect. Cysteine-glycine only increased after placebo- but not after levodopa- or rotigotine. DISCUSSION: Homocysteine elevation results from hepatic and gastrointestinal methylation processes. Transdermal rotigotine circumvents these methylation locations. Turnover of segregated alkyl residuals from rotigotine serves as methyl group donors, which counteract homocysteine increment. The placebo-related cysteinyl-glycine increase results from reduced free radical exposure. Low levodopa dosing and antioxidants in the rotigotine patch matrix prevented cysteinyl-glycine fall.

Beneficial effects of natural phenolics on levodopa methylation and oxidative neurodegeneration.

Levodopa (L-DOPA) is widely used for symptomatic management in Parkinson's disease. We recently showed that (-)-epigallocatechin-3-gallate, a tea polyphenol, not only inhibits L-DOPA methylation, but also protects against oxidative hippocampal neurodegeneration. In the present study, we sought to determine several other common dietary phenolics, namely, tea catechins [(+)-catechin and (-)-epicatechin] and a representative flavonoid (quercetin), for their ability to modulate L-DOPA methylation and to protect against oxidative hippocampal injury. A combination of in vitro biochemical assays, cell culture-based mechanistic analyses, and in vivo animal models was used. While both tea catechins and quercetin strongly inhibit human liver catechol-O-methyltransferase (COMT)-mediated O-methylation of L-DOPA in vitro, only (+)-catechin exerts a significant inhibition of L-DOPA methylation in both peripheral compartment and striatum in rats. The stronger in vivo effect of (+)-catechin on L-DOPA methylation compared to the other dietary compounds is due to its better bioavailability in vivo. In addition, (+)-catechin strongly reduces glutamate-induced oxidative cytotoxicity in HT22 mouse hippocampal neurons in vitro through inactivation of the nuclear factor-κB signaling pathway. Administration of (+)-catechin also exerts a strong neuroprotective effect in the kainic acid-induced oxidative hippocampal neurodegeneration model in rats. In conclusion, (+)-catechin is a dietary polyphenolic that may have beneficial effects in L-DOPA-based treatment of Parkinson patients by inhibiting L-DOPA methylation plus reducing oxidative neurodegeneration.

Protein oxidation injury occurs during pediatric cardiopulmonary bypass.

OBJECTIVE: Proteins are the major effectors of biological structure and function. Oxidation-induced changes to protein structure can critically impair protein function, with important pathologic consequences. This study was undertaken to examine whether oxidation-induced changes to protein structure occur during pediatric cardiopulmonary bypass and to examine the association with postoperative outcome. METHODS: Elevation of the 3,4-dihydroxyphenylalanine content of a protein relative to its native tyrosine content indicates structural damage due to oxidation. Protein 3,4-dihydroxyphenylalanine/native tyrosine ratios were measured before surgery and up to 6 hours after institution of cardiopulmonary bypass in 24 children undergoing repair of congenital heart disease, who were prospectively selected to form a cyanotic and comparable acyanotic control group. Results were correlated with perioperative variables and postoperative outcomes. RESULTS: Elevation of protein 3,4-dihydroxyphenylalanine/tyrosine ratios above baseline (0.48 mmol/mol [SD, 0.11 mmol/mol] vs 0.36 mmol/mol [SD, 0.13 mmol/mol]; P = .001) occurred within 30 minutes of initiating cardiopulmonary bypass in cyanotic but not in acyanotic children and correlated inversely with preoperative arterial oxygen saturation (R = -0.52; P = .03). Protein 3,4-dihydroxyphenylalanine/tyrosine ratios were also increased above baseline at 120 minutes (0.44 mmol/mol [SD, 0.12 mmol/mol]; P = .007) and 180 minutes (0.40 mmol/mol [SD, 0.14 mmol/mol]; P = .01) after the institution of cardiopulmonary bypass in children who underwent prolonged procedures. Elevation of 3,4-dihydroxyphenylalanine/tyrosine during prolonged procedures was associated with postoperative arrhythmias and the need for increased inotropic support (P = .001). CONCLUSIONS: Oxidative injury to proteins occurs during pediatric cardiopulmonary bypass. Cyanotic children are most at risk, particularly those undergoing prolonged procedures, in whom elevation of the protein 3,4-dihydroxyphenylalanine/tyrosine ratio is associated with increased postoperative morbidity.

Biochemical and clinical manifestations of dopamine-producing paragangliomas: utility of plasma methoxytyramine.

Measurements of plasma-free normetanephrine and metanephrine provide a sensitive test for diagnosis of pheochromocytoma but may fail to detect tumors that produce predominantly dopamine. Such tumors are extremely rare, usually found as extraadrenal paragangliomas. This report describes measurements of plasma concentrations of free methoxytyramine, the O-methylated metabolite of dopamine, in 120 patients with catecholamine-producing tumors, including nine with extraadrenal paragangliomas secreting predominantly dopamine. In seven of these nine patients, tumors were found incidentally or secondary to the space-occupying complications of the lesions. Plasma concentrations of free methoxytyramine and dopamine were increased in all nine patients, including two with normal plasma and urinary normetanephrine and metanephrine and normal urinary outputs of dopamine. Relative increases above normal for plasma methoxytyramine (104-fold) and dopamine (56-fold) were much greater (P < 0.001) than those for urinary dopamine (3-fold). Insensitivity of the latter for identification of dopamine-secreting tumors was due to dependence of the urinary amine on renal extraction and decarboxylation of circulating 3,4-dihydroxyphenylalanine. Measurements of plasma-free methoxytyramine, in addition to normetanephrine and metanephrine, are unlikely to improve diagnosis of pheochromocytomas in hypertensive patients with symptoms of catecholamine excess but may be useful in selected patients for identification of tumors that produce predominantly dopamine.

Rectal malignant melanoma diagnosed by N-isopropyl- p-123I-iodoamphetamine single photon emission computed tomography and 5-S-cysteinyl dopa: report of a case.

Malignant melanoma in the anorectal region is a rare disease associated with a very poor prognosis. Taking a biopsy of malignant melanoma is generally contraindicated because of the high risk of inducing metastasis. Although clinical examination and imaging findings are important for the preoperative diagnosis, conventional imaging techniques sometimes fail to provide information from which an accurate diagnosis can be made. We recently treated an 84-year-old woman with rectal malignant melanoma, in whom magnetic resonance imaging showed atypical findings. On the other hand, N-isopropyl- p-(123)I-iodoamphetamine single photon emission computed tomography and 5- S-cysteinyl dopa in blood serum, as a tumor marker of malignant melanoma, proved very effective for establishing the preoperative diagnosis. Despite radical abdominoperineal resection, the patient died of multiple liver and lung metastases about 5 months after surgery.

3-OMD and homocysteine plasma levels in parkinsonian patients.

One main metabolizing pathway of levodopa is O-methylation to 3-O-methyldopa (3-OMD) by catechol-O-methyltransferase (COMT). Since COMT requires Mg2+ and S-adenosylmethionine as methyl donor for this transmethylating process, COMT converts S-adenosylmethionine to S-adenosylhomocysteine and subsequent homocysteine. Objective of this study was to demonstrate relations between plasma levodopa, 3-OMD and total homocysteine in treated parkinsonian subjects. We measured homocysteine, levodopa and 3-OMD by HPLC. We compared plasma homocysteine in two groups of treated parkinsonian subjects subdivided according to their 3-OMD level. Homocysteine was significantly (p = 0.002) elevated in the group with higher 3-OMD concentrations and positively (r = 0.52, p = 0.0006) correlated to 3-OMD. Homocysteine induces vascular disease. Previous studies showed an increase of ischaemic heart- and cerebrovascular disease in treated parkinsonian patients.

Evaluation of plasma 3,4-dihydroxyphenylacetic acid (DOPAC) and plasma 3,4-dihydroxyphenylalanine (DOPA) as tumor markers in children with neuroblastoma.

Catecholamines and their metabolites are important in the diagnosis of neuroblastoma (NB). Plasma (p-) levels of 3,4-dihydroxyphenylalanine (DOPA) are increased in most NB, probably reflecting decreased DOPA decarboxylase activity. Urine (u-) homovanillic acid (HVA), a DOPA and dopamine (DA) metabolite. is also increased in most NB. DOPAC (3,4-dihydroxyphenylacetic acid) is an important metabolite of DA in tissues with monoamine oxidase (MAO) activity. Because MAO is expressed in NB tumor cells, we studied the importance of measuring p-DOPAC and p-DOPA as compared to u-HVA and u-vanillylmandelic acid (VMA) in the diagnosis and follow-up of NB. DOPAC, DOPA, dopamine, noradrenaline, adrenaline, VMA and HVA were measured by reverse-phase HPLC with electrochemical detection in 106 children (28 with NB (13 newly diagnosed), 25 with other solid tumors, 28 hospitalized for nonneoplastic diseases, and 25 healthy children). P-DOPAC or p-DOPA concentrations were above the upper normal range in 92% of untreated NB patients, as were u-HVA or u-VMA levels. None of these tumor markers was correlated to tumor stage or survival. P-DOPA but not p-DOPAC was correlated to age in NB children. Increased values of p-DOPAC and p-DOPA were found in one patient surviving NB for 10 years. Plasma DOPAC concentrations were decreased in children hospitalized for non-NB diseases, probably reflecting reduced food intake. Plasma analyses of DOPA and DOPAC seem to be useful alternatives in the diagnosis and follow-up of NB if urine sampling is to be avoided. Plasma DOPAC may be an index of nutritional status in various diseases.

5-S-cysteinyldopa as diagnostic tumor marker for uveal malignant melanoma.

PURPOSE: To evaluate the clinical significance of 5-S-cysteinyldopa (5-S-CD), a major intermediate in melanin synthesis, as a potential diagnostic tumor marker for uveal malignant melanoma. METHODS: The levels of 5-S-CD in the serum were measured by high-performance liquid chromatography in 16 patients with primary uveal melanoma. The levels of 5-S-CD were also measured in both aqueous and vitreous humor in 10 patients with uveal melanoma. The serum of healthy volunteers and patients with skin diseases other than melanoma, and the intraocular fluids of patients with cataract and vitreoretinal diseases were used as controls. RESULTS: Serum concentrations of 5-S-CD in patients with uveal melanoma in the absence of extraocular metastases were close to those of controls; however, serum concentrations of 5-S-CD were significantly elevated in patients with extraocular metastases of melanoma. Concentrations of 5-S-CD in the intraocular fluids, especially vitreous humor, were higher in patients with uveal melanoma than in controls. CONCLUSIONS: 5-S-CD in intraocular fluids may serve as a useful biochemical marker for the diagnosis of uveal melanoma. Serum 5-S-CD may contribute to the assessment of the presence and progression of extraocular metastases in patients with uveal melanoma.

Prospective evaluation of circulatory levels of catecholamines and serotonin in neuroleptic malignant syndrome.

OBJECTIVE: Neuroleptic malignant syndrome (NMS) may be associated with a dysregulation of the catecholaminergic and serotonergic systems. The objective of the present study was to evaluate prospectively the circulatory levels of serotonin (5-HT), epinephrine (E) and dopa in patients suffering from NMS. METHOD: Platelet-poor plasma (PPP) levels of serotonin, epinephrine and dopa in eight NMS patients were measured twice: in the acute state and in the state of remission. RESULTS: PPP dopa concentration was significantly lower in acute NMS state compared to the remission state (P = 0.023). In contrast, PPP E level was significantly higher (P = 0.019) in the acute NMS state and PPP 5-HT concentrations in the acute state tended to be higher than those at remission (P = 0.078). 5-HT/dopa ratio was significantly higher in the acute NMS (P = 0.015). CONCLUSION: These results may reflect reduction in dopaminergic function and increase in adrenergic and serotonergic activity in the acute NMS state.

Pharmacokinetics of radiotracers in human plasma during positron emission tomography.

Many radiopharmaceuticals for positron emission tomography (PET) are substantially metabolized in peripheral organs. Pharmacological treatments intended to alter cerebral metabolism might also alter radiotracer metabolism, consequently altering the cerebral uptake. First-order rate constants for the metabolism of PET tracers can be calculated by a linear graphical method from the precursor and metabolite concentrations measured in plasma extracts fractionated by HPLC. We tested the effects of specific pharmacological challenges on the plasma kinetics of six tracers used for PET studies of neurotransmission. The rate of O-methylation of circulating [(18)F]fluorodopa, a tracer of dopa decarboxylase activity in brain, was unaffected by pretreatment with amantadine, an antagonist of glutamate receptors. [(11)C]Deprenyl, a tracer of monoamine oxidase activity, was rapidly metabolized to [(11)C]methamphetamine and polar metabolites in healthy volunteers. The net rate constant of this metabolism was three times higher in a group of subjects under treatment for epilepsy, consistent with induction of hepatic microsomal enzymes by antiepileptic drugs. [(11)C]Sch 23390, a ligand for dopamine D1 receptors, was rapidly metabolized to polar metabolites. The net rate constant of metabolism was unaffected by pretreatment with lorazepam. [(11)C]-(S)-Nicotine, a ligand for nicotinic receptors, was rapidly metabolized to [(11)C]-(S)-cotenine, which is less polar than the parent compound. Pretreatment with mazindol, a dopamine uptake inhibitor, was without effect on peripheral metabolism of [(11)C]-(S)-nicotine. [(11)C]WIN 35,428, a tropane derivative which labels dopamine uptake sites, was metabolized to a nonpolar metabolite, but so slowly that the rate constant of this process could not be calculated. [(11)C]Raclopride, a ligand for dopamine D2 receptors, was first metabolized to a nonpolar metabolite, which then yielded two hydrophilic metabolites. The initial metabolic step was substantially blocked by pretreatment with amphetamine, possibly indicative of competitive inhibition of microsomal oxidation. Together, these results indicate that the linear graphic method is useful for estimating the kinetics of the plasma metabolism of many widely used PET tracers. Drug-drug interactions were revealed in subjects treated with specific pharmacological agents prior to tracer administration.

[Determination of serum S-100 protein and 5-S-cysteinyl-DOPA levels in melanoma patients]. / S-100 protein és 5-S-ciszteinil-DOPA vizsgálata melanomás betegekben.

This was the first time that authors detected se-S-100 and 5-SCD values with patients with malignant melanoma in Hungary. They examined the change of serum S-100 and 5-SCD value parallel. Sera were obtained with 184 melanoma patients 326 times. Patients were ranked into groups on the basis of clinical symptoms: free of symptoms and suffering from it (primary tumour, regional lymph node metastasis, soliter or multiplex distant metastasis). On the basis of the initial results the following have been found: S-100 protein and 5-SCD serum levels had no prognostic value in patients with primary melanoma. Patients without symptoms showed values around the normal level. There was significant difference in both markers between patients with or without symptoms. Significant differences were found between clinical stage I and II, as well as in clinical stage II and III. In the case of S-100 protein there was significant difference between the values of patients with soliter and multiplex distant metastasis.

Peripheral catecholamine alterations in adolescents with polycystic ovary syndrome.

OBJECTIVE: Polycystic ovary syndrome (PCO) is one of the most common endocrine disorders affecting women. Several lines of evidence have suggested the involvement of the sympathetic nervous system (SNS) in this condition. The present work was designed to assess neurochemically SNS activity in patients during the early stages of PCO. DESIGN AND PATIENTS: Fourteen patients with PCO (aged 14 to 21 years) were studied on a random day and 9 normal regularly cycling adolescents (aged 14 to 20 years) were studied during the early follicular phase (days 2 to 5). MEASUREMENTS: Hormonal profile was determined in basal conditions. LH and FSH were also measured after i.v. administration of 100 micrograms GnRH. Plasma concentrations of dihydroxyphenylalanine (Dopa), noradrenaline (NA), adrenaline (A), total dopamine (DA) and dihydroxyphenylglycol (DHPG) were determined in basal conditions and in response to GnRH by HPLC with electrochemical detection or a radioenzymatic method. Basal urinary Dopa, catecholamines and catechol metabolites (DHPG, vanillylmandelic acid (VMA), 3-methoxy-4-hydroxyphenylglycol (MHPG), homovanillic acid (HVA), metanephrine (MN) and normetanephrine (NMN)) were determined by HPLC with electrochemical detection. RESULTS: Basal plasma LH, testosterone, androstenedione, oestrone and LH/FSH ratio were higher (P < 0.01) and serum sex hormone-binding globulin (SHBG) were lower (P < 0.01) in PCO patients than in control subjects. Basal and GnRH-stimulated plasma free Dopa, A, NA and total DA were similar in patients and controls. Plasma DHPG was lower (P < 0.01) in PCO patients (4.20 +/- 0.30 nmol/l) than in controls (8.0 +/- 1.0 nmol/l) throughout the study. Urinary A, NA, DA, Dopa, MN, MHPG, HVA, and VMA were similar in patients and controls. Urinary DHPG was lower (P < 0.01) in PCO patients (0.50 +/- 0.02 mumol/d) than in controls (0.73 +/- 0.09 mumol/d). On the other hand PCO patients had a higher urinary excretion of NMN than controls (PCO: 1.20 +/- 0.10; C: 0.78 +/- 0.10 mumol/d, P < 0.05). CONCLUSIONS: Our results show the same endocrinological features in adolescent PCO patients as those reported in adults. The results also demonstrate a peripheral catecholaminergic alteration which suggests an alteration in noradrenaline deamination and/or uptake in adolescent patients. This study however does not permit us to conclude that PCO is primarily caused by this sympathetic alteration.

Pallidotomy improves motor responses and widens the levodopa therapeutic window in Parkinson's disease.

Stereotactic posteroventral pallidotomy (PVP) as a treatment for Parkinson's disease (PD) symptoms has been increasingly used in moderate-advanced disease. We examined the pharmacodynamic responses of PD patients to single oral levodopa doses and intravenous levodopa infusions before and after PVP surgery. Nine subjects with advanced PD received a single oral dose and ramped intravenous levodopa infusions before and 3-5 weeks after unilateral PVP. Timed motor tasks, Unified Parkinson's Disease Rating Scale (UPDRS) evaluations, and ordinal dyskinesia rating were performed after oral levodopa and during i.v. levodopa infusions. Serum prolactin and dopa levels were measured during the levodopa infusions. Overall timed motor but not motor UPDRS scores were improved after PVP in both the worst ("off") and best ("on") states. Contralateral but not ipsilateral limb dyskinesias were substantially reduced at all serum (dopa) levels after PVP. Ipsilateral and contralateral timed motor performance at low serum (dopa) levels was improved by PVP. Walking speeds at all serum (dopa) levels were not changed by PVP. Serum prolactin was reduced equally by increasing (dopa) preoperatively and postoperatively. PVP significantly and favorably altered oral and intravenous levodopa pharmacodynamics by improving bilateral limb motor function and contralateral dyskinesia but did not alter walking speed. PVP appears to widen significantly the therapeutic window for levodopa in PD.

Generalized melanosis in metastatic malignant melanoma: the possible role of DOPAquinone metabolites.

Generalized melanosis occurs very rarely as a complication of malignant melanoma, and the pathogenesis of this condition is still unclear. Histological examination of pigmented skin and measurements of the DOPAquinone metabolites 5-S-cysteinyldopa (5-S-CD) and 6-hydroxy-5-methoxyindole-2-carboxylic acid (6H5MI2C) in the patient's serum and urine were carried out. Histological examination revealed basal hyperpigmentation, discrete melanoma cells and melanophages around the blood vessels and an unusual melanin deposition within collagen bundles in the dermis. The levels of 5-S-CD and 6H5MI2C were dramatically increased both in the patient's serum and urine. The deposition of DOPAquinone metabolites secreted by the melanoma cells may contribute to the unusual melanin deposition within collagen bundles in the affected dermis.

Determination of unchanged [18F]dopamine in human and non-human primate plasma during positron emission tomography studies: a new solid-phase extraction method comparable to radio-thin-layer chromatography analysis.

Routine determination of [18F]DOPA and its metabolites in plasma is essential for assessment and quantification of presynaptic dopamine function in vivo using a modeling approach with positron emission tomography (PET). The determination of unchanged [18F]DOPA from human and non-human primate plasma using solid-phase extraction (SPE) with Sep-Pak cartridges during PET dopaminergic studies is described here. The results from the studies showed that this new approach in comparsion to a method such as thin-layer chromatography (TLC) possessed a simplicity, rapidity and accuracy as well as good correlation between the two techniques (p<0.0001). A proposed procedure involving radioanalysis on alumina plates (Al2O3) was also developed with an excellent correlation compared to the conventional C18 plates (r=0.96). Thus it could be concluded that the SPE on either C18 or alumina cartridges (Waters) compared to radio-TLC analysis on C18 and alumina systems, appears to be a useful analytical method suitable for correcting the input arterial function in routine clinical PET neurotransmission studies.

Malignant phaeochromocytoma with high circulating DOPA, and clonidine-suppressible noradrenaline.

Phaeochromocytoma, "perhaps the most fascinating of all tumors" [1], can present with a broad range of clinical manifestations [2]. Once suspected, the biochemical diagnosis is straightforward in most patients since plasma and urinary levels of noradrenaline and/or adrenaline, and urinary metabolites are well above those encountered in healthy subjects or patients with essential hypertension [3,4]. Exceptions to this general rule are well known, however, hence suppression tests have found favour, particularly in cases where catecholamine levels are within, or close to, the normal range [5-7]. We present a unique patient with malignant phaeochromocytoma whose plasma noradrenaline levels were in the high-normal range, and suppressed normally with clonidine administration. He had extremely high circulating levels of dihydroxyphenylalanine (dopa) which were not affected by clonidine, and different patterns of catecholamine concentrations in tumour tissue and plasma.

3,4-dihydroxyphenylalanine (DOPA) decarboxylase deficiency and resultant high levels of plasma DOPA and dopamine in unfavorable neuroblastoma.

Neuroblastoma (NB) is a tumor which arises from neural crest cells. In the developing neural crest cells, the induction of 3,4-dihydroxyphenylalanine (DOPA) decarboxylase is more delayed than that of tyrosine hydroxylase and dopamine-beta-hydroxylase. If NB cells are arrested in an early stage of neural crest development, the induction of DOPA decarboxylase is insufficient and the accumulation and secretion of DOPA can be caused. The biochemically immature phenotype is thought to represent the undifferentiated characteristics of the cells and might correlate with the grade of malignancy. To investigate whether the hypothesis is clinically applicable or not, we have measured plasma DOPA, dopamine and urinary catecholamine metabolites in NB patients. The levels of plasma DOPA, dopamine, urinary homovanillic acid (HVA) and vanillactic acid (VLA) were significantly higher in patients with unfavorable NBs and the higher plasma DOPA level was significantly associated with the patients' age (> 1 year old), tumor stage (III, IV) and DNA diploidy. Serial determination of plasma DOPA was a good monitor of the disease course. These results are compatible with the hypothesis on DOPA decarboxylase deficiency and DOPA secretion in undifferentiated, unfavorable NBs. In conclusion, the plasma DOPA can be used to predict patients' prognosis as well as to follow up patients with NB.

Occipital horn syndrome and a mild Menkes phenotype associated with splice site mutations at the MNK locus.

We have found mutations in the Menkes disease gene (MNK) which impair, but do not abolish, correct mRNA splicing in patients with less severe clinical phenotypes. In one family, four males aged 2-36 years with a distinctive Menkes variant have a mutation at the +3 position of a splice donor site near the 3' end of the Menkes coding sequence that is associated with exon skipping and a stable mutant transcript. In an unrelated 15-year-old male with typical occipital horn syndrome, a point mutation at the -2 exonic position of a splice donor site in the middle of the gene causes exon-skipping and activation of a cryptic splice acceptor site. In both mutations, maintenance of some normal splicing is demonstrable by RT-PCR, cDNA sequencing and ribonuclease protection.

Clinical application of noradrenaline spillover methodology: delineation of regional human sympathetic nervous responses.

The proportionality which in general exists between rates of sympathetic nerve firing and the overflow of noradrenaline into the venous drainage of an organ provides the experimental justification for the use of measurements of noradrenaline in plasma as a biochemical measure of sympathetic nervous function. Static measurements of noradrenaline plasma concentration have several limitations. One is the confounding influence of noradrenaline plasma clearance on plasma concentration. Other drawbacks include the distortion arising from antecubital venous sampling (this represents but one venous drainage, that of the forearm), and the inability to detect regional differentiation of sympathetic responses. Clinical regional noradrenaline spillover measurements, performed with infusions of radiolabelled noradrenaline and sampling from centrally placed catheters, and derived from regional isotope dilution, overcome these deficiencies. The strength of the methodology is that sympathetic nervous function may be studied in the internal organs not accessible to nerve recording with microneurography. Examples of the regionalization of human sympathetic responses disclosed include the preferential activation of the cardiac sympathetic outflow with mental stress, cigarette smoking, aerobic exercise, cardiac failure, coronary insufficiency, essential hypertension and in ventricular arrhythmias, and the preferential stimulation or inhibition of the renal sympathetic nerves with low salt diets and mental stress, and with exercise training, respectively. By application of the same principles, regional release of the sympathetic cotransmitters neuropeptide Y and adrenaline can be studied in humans. Cotransmitter release, however, is detected only with some difficulty. In restricted circumstances we find evidence of regional cotransmitter release to plasma, such as the release of neuropeptide Y from the heart at the very high rates of sympathetic nerve firing occurring with aerobic exercise, and cardiac adrenaline release also with exercise and after loading of the neuronal adrenaline pool by intravenous infusion of adrenaline.