Cardiac Spillover Research Articles

Abstract 651: Myocardial NEFA Uptake is Enhanced to a Greater Extent Than NEFA Oxidation in Conscious Senescent Beagles with Myocardial Insulin Resistance

Background : Prior studies have demonstrated that aging is associated with myocardial insulin resistance, mitochondrial ultra-structural changes and decreased UCP-3 protein. We sought to determine the relationship between these metabolic limitations and myocardial NEFA uptake and oxidation in senescent beagles. Methods : Eight young (age 3– 4 years) and 8 senescent (age 11–12 years) beagles underwent chronic instrumentation for measurement of hemodymics and myocardial substrate balance. All dogs were subjected to an 8 week exercise training program prior to instrumentation to control for activity level. In the fasting, conscious state, myocardial NEFA uptake and oxidation was measured using 3 H-oleate, and insulin resistance was assessed via hyperinsulinemic-euglycemic clamps. Samples of LV myocardium were harvested at euthanasia to measure components of NEFA transport. Results : There was no difference in body weight or adiposity between groups. Older beagles had marked (p<0.01) increases in plasma NEFA (Y:426± 65; O:902±78 μMol/min) and increased cardiac NE spillover (Y:21±4; O: 221±36 pmol/min). However, myocardial external efficiency was reduced (p<0.05) in older dogs (Y:15±3; O: 9±3 g·m 2 /ml O 2 /min). Hearts from older beagles demonstrated significant increases (p<0.01) in NEFA uptake (Y: 5.2±0.9; O: 9.3±0.8 μMol/min/100g), and only modest increases in oxidation (Y: 4.9±0.6; O: 7.0±1.1 μMol/min/ 100g). In sarcolemmal membranes, expression of Fatty Acid Transport Protein-1 (FATP-1) was upregulated (Y:72 11 DU; O: 235 21 DU, p<0.05), whereas that of FAT/CD36 was markedly reduced (Y: 1,312±123; O: 387±12 DU, p<0.001). There was no difference in PPARα (Y: 425±25; O: 475±116 DU) or PGC-1α (Y: 637±53; O: 633±14 DU) expression. The cumulative result of these changes was an accumulation of neutral lipids within older hearts and significant (p<0.02) insulin resistance (Y: 8.3 ±0.6; O: 4.3±0.8 μMol/min/100g). Conclusion : In older beagles, myocardial NEFA uptake exceeds oxidation, resulting in intramyocardial lipid accumulation. Increased FA uptake occurs despite decreased FAT/CD36 expression, and may be mediated by an upreguation of FATP-1 expression. These changes are seen in association with progressive myocardial insulin resistance.

Acute electrophysiologic effects of intravenous amiodarone are independent of a sympatholytic action in humans.

Previous experiments in animals demonstrated a novel sympatholytic action of acute intravascular amiodarone (AM). It is not known if this action also occurs in humans. Twelve male volunteers performed handgrip for 10 min before and after 300 mg intravenous (IV) AM over 60 min. The effect of handgrip was determined from changes in blood pressure (BP), heart rate (HR), and cardiac noradrenaline (NA) spillover. Changes in cardiac spillover of dihydroxyphenylglycol (DHPG), the metabolite of NA, were measured during AM infusion. The electrophysiological effects of AM were determined from changes to the A-H intervals during right atrial stimulation (100 beats/min). Handgrip increased HR (63 +/- 2 to 84 +/- 5 beats/min and 65 +/- 3 to 84 +/- 4 beats/min), systolic BP (141 +/- 4 to 179 +/- 6 mm Hg and 140 +/- 4 to 179 +/- 7 mm Hg), and cardiac NA spillover (11.9 +/- 4 to 44.3 +/- 13 ng/min and 17.3 +/- 4 to 55.5 +/- 11 ng/min) before and after AM, respectively (P < 0.02 in all groups). There was good correlation between increases in cardiac NA spillover and HR (r2 = 0.86) and systolic BP (r2 = 0.87). AM increased the A-H interval (95.5 +/- 18 to 107.8 +/- 20 ms, P < 0.02). There was no difference in hemodynamic or NA response to handgrip before or after the AM infusion. There was also no change in DHPG cardiac spillover during AM infusion. Acute IV AM did not exert a sympatholytic action in humans, with no attenuation in hemodynamic or NA response to handgrip or increase in DHPG production, despite producing an electrophysiologic response.

Cardiac-specific overexpression of tumor necrosis factor-alpha causes oxidative stress and contractile dysfunction in mouse diaphragm.

We have developed a transgenic mouse with cardiac-restricted overexpression of tumor necrosis factor-alpha (TNF-alpha). These mice develop a heart failure phenotype characterized by left ventricular dysfunction and remodeling, pulmonary edema, and elevated levels of TNF-alpha in the peripheral circulation from cardiac spillover. Given that TNF-alpha causes atrophy and loss of function in respiratory muscle, we asked whether transgenic mice developed diaphragm dysfunction and whether contractile losses were caused by oxidative stress or tissue remodeling. muscles excised from transgenic mice and littermate controls were studied in vitro with direct electrical stimulation. Cytosolic oxidant levels were measured with 2', 7'-dichlorofluorescin diacetate; emissions of the oxidized product were detected by fluorescence microscopy. Force generation by the diaphragm of transgenic animals was 47% less than control (13.2+/-0. 8 [+/-SEM] versus 25.1+/-0.6 N/cm(2); P:<0.001); this weakness was associated with greater intracellular oxidant levels (P:<0.025) and was partially reversed by 30-minute incubation with the antioxidant N:-acetylcysteine 10 mmol/L (P:<0.01). Exogenous TNF-alpha 500 micromol/L increased oxidant production in diaphragm of wild-type mice and caused weakness that was inhibited by N:-acetylcysteine, suggesting that changes observed in the diaphragm of transgenic animals were mediated by TNF-alpha. There were no differences in body or diaphragm weights between transgenic and control animals, nor was there evidence of muscle injury or apoptosis. Elevated circulating levels of TNF-alpha provoke contractile dysfunction in the diaphragm through an endocrine mechanism thought to be mediated by oxidative stress.

Antiadrenergic effect of chronic amiodarone therapy in human heart failure

OBJECTIVESThe aim of the present study was to evaluate the influence of amiodarone on neurochemical parameters of sympathetic nervous activity in patients with congestive heart failure.BACKGROUNDUnlike most antiarrhythmic agents, amiodarone has been shown to exert a beneficial effect on survival in some studies of patients with congestive heart failure. The pharmacology of this agent is complex, and as such, the mode of its action is unclear in humans. Some experimental studies suggest that amiodarone exerts a sympatholytic effect.METHODSTo evaluate the effect of amiodarone on sympathetic nervous activity, we measured the total systemic and cardiac norepinephrine (NE) spillover rate by isotope dilution in 58 patients with severe heart failure (left ventricular ejection fraction 20 ± 1%), 22 of whom were receiving chronic amiodarone treatment. Release rates for dihydroxyphenylalanine (DOPA, a precursor of NE), and endogenous and radiolabeled dihydroxyphenylglycol (DHPG and 3H-DHPG, intraneuronal metabolites of NE and 3H-NE, respectively) were also determined to assess sympathetic neuronal integrity.RESULTSAmiodarone-treated patients had significantly lower cardiac spillover rates for NE (42%, p = 0.001), DOPA (74%, p < 0.001), DHPG (44%, p < 0.01) and 3H-DHPG (51%, p < 0.01) than those patients not treated with amiodarone. Hemodynamic assessment of amiodarone-treated patients revealed higher cardiac output (4.4 ± 0.2 vs. 3.7 ± 0.2 liters/min, p < 0.01), and slightly lower pulmonary capillary wedge pressure (18 ± 2 vs. 22 ± 1, p = NS) than in untreated patients. After correction for the potential confounding effect of hemodynamic differences, amiodarone-treated patients continued to demonstrate significantly lower spillover rates of NE, DOPA and DHPG from the heart.CONCLUSIONSThese data indicate that amiodarone may exert beneficial effects on the failing human heart through a sympatholytic process, and this action appears to be relatively cardioselective.

Cardiorenal epinephrine kinetics: evidence for neuronal release in the human heart.

There are experimental data suggesting that epinephrine (Epi) may act as a cotransmitter in sympathetic nerves, stimulating presynaptic beta 2-receptors and enhancing norepinephrine (NE) release. To examine neuronal Epi release, patients with congestive heart failure and hypertension and healthy subjects were examined with the isotope-dilution method. At baseline, small cardiac and renal Epi spillovers were found in patients. During intense supine exercise, cardiac NE and Epi spillovers increased concomitantly with similar magnitude, whereas no renal Epi spillover could be detected. Blockade of neuronal uptake 1 caused a consistent decrease in both cardiac and renal fractional extractions of NE and Epi. The present study demonstrates baseline cardiorenal Epi release in patients with congestive heart failure and renal Epi release in hypertensive patients. Furthermore, Epi is removed by neuronal uptake in both the heart and kidney, and cardiac Epi spillover increases during exercise. This study, in contrast to other results, provides evidence for cardiac neuronal Epi release.

Cardiac sympathetic responses to acute vasodilation. Normal ventricular function versus congestive heart failure.

Although there is indirect evidence of impaired baroreflex control of sympathetic outflow directed at heart muscle, the regulation of cardiac sympathetic activity in the setting of heart failure is largely unexplored. We used the norepinephrine spillover method to address the hypothesis that baroreflex control of cardiac sympathetic activity is reduced in heart failure. Twenty-three patients were studied, 17 in a group with heart failure and 6 in a group with normal ventricular function. In both groups, cardiac norepinephrine spillover was assessed in response to nitroprusside infused to steady-state conditions. Nitroprusside resulted in significant reductions in mean systemic arterial pressure (normal group, -15 +/- 3% [mean +/- SEM]; heart failure group, -13 +/- 1%) and mean pulmonary artery pressure (normal group, -25 +/- 10%; heart failure group, -29 +/- 4%). In response to nitroprusside, there was a 98 +/- 16% increase in cardiac norepinephrine spillover in the normal group (from 81 +/- 10 to 159 +/- 25 pmol/min, P < .05). Despite similar hemodynamic responses to nitroprusside in the heart failure group, there was only a 28 +/- 14% increase in cardiac spillover (from 211 +/- 71 to 245 +/- 59 pmol/ min, P = NS), a response that was significantly smaller than that seen in the normal group. In patients with heart failure compared with subjects with normal ventricular function, there was a significantly smaller increase in cardiac sympathetic activity in response to a steady-state infusion of nitroprusside. This result provides evidence for reduced baroreflex control of cardiac sympathetic activity in heart failure.

Cardiac sympathetic nerve function in congestive heart failure.

Increased availability of norepinephrine (NE) for activation of cardiac adrenoceptors (increased cardiac adrenergic drive) and depletion of myocardial NE stores may contribute to the pathophysiology and progression of congestive heart failure. This study used a comprehensive neurochemical approach to examine the mechanisms responsible for these abnormalities. Subjects with and without congestive heart failure received intravenous infusions of [(3)H]NE. Cardiac spillover, reuptake, vesicular-axoplasmic exchange, and tissue stores of NE were assessed from arterial and coronary venous plasma concentrations of endogenous and [(3)H]-labeled NE and dihydroxyphenylglycol. Tyrosine hydroxylase activity was assessed from plasma dopa, and NE turnover was assessed from measurements of NE metabolites. NE release and reuptake were both increased in the failing heart; however, the efficiency of NE reuptake was reduced such that cardiac spillover of NE was increased disproportionately more than neuronal release of NE. Cardiac NE stores were 47% lower and the rate of vesicular leakage of NE was 42% lower in the failing than in the normal heart. Cardiac spillover of dopa and NE turnover were increased similarly in congestive heart failure. Increased neuronal release of NE and decreased efficiency of NE reuptake both contribute to increased cardiac adrenergic drive in congestive heart failure. Decreased vesicular leakage of NE, secondary to decreased myocardial stores of NE, limits the increase in cardiac NE turnover in CHF. Decreased NE store size in the failing heart appears to result not from insufficient tyrosine hydroxylation but from chronically increased NE turnover and reduced efficiency of NE reuptake and storage.

Neurochemical evidence of cardiac sympathetic activation and increased central nervous system norepinephrine turnover in severe congestive heart failure

Objectives. The aim of this study was to characterize cardiac sympathetic nervous function in patients with severe heart failure and to investigate the influence of the cause of heart failure, hemodynamic variables and central nervous system catecholamine release on cardiac sympathetic tone.Background. Although heart failure is generally accompanied by sympathoexcitation, the integrity of cardiac sympathetic nerve function in heart failure remains controversial, particularly in relation to nerve firing activity and to the capacity of sympathetic nerves to recapture norepinephrine. Additionally, the location of the afferent and central neural pathways implicated in heart failure-induced sympathoexcitation remains unclear.Methods. Radiotracer techniques were applied in 41 patients with severe heart failure and 15 healthy control subjects to study the biochemical aspects of whole body and cardiac sympathetic activity. Hemodynamic indexes of cardiac performance were measured in the heart failure group, and their association with sympathetic activity was studied. Jugular venous catechol spillover was measured to study the central noradrenergic control of sympathetic outflow.Results. Sympathoexcitation was evident in the heart failure group, reflected by a 62% increase (p < 0.001) in total body and a 277% increase (p < 0.001) in cardiac norepinephrine spillover rates. These changes were accompanied by significant increases in the cardiac spillover of the norepinephrine precursor dihydroxyphenylalanine, the sympathetic cotransmitter neuropeptide Y and the extraneuronal metabolite 3-methoxy-4-hydroxyphenylglycol. The level of cardiac sympathetic activity was significantly correlated (r = 0.59, p < 0.001) with the mean pulmonary artery pressure. An increase in the spillover of dihydroxyphenylalanine and 3-methoxy-4-hydroxyphenylglycol from the brain was present, suggesting activation of central noradrenergic neurons.Conclusions. Cardiac sympathetic activation is present in severe heart failure, bearing a close relation with pulmonary artery pressures, independent of heart failure etiology. Activation of noradrenergic neurons in the brain is also present and may be the underlying central nervous mechanism of the sympathoexcitation observed in heart failure.

Silent ischemia in unstable angina is related to an altered cardiac norepinephrine handling.

Inferential evidence suggests that silent ischemia might be related to sympathetic activity. Study of [3H]norepinephrine kinetics is a suitable tool to assess the regional sympathetic activity. This method was applied to investigate whether silent myocardial ischemia in unstable angina is related to and depends on cardiac sympathetic overactivity. Patients with active unstable angina were compared with patients with inactive unstable angina, stable effort angina, and controls. Silent myocardial ischemia was evaluated by three 24-hour Holter monitoring periods on alternate days, and [3H]norepinephrine kinetics was assessed under rest conditions and following the cold pressor test. Simultaneously, catecholamine concentrations were measured in the aortic, coronary sinus, and peripheral venous blood. Different than the other groups (p = 0.0013), in patients with active unstable angina, the majority of silent ischemic episodes occurred without increase in heart rate. These patients had a positive coronary sinus-aorta norepinephrine gradient, both at rest and following the cold pressor test. [3H]Norepinephrine kinetics demonstrated an increased selective cardiac spillover, both at rest and, even more, after the cold pressor test. Reduced cardiac [3H]norepinephrine extraction also was found. A significant relation was found between the number of ischemic episodes or the overall duration of silent ischemia and norepinephrine spillover, both at rest and following cold application. During the acute phase of unstable angina (but not in the quiescent phase or in stable effort angina), a disorder in cardiac norepinephrine handling occurs. This results in a reflex cardiac sympathetic overactivity that plays a major role in the occurrence of silent myocardial ischemia.

Cardiac noradrenergic function one year following cardiac transplantation.

Controversy exists whether or not cardiac nerves regrow into a human transplanted heart. The present study examined cardiac noradrenergic function one year following cardiac transplantation. Eight male cardiac transplant recipients having standard triple immunosuppressive treatment were catheterized diagnostically one year after heart transplantation. They had normalized cardiac hemodynamics and showed no histological rejection in biopsies. A second study group, with intact cardiac innervation, consisted of 19 patients with stable angina pectoris class I to III. Cardiac sympathetic nerve activity was assessed by measurements of cardiac noradrenaline (NA) extraction and spillover, using infusion of tritium-labelled NA (3H-NA). To further characterize cardiac noradrenergic function (synthesis and metabolism) arterial and coronary venous plasma concentrations of the NA precursor, dihydroxyphenylalanine (DOPA), the dopamine metabolite, dihydroxyphenylacetic acid (DOPAC), and the intra-neuronal NA metabolite, dihydroxyphenylglycol (DHPG), were also examined. In transplant recipients, cardiac sympathetic function was also examined before and during supine bicycle exercise and after administration of desipramine to block neuronal uptake. Significant dilution of 3H-NA with endogenous NA from arterial to coronary venous plasma in cardiac transplant patients indicated detectable spillover of NA into plasma from transplanted hearts. However, cardiac spillover of NA in heart transplant patients was only a quarter of that observed in angina patients. Also, transplanted hearts showed no production of DOPA, DOPAC, DHPG or 3H-labelled DHPG, whereas there were positive arterial-venous plasma gradients in all these catechols across the hearts of angina patients. Cardiac fractional extraction of NA was 74% in the angina patients, but only 13% in transplant patients. In transplant patients, cardiac spillover of NA and extraction of 3H-NA were not affected by desipramine or exercise. The present study shows little evidence of any functional activity of cardiac sympathetic nerves in the transplanted heart, as assessed by biochemical indices of release, reuptake, metabolism and synthesis of NA. This argues against cardiac reinnervation of functional importance, at least within the first postoperative year.

Increased cardiac production of dihydroxyphenylalanine (DOPA) during sympathetic stimulation in anaesthetized dogs

Entry of dihydroxyphenylalanine (DOPA) into plasma from specific organs may reflect regional activity of tyrosine hydroxylase, the enzyme responsible for the immediate synthesis of DOPA and rate-limiting for subsequent formation of catecholamines. Therefore, cardiac spillovers of DOPA, noradrenaline and the intraneuronal metabolite of noradrenaline, dihydroxyphenylglycol (DHPG), were examined during two periods of graded electrical stimulation of the sympathetic nerves to the heart in anaesthetized dogs. Responses were examined before and after neuronal uptake blockade with desipramine. Cardiac spillover of DOPA increased by 1.8- and 4.4-fold during sympathetic stimulation before desipramine and by 1.6- and 3.3-fold after desipramine. Fold increases in cardiac spillover of DOPA were much lower than but positively related with fold increases in noradrenaline spillover (5.9- and 13.8-fold increases before and 9.0- and 15.8-fold increases after desipramine). Increases in cardiac spillover of DHPG (1.5- and 2.3-fold increases) were blocked by desipramine so that fold changes in spillover of DOPA were greater than and poorly related to changes in spillover of DHPG. Fold increases in cardiac spillover of DOPA showed a close one-to-one positive relationship with fold increases in the sum of cardiac spillovers of noradrenaline and dihydroxyphenylglycol before and after desipramine. For a given fold increase in noradrenaline release, transmitter turnover is increased fractionally and noradrenaline synthesis need also only increase fractionally to maintain transmitter stores constant. The close relationship between fold increases in cardiac spillover of DOPA and combined spillovers of noradrenaline and DHPG is consistent with regulation of tyrosine hydroxylase activity to match changes in noradrenaline synthesis with changes in noradrenaline turnover. Changes in cardiac spillover of DOPA appear to reflect local changes in tyrosine hydroxylase activity.

Sympathetic nervous function in human heart as assessed by cardiac spillovers of dihydroxyphenylglycol and norepinephrine.

Measurement of cardiac norepinephrine spillover may indicate the amount of transmitter at neuroeffector sites but does not distinguish neuronal release or reuptake in determining this amount or provide information about other aspects of sympathetic function. This report examines how cardiac spillover of the norepinephrine metabolite dihydroxyphenylglycol (DHPG) provides additional distinct information about cardiac sympathetic function. Arterial and coronary venous blood samples were taken during cardiac catheterization and intravenous infusion of [3H]norepinephrine in 57 subjects. Subjects were given intravenous yohimbine or underwent mental stress, handgrip exercise, and cycling exercise to activate sympathetic nerves or were given intravenous desipramine to block norepinephrine reuptake. Cardiac DHPG spillover (601 +/- 41 pmol/min) was eightfold greater than norepinephrine spillover (78 +/- 10 pmol/min) at rest and increased during sympathetic activation by 65% of the increase of norepinephrine. This and the desipramine-sensitive cardiac production of [3H]-labeled DHPG from [3H]norepinephrine indicated that 10.5 times more endogenous norepinephrine is recaptured than escapes into plasma; that more than 90% of recaptured norepinephrine is sequestered into storage vesicles; and that under resting conditions, most cardiac spillover of DHPG and turnover of norepinephrine are from metabolism of transmitter leaking from vesicles; the latter process is independent of exocytotic transmitter release with a rate at rest over 100-fold that of norepinephrine spillover and over 10-fold that of norepinephrine reuptake. Cardiac spillover of DHPG provides information about processes close to or within sympathetic nerve endings that cannot be provided by measurements of norepinephrine spillover alone. This includes quantitative information about the role of neuronal uptake in terminating the actions of norepinephrine at neuroeffector sites and the importance of vesicular-axoplasmic exchange of norepinephrine as a dynamic process contributing to norepinephrine turnover.

Disposition of endogenous adrenaline compared to noradrenaline released by cardiac sympathetic nerves in the anaesthetized dog.

The fate of adrenaline released from cardiac sympathetic nerves was compared with that of noradrenaline before and during two periods of electrical stimulation of the left ansa subclavia in eight anaesthetized dogs. Cardiac spillovers and extractions of both catecholamines were estimated simultaneously using infusions of 3H-labelled adrenaline and noradrenaline. Animals were studied before and after neuronal uptake blockade with desipramine. Cardiac spillover of adrenaline, detectable at rest at 1.4 +/- 0.3 pmol/min, increased to 4.0 +/- 1.1 and 5.3 +/- 1.2 pmol/min during sympathetic stimulation. Cardiac noradrenaline spillover increased from 49 +/- 12 to 205 +/- 40 and 451 +/- 118 pmol/min. After desipramine, cardiac spillovers of adrenaline were decreased, whereas those of noradrenaline were increased so that the ratio of adrenaline to noradrenaline spillover, meaned before and during stimulation, decreased substantially from 1:42 to 1:166. The desipramine-induced decrease in cardiac extractions of 3H-labelled catecholamines indicated adrenaline was removed 60% less efficiently than noradrenaline by neuronal uptake, whereas the extractions remaining indicated adrenaline was removed 50% more efficiently by extraneuronal uptake. The differences in removal processes indicated that 35% of the adrenaline released by cardiac sympathetic nerves was recaptured compared to 88% for noradrenaline, leaving 53% to be removed extra-neuronally compared to 6.6% for noradrenaline, so that proportionally more released adrenaline than noradrenaline escaped to spillover into plasma (12% versus 5.4%). Since extra-neuronal uptake was more efficient for adrenaline than noradrenaline, proportionally less released adrenaline than noradrenaline escaped local removal to spillover into plasma when neuronal uptake was blocked (17% versus 45%). This reversed the situation before blockade so that desipramine substantially decreased the ratio of adrenaline to noradrenaline spillover. Thus, differences in the efficiencies of neuronal or extraneuronal uptake are important determinants of the amounts of locally released adrenaline and noradrenaline that escape removal processes to act at neuroeffector sites or spillover into plasma.

Coronary venous noradrenaline during coronary angioplasty

Coronary venous and arterial noradrenaline concentrations were measured during percutaneous transluminal coronary angioplasty in 14 patients. Coronary venous noradrenaline did not increase significantly during balloon inflation but was increased during early reperfusion in patients undergoing left anterior descending angioplasty ( n = 8), from 157 ± 38 pg/ml to 295 ± 94 pg/ml ( P < 0.05). Coronary blood flow, measured by thermodilution in 7 further patients during left anterior descending angioplasty, was 69 ± 9 ml/min, decreased to 80 ± 3% of basal flow during ballon inflation ( P < 0.01) and increased to 135 ± 5% during early reperfusion ( P < 0.01). It was estimated using these results that cardiac spillover of noradrenaline did not change during occlusion of the left anterior descending artery, but increased almost 3-fold during early reperfusion. During the period of ballon inflations, there was a modest increase in overall sympathetic tone, as assessed by total noradrenaline spillover to plasma (400 ± 77 ng/min to 473 ± 87 ng/min, P < 0.01). These results, suggesting an increase in release of noradrenaline during early reperfusion following brief occlusion of the left anterior descending artery, may be relevant to the genesis of reperfusion arrhythmias.

Biochemical evidence of sympathetic denervation of the heart in pure autonomic failure

Primary autonomic failure is a heterogenous group of diseases with evidence for lesions in both the central and peripheral elements of the autonomic nervous system. We determined the extent of peripheral sympathetic dysfunction in six patients with primary autonomic failure without clinical evidence of central nervous system involvement (pure autonomic failure) using biochemical methods for studying regional noradrenaline spillover and removal. The results were compared with those from 14 age-matched normal subjects, seven of whom were studied before and after pharmacological neuronal uptake-blockade with desipramine. Total, cardiac and renal noradrenaline spillover to plasma were 78%, 98% and 66% lower respectively in pure autonomic failure than in normal subjects (p less than 0.001). Total noradrenaline plasma clearance was 20% lower in pure autonomic failure (p less than 0.005) than in normal subjects and similar to the level observed in normal subjects following neuronal noradrenaline uptake-blockade with desipramine, mean transcardiac extraction of tritiated noradrenaline was 74% in normal subjects and 20% in pure autonomic failure, identical to the value post-desipramine in normal subjects. Cardiac spillover of the noradrenaline precursor, dihydroxyphenylanine, and the primary intra-neuronal metabolite dihydroxyphenylglycol, were 78% and 94% lower respectively in pure autonomic failure than in normal subjects (p less than 0.001). These data indicate a marked reduction in the apparent release rate and neuronal uptake of noradrenaline in the hearts of patients with pure autonomic failure, and provide biochemical evidence of almost total postganglionic sympathetic denervation in this condition.

Neuronal reuptake of norepinephrine and production of dihydroxyphenylglycol by cardiac sympathetic nerves in the anesthetized dog.

Reuptake of norepinephrine by cardiac sympathetic nerves before and during two levels of electrical stimulation of the left ansa subclavia was estimated in anesthetized dogs from the cardiac production of dihydroxyphenylglycol (DHPG), the intraneuronal metabolite of norepinephrine. The method depended on the effects of neuronal uptake blockade with desipramine on the cardiac production of [3H]DHPG from intravenously infused [3H]norepinephrine. The ratio of the desipramine-induced decrease in the cardiac extraction of [3H]norepinephrine to the production of [3H]DHPG was used to transform the cardiac production of DHPG from recaptured norepinephrine into a rate for norepinephrine reuptake. Cardiac spillover of norepinephrine into plasma increased from 49 +/- 12 to 205 +/- 40 and 451 +/- 118 pmol/min during sympathetic activation. Cardiac DHPG production increased from 108 +/- 18 to 166 +/- 34 and 240 +/- 47 pmol/min. Desipramine decreased resting cardiac DHPG production by 20% and completely blocked the stimulation-induced increase. Thus, most (80%) cardiac DHPG produced at rest was derived from norepinephrine leaking from storage vesicles. This amount remained constant, and that derived from recaptured norepinephrine increased during sympathetic activation. The cardiac extraction of [3H]norepinephrine (126,000 dpm/min) and production of [3H]DHPG (3,790 dpm/min) were decreased by 55-57% after desipramine. Thus, only 3% of the norepinephrine recaptured by cardiac sympathetic nerves appeared in plasma as DHPG. The remainder was sequestered into storage vesicles (more than 94%) or ultimately formed metabolites other than DHPG (less than 3%). Reuptake of norepinephrine by cardiac sympathetic nerves was 1,188 +/- 476 pmol/min and increased in parallel with cardiac norepinephrine spillover to 4,182 +/- 1,982 and 6,594 +/- 2,241 pmol/min during sympathetic stimulation. Of the norepinephrine released by cardiac sympathetic nerves, 16-fold more was recaptured than entered plasma. Combined estimation of norepinephrine reuptake and spillover offers an approach to assess the efficiency of neuronal reuptake in disorders of cardiac function.

Regional extraction of circulating norepinephrine, DOPA, and dihydroxyphenylglycol in humans

Dihydroxyphenylglycol (DHPG) is the main intraneuronal metabolite of the sympathetic neurotransmitter, norepinephrine (NE), and dihydroxyphenylalanine (DOPA) the immediate product of the rate-limiting step in catecholamine biosynthesis. Simultaneous measurements of regional rates of appearance (spillovers) of NE, DOPA, and DHPG in plasma have the potential to provide unique information about aspects of sympathoneural function but have not actually been measured in humans. In the present study, spillovers of DHPG, DOPA, and NE in the heart, head, leg, and lungs, were estimated from regional extraction fractions of infused [ 3H]-1-NE, DHPG, and [ 13C 6]DOPA or unlabelled DOPA in humans during cardiac catheterization. There was little cardiac extraction of DHPG (7 ± SEM 2%) or DOPA (8 ± 4%) but substantial extraction of NE (69 ± 4%). Values for cardiac spillover of DHPG and DOPA therefore were similar to values for the arteriovenous increment times plasma flow (arteriovenous production rate), whereas the cardiac spillover of NE averaged about 7-times the NE arteriovenous production rate. Cardiac DHPG spillover (28 ± 3 ng/min) exceeded the spillovers of NE (9 ± 2 ng/min) and DOPA (15 ± 4 ng/min). In contrast, cranial DOPA spillover (159 ng/min) exceeded those of NE and DHPG by 8- and 2-fold and accounted for about 1/10 of the total spillover of DOPA into arterial plasma. In the femoral vascular bed, arteriovenous production rates of NE and DHPG were unrelated to femoral spillovers of NE and DHPG. Arterial and regional clearances of [ 13 C 6]DOPA were similar to those of unlabelled DOPA. The results suggest that (1) endogenous NE, DOPA, and DHPG all are released into the bloodstream by the heart, head, and limbs of humans; (2) DHPG and DOPA are not co-released with NE; (3) cardiac arteriovenous production rates of DOPA and DHPG can be used to indicate cardiac spillover of these catechols, whereas the cardiac NE arteriovenous production rate substantially underestimates cardiac NE spillover; and (4) estimates of limb spillover of NE and DHPG require concurrent measurements of the corresponding regional clearances.