Unlabelled Noradrenaline Research Articles

Effects of iodoproxyfan, a potent and selective histamine H3 receptor antagonist, onα 2 and 5-HT3 receptors

We determined the affinity and/or potency of the novel H3 receptor antagonist iodoproxyfan at alpha 2 and 5-HT3 receptors. Iodoproxyfan and rauwolscine (a reference alpha 2 ligand) (i) monophasically displaced 3H-rauwolscine binding to rat brain cortex membranes (pKi 6.79 and 8.59); (ii) facilitated the electrically evoked tritium overflow from superfused mouse brain cortex slices preincubated with 3H-noradrenaline (pEC50 6.46 and 7.91) and (iii) produced rightward shifts of the concentration-response curve (CRC) of (unlabelled) noradrenaline for its inhibitory effect on the evoked overflow (pA2 6.65 and 7.88). In the guinea-pig ileum, iodoproxyfan 6.3 mumol/l failed to evoke a contraction by itself but depressed the maximum of the CRC of 5-hydroxytryptamine (pD'2 5.24). Tropisetron (a reference 5-HT3 antagonist) produced rightward shifts of the CRC of 5-hydroxytryptamine (pA2 7.84). In conclusion, the affinity/potency of iodoproxyfan at H3 receptors (range 8.3-9.7 [1]) exceeds that at alpha 2 receptors by at least 1.5 log units and that at 5-HT3 receptors by at least 3 log units.

The 5-HT3 receptor agonist 1-(m-chlorophenyl)-biguanide facilitates noradrenaline release by blockade of ?2-adrenoceptors in the mouse brain cortex

We analyzed the facilitatory effect of the 5-HT3 receptor agonist 1-(m-chlorophenyl)-biguanide (mCPBG) on the electrically evoked noradrenaline release in superfused mouse brain tissue. In addition, we determined the affinities of mCPBG and two other 5-HT receptor ligands, namely 2-methyl-5-hydroxytryptamine (2-methyl-5-HT; also a 5-HT3 receptor agonist) and 5-carboxamidotryptamine (5-CT; a 5-HT1 receptor agonist) for alpha 2 binding sites. The latter two 5-HT receptor agonists were included because of the claimed involvement of alpha 2-adrenoceptors in their effects on noradrenaline release. In superfusion experiments on mouse brain cortex slices preincubated with 3H-noradrenaline, tritium overflow evoked by 2-min periods of electrical field stimulation (3 Hz) was facilitated by mCPBG and, in addition, by rauwolscine (alpha 2-adrenoceptor antagonist) and tetraethylammonium (K+ channel blocker) (which were examined for comparison). The effect of mCPBG was not affected by the 5-HT3 receptor antagonist tropisetron or by desipramine but was abolished by rauwolscine. In slices superfused with medium containing desipramine, the concentration-response curve of unlabelled noradrenaline for its inhibitory effect on the electrically (0.3 Hz) evoked overflow was shifted to the right by mCPBG and rauwolscine (apparent pA2 5.35 and 7.88, respectively). In another series of superfusion experiments, 4 electrical pulses, administered at 100 Hz, were used to evoke tritium overflow. Tritium overflow evoked by this stimulation procedure (under which an endogenous tone of noradrenaline does not develop) was not affected by mCPBG and rauwolscine but still increased by tetraethylammonium.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of blockade of noradrenaline re‐uptake on evoked tritium overflow from mouse vasa deferentia and rat cortex slices

1. In tissues previously incubated with [3H]-noradrenaline exposure to cocaine (0.1 to 10 microM) or desmethylimipramine (0.01 to 1 microM) produced a concentration-dependent increase (up to 2 fold) in electrically evoked (3 Hz, 2 ms, 20 mA, 120s every 20 min) fractional overflow of tritium from rat brain cortex slices but not from mouse vas deferens (2.5 Hz, 2 ms, 400 mA, for 90s every 14 min). 2. Yohimbine and idazoxan (0.01 to 1 microM) increased fractional evoked overflow of tritium by up to 2 fold; in the presence of these drugs, cocaine (10 microM) produced an increase in both tissues (up to 3.5 fold over control). 3. In brain slice an increase in stimulation frequency (0.1, 0.5, 1, 3 and 6 Hz) decreased fractional evoked overflow of tritium per pulse but cocaine (10 microM) produced a significant enhancement at each frequency except 6 Hz. In vas deferens fractional tritium overflow per pulse changed little with increasing frequency and cocaine produced no effect. 4. In both tissues fractional evoked overflow of tritium was dependent on the stimulation current; cocaine (10 microM) increased fractional evoked overflow from brain slice at every current tested but was without effect in vas deferens. 5. Chromatographic separation of the released tritium showed there was little difference in the proportions of [3H]-noradrenaline and 3H-metabolites overflowing from the tissues. Cocaine increased the proportion of [3H]-noradrenaline and decreased the proportion of [3H]-DOPEG overflowing both at rest and during stimulation. 6. In brain slice an increase in electrically evoked overflow was produced by cocaine (10 microM) whether total tritium overflow (1.8 fold), overflow of [3H]-noradrenaline (1.8 fold) or overflow of unlabelled noradrenaline (1.8 fold) was measured. Evoked overflow from vas deferens was unaffected when assessed by any of these three methods. 7. The mechanism responsible for this differential effect of cocaine is unclear but may involve differences in the physical relationship between release sites, reuptake sites and presynaptic autoreceptors.

Total body, systemic and pulmonary clearance and fractional extraction of unlabelled and differently 3H-labelled noradrenaline in the anaesthetized rabbit

1. Rabbits were anaesthetized with urethane/chloralose and infused intravenously with trace amounts of 3H-2,5,6-, 3H-7,8- or 3H-7-(-)noradrenaline either without or with unlabelled (-)noradrenaline being simultaneously infused (0.2 micrograms kg-1 min-1). To obtain clearance values and extraction ratios for the pulmonary, systemic and total circulation, steady-state concentrations of infused noradrenaline were determined in mixed central venous (Cv) and arterial (Ca) plasma. Heart rate and blood pressure were recorded via the carotid artery, and the dye dilution method was used to determine the cardiac output of plasma. 2. The simultaneous infusion of unlabelled noradrenaline, which increased plasma levels of noradrenaline by a factor of 5, had no significant effect on either heart rate, blood pressure or cardiac output (when determined at steady state of the noradrenaline infusion). 3. The simultaneous infusion of unlabelled noradrenaline did not affect the clearance values of any of the three type of 3H-noradrenaline. Moreover, the clearances of the various types of 3H-noradrenaline were virtually identical and agreed with that of unlabelled noradrenaline. However, the clearance of labelled and unlabelled noradrenaline from arterial plasma was 1.15 times higher than that from central venous plasma. This factor corresponded to the ratio of Cv/Ca and pointed towards net removal of noradrenaline from the pulmonary circulation. 4. The fractional pulmonary extractions [1-(Ca/Cv)] of the three types of 3H-noradrenaline did not differ from each other and were not affected by the simultaneous infusion of unlabelled noradrenaline.(ABSTRACT TRUNCATED AT 250 WORDS)

Lack of selectivity between the uptake of [3H]adrenaline and [3H]noradrenaline into rat hypothalamic slices.

The uptake of [3H]adrenaline and [3H]noradrenaline into rat hypothalamic slices was compared for determination of whether adrenaline uptake was independent of uptake into noradrenergic neurones. Kinetic analysis revealed a similar high-affinity uptake process for both adrenaline and noradrenaline, with Km and Vmax values within similar ranges. These uptakes were inhibited by desipramine and maprotiline in a dose-dependent manner, but the selective dopamine and 5-hydroxytryptamine uptake inhibitors benztropine and fluoxetine, respectively, were without effect. Competition for uptake sites by unlabelled adrenaline with [3H]adrenaline and [3H]-noradrenaline and by unlabelled noradrenaline with [3H]-adrenaline and [3H]noradrenaline was very similar. Lesioning of the major adrenaline-containing cell group (C1 cell group) decreased the hypothalamic adrenaline concentration but had no effect on hypothalamic [3H]adrenaline or [3H]noradrenaline uptake. The results suggest that exogenous adrenaline is largely taken up by high-affinity sites on noradrenergic nerve terminals.

Modulation of noradrenaline release in human saphenous vein via presynaptic alpha 2-adrenoceptors.

Strips of human saphenous veins were incubated with [3H]noradrenaline and subsequently superfused with physiological salt solution containing cocaine, corticosterone and propranolol. The electrically (6 Hz) evoked overflow of tritium (78% of which was accounted for by unmetabolized [3H]noradrenaline) was abolished by tetrodotoxin or omission of Ca2+ from the superfusion fluid. Unlabelled noradrenaline, alpha-methylnoradrenaline, B-HT 920 and clonidine inhibited the evoked overflow (maximum effect of clonidine lower than that of the other compounds) whereas methoxamine was ineffective. The alpha 2-adrenoceptor antagonists, BDF 6143 and rauwolscine, facilitated the evoked overflow but no effect was obtained with prazosin. Rauwolscine produced a shift to the right of the concentration-response curve of B-HT 920 for its inhibitory effect on evoked outflow and BDF 6143 caused a shift to the right of the corresponding curve of clonidine. It is concluded that the stimulation-evoked release of noradrenaline from the sympathetic nerve fibres of the human saphenous vein is modulated via presynaptic alpha 2-adrenoceptors.

Radioenzymatic assay of catecholamines: real and apparent losses of labelled product licister mason and cyril weinkove

In the radioenzymatic assay of catecholamines, using catecho l- O-methyltransferase, the yield of labelled product is frequently less than the expected value. This has been attributed by some workers to losses during the periodate oxidation of the O-methylated derivatives. Using spectrophotometric methods, we have demonstrated that there is no oxidative demethylation of the methoxycatecholamines during periodate oxidation. In a novel technique designed to determine the specific activity of small quantities of tritiated S-adenosylmethionine, high performance liquid chromatography with electrochemical detection was used to measure the amount of adrenaline formed from unlabelled noradrenaline, in the presence of phenylethanolamine- N-methyltransferase and tritiated S-adenosylmethionine. As well as the real losses occurring during solvent extraction and thin layer chromatography (demonstrated spectrophotometrically), apparent ‘loss’ of radiolabelled product may be due to the assumption that the value for the specific activity of the tritiated S-adenosylmethionine and the determination of the product's radioactivity are both absolutely accurate, with no allowance being made for the normal and expected experimental errors in such measurements.

The influence of uptake2 on the inhibition by unlabelled noradrenaline of the stimulation-evoked overflow of 3H-noradrenaline in rabbit aorta with regard to surface of amine entry.

The release by electrical field stimulation of 3H-noradrenaline from the adrenergic nerve endings in the rabbit aorta was studied in a special double-chambered organ bath. Independently of the frequency (1-10 Hz) and the number of pulses used (300-3,000 pulses), only 10-20% of the stimulation-evoked overflow of tritium (total evoked overflow) left the aortic wall via the intimal surface (intimal evoked overflow). The corresponding percentage value for the basal efflux was twice that for the evoked overflow, thus indicating that part of the radioactivity in the basal efflux originated from an extraneuronal compartment. The radioactivity in the total evoked overflow (in response to stimulation at 10 Hz) originated from at least two compartments (compartment I and II) with half times for efflux of about 2 and 6 min, respectively. In the intimal evoked overflow, compartment II (but not compartment I) was involved. When uptake1 was inhibited, 70% of the radioactivity in the intimal evoked overflow (stimulation at 1 Hz) consisted of metabolites, while unchanged amine prevailed by far in the total and adventitial evoked overflow, respectively. Additional inhibition of uptake2 thus had a striking effect only on the composition of the radioactivity in the intimal evoked overflow. The intimal surface was exposed to unlabelled noradrenaline in order to inhibit the evoked overflow of tritium (stimulation at 1 Hz; uptake1 inhibited). When uptake2 was inhibited additionally, the dose-response curve for the inhibitory effect of noradrenaline was shifted to the left by a factor of 4.(ABSTRACT TRUNCATED AT 250 WORDS)

Noradrenaline release and clearance in relation to age and blood pressure in man.

Plasma noradrenaline concentration increases with age. This study was designed to investigate whether an increased rate of noradrenaline release into the circulation or a decrease in clearance is primarily responsible for this age related change in concentration. Sixteen healthy male subjects were studied, eight young (21-36 years) and eight old (65-78 years). Clearance was calculated from steady state noradrenaline concentrations during constant rate infusions of unlabelled noradrenaline. Clearance did not differ between the two groups: young 4.8 l/min (range 2.7-6.1), old 4.1 (range 2.6-8.2). The old subjects had significantly greater rates of release. Supine: young 10.3 nmol/min (range 5.3-17.6), old 19.7 (range 10.1-30), P less than 0.05. Standing: young 17.2 (range 11-36.4), old 29.2 (range 21.8-47.9), P less than 0.01. No significant relationship was found in either supine or standing position between rate of noradrenaline release and either systolic or diastolic blood pressure. These results indicate that plasma noradrenaline concentration rises with age because of an increased rate of release, but that this increased release is not responsible for the higher blood pressure seen in the elderly.

Assessment of sympathetic nervous function in humans from noradrenaline plasma kinetics.

Noradrenaline is the neurotransmitter of the sympathetic nervous system. Many attempts have been made, in research spanning two decades 11-41, to utilize tracer methodology and the mathematical techniques of compartmental analysis 15, 61 to elucidate the kinetics of noradrenaline in humans. These studies have often been hampered by technical problems, such as the impracticability of assaying tissue noradrenaline levels in humans, and in earlier experiments, by the unavailability of radiolabelled L-noradrenaline, and the lack of a sufficiently sensitive and specific assay for plasma noradrenaline. The disposition of noradrenaline in fact is complex, defying precise compartmental analysis 141, and in the past this has frustrated attempts to measure noradrenaline turnover. Noradrenaline in plasma is derived in the main from transmitter released by sympathetic nerves, to a very small degree from the adrenal medulla 171, and probably not at all from the central nervous system [81. Faced with this lack of an acceptable method for determining noradrenaline turnover, investigators have turned to the plasma concentration of the transmitter as an index of overall sympathetic nervous system ‘activity’ (nerve firing rate) in studies of human physiology and disease [9-141. There are several possible objections, however, to this application of plasma noradrenaline values, not the least being that the plasma concentration is determined in part by the rate at which noradrenaline is removed from the circulation, and not solely by the rate of noradrenaline release. provide such an ambiguous guide to sympathetic nervous system activity, kinetic techniques for estimating the rate of release of noradrenaline to plasma have recently been developed 115-191. These methods for studying plasma noradrenaline kinetics differ from the earlier, more ambitious studies of whole-body noradrenaline kinetics in several respects, in particular in their independence from complex mathematical models of noradrenaline disposition. The central feature of these methods is the determination of the metabolic clearance rate of plasma noradrenaline, clearance being calculated after either a bolus intravenous injection of noradrenaline I171 or intravenous infusion to steady state 115, 16, 18, 191. Published methods differ in their use of either radiolabelled 115, 17, 181 or unlabelled noradrenaline 116, 191, and if a tracer is used, in the use of either Lor DL-noradrenaline. The use of either pharmacological doses of unlabelled noradrenaline [ 16, 191 or radiolabelled DL-IIOTadrenaline [ 151 carries potential disadvantages. If pressor doses of noradrenaline are used, clearance estimates may possibly be distorted by saturation of removal mechanisms or alteration of organ blood flow rates, and reflex suppression of endogenous noradrenaline release could occur [ 16,201. Differences in the body’s handling of the D-isomer, particularly for noradrenaline uptake [ 2 11, could influence results obtained with racemic radiolabelled noradrenaline. For infused radiolabelled noradrenaline (NA) the following relationships hold, under steadystate conditions 15, 181:

Alpha-Adrenoceptor-mediated inhibition of noradrenaline release in rabbit brain cortex slices. Receptor properties and role of the biophase concentration of noradrenaline.

Brain cortex slices from rabbits were preincubated with [3H]noradrenaline and then superfused and stimulated electrically at 3Hz. In the presence of cocaine 30 microM, unlabelled noradrenaline, alpha-methylnoradrenaline, clonidine, oxymetazoline, xylazine and guanabenz decreased, whereas yohimbine, corynanthine, phentolamine, tolazoline and azapetine increased the stimulation-evoked overflow of tritium. Phenylephrine and prazosin had no effect on the evoked overflow except at concentrations that greatly accelerated the basal outflow of tritium. The results indicate that the noradrenergic axons of rabbit brain cortex are endowed with presynaptic alpha-adrenoceptors which are exclusively of the alpha 2-type. Addition of various concentrations of cocaine, addition of pargyline, or stimulation at different current strengths was used to obtain either a high or low stimulation-evoked overflow of tritium. Independently of the method used, a low evoked overflow coincided with a large percentage inhibition produced by 0.1 micro M clonidine, whereas a high evoked overflow coincided with a smaller percentage inhibition produced by clonidine. The results indicate that drugs which block the re-uptake of noradrenaline diminish the presynaptic inhibitory effect of alpha-adrenergic agonists by increasing the biophase concentration of released noradrenaline.

Antagonistic properties of quipazine at presynaptic serotonin receptors and alpha-adrenoceptors in rat brain cortex slices.

Rat brain cortex slices preincubated with 3H-serotonin or 3H-noradrenaline were superfused with physiological salt solution, and the effect of quipazine on the 3H overflow evoked by electrical field stimulation was examined. 1. The electrically evoked tritium overflow from brain slices preloaded with 3H-serotonin was increased by quipazine in a concentration-dependent manner. The concentration-response curves of unlabelled serotonin and noradrenaline for their inhibitory effects on impulse-evoked 3H overflow were shifted to the right by quipazine. 2. In brain slices preincubated with tritiated noradrenaline, quipazine caused an increase in electrically stimulated 3H overflow which was less marked than the effect on brain slices preloaded with 3H-serotonin. The concentration-response curve of unlabelled noradrenaline for its decreasing effect on stimulation-evoked 3H overflow was shifted to the right by quipazine. We conclude that quipazine blocks presynaptic inhibitory serotonin receptors and alpha-adrenoceptors on monoaminergic neurones of the rat brain cortex.

Functional characterization of central α-adrenoceptors by yohimbine diastereomers

Rat occipital cortex slices were preincubated with [ 3H]noradrenaline and then superfused with medium containing 30 μM cocaine. They were stimulated electrically at 3 Hz. Unlabelled noradrenaline (0.1 μM), α-methylnoradrenaline (0.01–0.1 μM), xylazine (0.1–10 μM) and guanabenz (0.01 μM) decreased, whereas yohimbine (0.01–1 μM), rauwolscine (0.01–1 μM), corynanthine (10 μM), tolazoline (0.1–10 μM) and azapetine (0.1–1 μM) increased the stimulation-evoked overflow of tritium without a change in basal outflow. Pseudoyohimbine and prazosin at up to 0.1 μM did not change the evoked overflow, and at higher concentrations enhanced the basal outflow of tritium. In vivo, yohimbine 10 mg/kg and rauwolschine 10 mg/kg markedly, and corynanthine 10 mg/kg slightly accelerated the α-methyltyrosine-induced disappearance of noradrenaline from rat whole brain. Yohimbine and rauwolscine but not corynanthine also accelerated the α-methyltyrosine-induced depletion of dopamine. The results add four compounds to the list of drugs with α-adrenoceptor affinity which inhibit (agonists) or facilitate (antagonists) action-potential-evoked release of noradrenaline in rat brain cortex. The presynaptic receptors are of the α 2-type. The receptors which control the activity of noradrenaline and dopamine neurons in vivo also appear to be α 2.

Adrenaline activation of prejunctional beta-adrenoceptors in guinea-pig atria.

1. Adrenaline in a concentration of 1.0 microM depressed the stimulation-induced efflux of tritium from the guinea-pig atria incubated with [3H]-noradrenaline, whereas adrenaline in a concentration of 0.5 nM significantly enhanced the stimulation-induced efflux of tritium. This enhancement was blocked by metoprolol (0.1 microM) and thus appears to be mediated by beta-adrenoceptors. 2. In guinea-pig atria incubated with unlabelled adrenaline and then with [3H]-noradrenaline, both catecholamines were released by field stimulation. In such atria metoprolol, practolol, oxprenolol or propranolol decreased the stimulation-induced efflux of tritium. These effects did not occur if the atria were incubated with unlabelled noradrenaline and then with [3H]-noradrenaline, suggesting that neuronally released adrenaline activates prejunctional beta-adrenoceptors. 3. The effect of oxprenolol in decreasing the release of tritium from guinea-pig atria, incubated with unlabelled adrenaline and then with [3H]-noradrenaline was greater in the presence of phentolamine. This may reflect the alpha-adrenoceptor blocking activity of oxprenolol.

The neuronal and extraneuronal distribution of 3H(-)-noradrenaline in the perfused rat heart.

1. After inhibition of both COMT and MAO, rat hearts were perfused with 1 μM 3H-(−)-noradrenaline for 30 min. Cocaine-sensitive neuronal and corticosterone-sensitive extraneuronal compartments were of about equal importance for the distribution of the amine into the heart. The absence of calcium from the perfusion fluid increased the axoplasmic and decreased the extraneuronal distribution of noradrenaline. 2. Under the same experimental conditions, hearts were washed out after the initial perfusion, and the efflux curves were subjected to compartmental analysis. For the nerve endings three distribution compartments were identified (the axoplasmic compartments B and C, characterized by half times of 8 and 45 min, respectively, as well as the vesicular “bound fraction”), while there was only one important extraneuronal distribution compartment (compartment C with a half time of about 45 min). 3. Efflux from the axoplasmic compartment C was accelerated by unlabelled noradrenaline, not affected by cocaine, reduced by lowering of the temperature from 37° to 27° C. Moreover, it was greatly accelerated by the omission of sodium from the wash-out solution. 4. Efflux from the extraneuronal compartment C, on the other hand, was not affected by the presence of unlabelled noradrenaline, and impaired by lowering of the temperature as well as by corticosterone. The omission of sodium from the wash-out solution caused an acceleration of the extraneuronal efflux which was significantly smaller than that of axoplasmic efflux. 5. Other hearts were perfused with either 1 or 16.7 μM 3H-(−)-noradrenaline (plus 30 μM cocaine) for 30 min and then washed out with amine-free solution; in these experiments either MAO or COMT was inhibited. When MAO was inhibited, the efflux curves for normetanephrine indicated that the accumulation of the amine in the extraneuronal compartment C can easily exceed that concentration which saturates the intracellular COMT. When MAO was not inhibited, afflux curves for DOPEG failed to provide any evidence for saturation of extraneuronal MAO. These results are in good agreement with earlier determinations of the kinetic constants of the “extraneuronal metabolizing systems” of the rat heart (Fiebig and Trendelenburg, 1978b). 6. The results indicate that extraneuronal mechanisms play an important role in the rat heart, and also that the extraneuronal compartment C (for noradrenaline) corresponds to the compartment III (for isoprenaline described by Bonisch et al. (1974).

Effects of presynaptic modulators on Ca2+-induced noradrenaline release from central noradrenergic neurons. Noradrenaline and enkephalin inhibit release by decreasing depolarization-induced Ca2+ influx.

Slices of rat occipital cortex and hypothalamus were preincubated with 3H-noradrenaline. Tritium overflow was induced by introduction of 0.65 or 1.3 mM CaCl2 for 2–5 min after superfusion of the slices with Ca2+-free solution; Ca2+-induced overflow was promoted either by the presence of 30 mM K+ throughout superfusion or by incubation with the ionophore A 23187 (300 μM; subsequent to preincubation with 3H-noradrenaline) before onset of superfusion. The effects of drugs interfering with presynaptic α-adrenoceptors and opiate receptors on CaCl2-induced tritium overflow and on the metabolism of 3H-noradrenaline released were investigated. 1. The CaCl2-evoked tritium overflow promoted by 30 mM K+ was decreased by methionineenkephalin (1 μM). This effect was antagonized by simultaneous administration of naloxone (10 μM). When given alone, naloxone (10 μM) did not alter the CaCl2-induced overflow of tritium. 2. When cocaine (100 μM) was present in the superfusion fluid throughout the experiments, the CaCl2-evoked tritium overflow promoted by 30 mM K+ was decreased by unlabelled noradrenaline (1 μM) and increased by phentolamine (1 μM); the inhibition of CaCl2-induced tritium overflow caused by unlabelled noradrenaline was reversed by simultaneous administration of phentolamine (1 μM each). 3. The CaCl2-evoked tritium overflow promoted by ionophore A 23187 was not affected by 1 μM methionine-enkephalin or 1 μM unlabelled noradrenaline (the latter in the presence of 100 μM cocaine throughout superfusion). 4. Methionine-enkephalin, unlabelled noradrenaline and phentolamine did not significantly alter the metabolism of 3H-noradrenaline released by 1.3 mM CaCl2 under both conditions (30 mM K+ and 300 μM A 23187, respectively).

Chronic methods to study the release of catecholamines from the neostriatum in nonanaesthetized monkeys (Macaca mulatta).

The present report describes two methods allowing the chronic collection of catecholamine (CA) released from the neostriatum in the nonanaesthetized monkey (Macaca mulatta). In both methods the monkeys were placed in a restraining chair. Indwelling electrodes allowed the correlation of behavioural observations with polygraphic recordings. The first method uses a special superfusing cup permanently implanted on the lateral surface of the caudate nucleus, which permits the demonstration of spontaneous and D-amphetamine-induced release of (3H)-dopamine (DA) newly synthesized from L-(3,5-3H)-tyrosine over a period of six days. In the second method the collection of unlabelled CA is carried out by a localized ventricular perfusion. A radio-enzymatic estimation enabled the measurement of unlabelled DA and noradrenaline. With this second method the animals were kept for 25 to 30 days.

The Kinetics of Plasma Noradrenaline in Normal and Hypertensive Subjects

1. The kinetics of plasma noradrenaline have been determined in normal and essential hypertensive patients by intravenous injection of tritiated noradrenaline and serial mixed venous sampling. 2. The metabolic clearance rate of plasma noradrenaline in normal subjects was approximately 1 1 min-1 m-2, whereas in essential hypertensive patients it was significantly reduced to approximately 0.6 1 min-1 m-2. 3. Metabolic clearance rate was negatively correlated to mean arterial blood pressure and total peripheral resistances. 4. Particularly low values of metabolic clearance rate were found in two patients with congestive heart failure and one with phaeochromocytoma. 5. We propose that the access of plasma noradrenaline to the main removal mechanisms takes place in competition with the flow of unlabelled endogenous noradrenaline directly released by nerve endings. The slower removal of plasma noradrenaline in essential hypertension could then express a larger release of endogenous noradrenaline in this condition.

Presynaptic receptor systems on the noradrenergic neurones of rat brain.

Occurrence and properties of receptors modulating release of noradrenaline were studied in slices of rat occipital cortex and, less extensively, of rat hypothalamus and cerebellar cortex. The slices were preincubated with 3H-noradrenaline and then superfused and stimulated either electrically at 1–3 Hz or by 15–20 mM potassium. (1) Omission of calcium abolished the electrically and potassium-evoked overflow of tritium. Tetrodotoxin blocked the electrically evoked overflow and reduced the response to potassium. The electrically evoked overflow of total tritium consisted of 80% 3H-noradrenaline and 19% 3H-3,4-dihydroxyphenylglycol (DOPEG). Cocaine strongly decreased the electrically evoked overflow of 3H-DOPEG. The overflow of tritium evoked by 15 mM potassium also contained mainly 3H-noradrenaline with only a minor part of 3H-DOPEG. (2) Both the electrically and the potassium-evoked overflow of tritium were reduced by unlabelled noradrenaline (in the presence of cocaine) and tramazoline, and increased by phentolamine and yohimbine. The changes in the evoked overflow of total tritium reflected approximately proportionate changes of 3H-noradrenaline and 3H-DOPEG. (3) Both the electrically and the potassium-evoked overflow of tritium were reduced by morphine and methionine-enkephalin. 3H-noradrenaline and 3H-DOPEG were proportionately decreased. Dextrorphan, in contrast to its analgesically active enantiomer levorphanol, had no effect. Inhibition by morphine persisted after pretreatment with indometacin. (4) Prostaglandin E1 reduced both the electrically and the potassium-evoked overflow of tritium. 3H-noradrenaline and 3H-DOPEG were proportionately decreased. (5) Phentolamine antagonized the inhibitory effect of noradrenaline, but not that of morphine, enkephalin, and prostaglandin E1. Naloxone antagonized the effect of enkephalin, but not that of tramazoline and prostaglandin E1. (6) Acetylcholine (up to 10−5 M) and oxotremorine (up to 10−4 M) failed to reduce the electrically and potassium-evoked overflow. Moreover, atropine (up to 3×10−6 M) had no effect. Up to 10−3 M acetylcholine (in the presence of atropine) as well as up to 10−4 M nicotine did not change the basal outflow of tritium in the absence of releasing stimuli. 10−3 M nicotine caused an increase which was not changed by hexamethonium or omission of calcium. In contrast to the overflow evoked by electrical stimulation or high potassium, the overflow evoked by nicotine mainly consisted of 3H-DOPEG with only a minor part attributable to 3H-noradrenaline. (7) γ-Aminobutyric acid slightly enhanced both the basal and (maximally by 27%) the electrically evoked overflow. Histamine (in the presence of cocaine) slightly decreased the electrically evoked overflow (maximally by 17%). (8) The following drugs did not change the electrically evoked overflow of tritium: dopamine (in the presence of cocaine), isoprenaline (up to 10−6 M), propranolol (up to 10−6 M), serotonin (in the presence of cocaine), angiotensin I, angiotensin II, saralasin, and substance P. (9) It is concluded that transmitter release from the noradrenergic neurones of several brain areas is modulated by drugs acting on α-adrenoceptors, opiate receptors, and prostaglandin receptors. It seems likely, though by no means certain, that the receptors involved are located on the noradrenergic nerve endings themselves, i.e., that they are presynaptic receptors. No evidence was obtained for presynaptic dopamine, β-, muscarine, nicotine, and angiotensin receptors. This pattern differs from that found in other tissues, thus substantiating marked tissue differences in presynaptic receptor systems.

Selectivity for catecholamines of presynaptic alpha-receptors involved in feedback control of sympathetic neurotransmitter secretion in guinea-pig vas deferens

The aim of the study was to quantitatively compare the relative affinities of noradrenaline, adrenaline, dopamine and isoprenaline for the, probably neural, receptors mediating feedback control of sympathetic neurotransmitter secretion. The experiments were carried out in isolated superfused field stimulated guinea-pig vas deferens, in which the noradrenaline stores had been labelled by preincubation with tritiated (-)-noradrenaline. Desipramine and normetanephrine were added to prevent rebinding of transmitter. Exogenous noradrenaline was found to cause a dose-dependent and reversible depression of the secretion of tracer noradrenaline evoked by field stimulation. Since the depressing effect was not affected by a ten-fold rise in the desipramine concentration, it seems likely that it was not due to uptake and preferential secretion of unlabelled exogenous noradrenaline, but was truly due to depression of the secretory mechanism. Adrenaline was significantly more potent than noradrenaline, as inhibitor of the secretion of tracer transmitter, while dopamine, at the same molar concentration, was without effect. The beta-agonist isoprenaline did not depress, but rather tended to enhance, the secretion of tracer noradrenaline. It is concluded that the receptors controlling the secretion of noradrenaline from the sympathetic nerves of guinea-pig vas deferens quantitatively-with regard to sensitivity-as well as qualitatively-with regard to order of preference for different catecholamines-resemble the "classical" alpha-receptors of e.g. smooth muscle in the same tissue.