Coronary arteries of cardiac patients are hyperreactive and contain stores of amines: a mechanism for coronary spasm.

Coronary arteries from hearts of cardiac patients contain significantly higher concentrations of histamine than do those from noncardiac patients. The coronary vessels of cardiac patients are also hyperresponsive to histamine and serotonin. These differences between groups of patients suggest an explanation for coronary artery spasm in heart disease.

Non-neurogenic contractions in isolated coronary arteries by brief electrical pulses.

OBJECTIVE: The aim was to describe a novel form of non-neurogenic coronary artery contraction. METHODS: Superfused cattle coronary artery preparations in vitro were placed between platinum electrodes and stimulated. RESULTS: The preparations responded to exceedingly brief transmural stimulation (a single ten thousandth of a second pulse) with long lasting [150(SEM 18.5) s] contractions. These previously undescribed contractions were not reduced by neural blockade nor did they involve free radicals, prostaglandins, or the endothelium. In contrast to the rapid loss of responses to KCl in zero calcium Krebs solution, responses to stimulation were only progressively diminished over many minutes, suggesting involvement of internal calcium stores. The L channel calcium antagonists nifedipine and diltiazem markedly reduced contractions to KCl but did not materially alter those to 1, 5, or 10 stimulation pulses, nor did the T channel antagonist tetramethrin, further supporting the involvement of stored calcium in contractions to stimulation. Evidence was obtained that neither Na+/K(+)-ATPase nor the Na+/Ca+ exchanger are involved in the contractions to stimulation. Ryanodine in zero calcium Krebs solution potentiated contractions to stimulation with 10 pulses and also to endothelin, but depressed contractions to U 46619. CONCLUSIONS: Stimulation at excitation parameters well below those associated with neurotransmitter release activates a highly effective contraction sequence that may use a ryanodine insensitive pool of bound calcium. This novel process may have physiological and pathophysiological relevance and provides a means of activating contraction in a coronary artery and studying its time course and contributory components, without the complicating participation of an agonist drug.

Single pulse stimulation of guinea-pig vas deferens and the presynaptic receptor hypothesis.

1 The effect of phenoxybenzamine on the efflux of [3H]-noradrenaline and the mechanical response to single pulse excitation of superfused guinea-pig vas deferens was determined to examine the validity of the currently accepted hypothesis of a presynaptic negative feedback system on adrenergic nerve terminals. 2 The adrenoceptor antagonist enhanced both the outflow of tritium and the mechanical response to single pulse stimulation. The efflux of labelled material and the responses to 4 pulses were also enhanced, as expected. 3 Blockade of neuronal and extraneuronal uptake did not by itself increase nerve-induced outflow or the mechanical response nor dated within the framework of a hypothesis that proposes that the enhancement of response and tritium efflux by phenoxybenzamine results from blockade of a feedback system whereby noradrenaline released by previous impulses inhibits its own subsequent release.

The role of calcium in the effects of noradrenaline and phenoxybenzamine on adrenergic transmitter release from atria: no support for negative feedback of release.

1 The relation of calcium ion influx into nerve terminals to presynaptic adrenoceptor function and the possible masking, by desensitization due to intraneuronal calcium accumulation, of the effects of adrenoceptor agonists and antagonists on presynaptic alpha-adrenoceptors was investigated in guinea-pig atria previously incubated with [(3)H]-noradrenaline.2 Atria were stimulated with 100 pulses at various frequencies (1 to 15 Hz) in standard (2.3 mm), low (0.26 mm) and high (6.9 mm) calcium-Krebs solution in the absence and then the presence first of noradrenaline and subsequently phenoxybenzamine.3 The per pulse overflow of tritium was directly related to the calcium concentration of the Krebs solution, being much reduced and substantially increased in 0.26 and 6.9 mm calcium-Krebs solutions respectively.4 Noradrenaline inhibited the overflow of tritium in low calcium-Krebs solution, to a relatively constant extent, independently of frequency. In addition, the agonist had a greater maximal inhibitory effect in standard than in reduced calcium-Krebs. The catecholamine was as effective an inhibitor of overflow at the lowest and highest frequencies in high as it was in standard calcium-Krebs solution. Phenoxybenzamine invariably increased the tritium overflow but was generally less effective both in low and in high calcium-Krebs solution. The patterns of inhibition and enhancement of stimulation-induced tritium overflow by these two agents do not indicate an intimate relationship between calcium influx and adrenoceptor activation; nor does desensitization appear to be an adequate explanation of the relationship between frequency of stimulation and the intensity of agonist and antagonist effect in the three different calcium concentrations.5 It is concluded that the perineuronal levels of adrenergic transmitter do not establish the magnitudes of effect of exogenous adrenoceptor agonists and antagonists on tritium overflow and that a negative feedback regulation of release by transmitter is exceedingly unlikely under ordinary conditions of neurotransmission.

Mechanisms of inactivation of noradrenaline in the iris sphincter, tracheal muscle and facial artery of cattle: implications for beta-adrenoceptor-mediated responses.

1 The role of neuronal and extraneuronal pathways of amine inactivation in regulating the inhibitory actions of noradrenaline was investigated in three bovine smooth muscle preparations in which the primary adrenoceptor is of the beta-type.2 The extraneuronal uptake inhibitor, 17beta-oestradiol, sensitized the inhibitory responses to noradrenaline in the facial artery, the iris sphincter and in tracheal muscle preparations, indicating a major role for non-neuronal processes in agonist-inactivation in all three preparations. Cocaine also increased responses to noradrenaline, pointing to a role for neuronal uptake either as a terminating mechanism or as a process limiting access of exogenous agonist molecules to their site of action.3 Cocaine did not enhance significantly responses to isoprenaline, a potent beta-adrenoceptor agonist which is not taken up neuronally. Further, relaxations to metaraminol, a sympathomimetic amine which is taken up extraneuronally, but much less so than noradrenaline, were also less enhanced by 17beta-oestradiol in the three preparations tested. These findings support the specificity of action of cocaine and 17beta-oestradiol as neuronal and extraneuronal uptake inhibitors in the present experiments.4 Studies of the uptake of [(3)H]-noradrenaline revealed that 17beta-oestradiol reduced the uptake of amine in the presence of cocaine, confirming a cocaine-resistant site of action for the steroid in all three preparations.5 It is concluded that extraneuronal uptake sites are located sufficiently close to the beta-adrenoceptors to modulate the concentration and duration of action of noradrenaline at these sites of action. It is proposed that in smooth muscles which contain a preponderance of beta-receptors, extraneuronal metabolism is a key event in terminating the inhibitory effects produced.

Yohimbine and prolongation of stimulation pulse duration alter similarly 3H-transmitter efflux in heart: an alternative to the negative feedback hypothesis.

The hypothesis of negative feedback regulation of noradrenaline release was studied in guinea-pig left atrial halves mounted in vitro. Tissues were transmurally stimulated with 30, 100 or 300 pulses at 2 Hz with pulse durations ranging from 50 mus to 2,000 mus, and the efflux of 3H-transmitter determined. The efflux of tritium increased with increasing pulse duration as was anticipated, but the effects of supposed presynaptic antagonism by yohimbine were opposite to expectations for a negative feedback system. The magnification of efflux by yohimbine, compared to untreated controls was less rather than more as stimulation-induced transmitter efflux climbed with increases in pulse duration, and with all other parameters of stimulation held constant. It is concluded that the neuronal effect of yohimbine is not linked to negative feedback or to any other system sensing the perineuronal concentration of previously released transmitter. Analysis of the effects on tritium efflux of yohimbine and of prolongation of the stimulation pulse duration, reveals a similarity in the way that they promote transmitter release. Yohimbine increased efflux to approximately the same value at all pulse durations between 50 and 1,000 mus and the value reached was equivalent to that obtained in untreated atria during stimulation with very long pulses (2,000 mus duration). It is suggested that yohimbine prolongs the outward current attributable to the efflux of potassium from axon terminals, and by this means prolongs depolarization and the period of transmitter release. Tetraethylammonium (TEA), a quaternary ion known to plug potassium efflux channels, had an effect on transmitter efflux that was, in some ways, similar to that of yohimbine but of greater magnitude. The present findings provide, for the first time, an alternative to the hypothesis of negative feedback, that might explain the presynaptic effects of adrenoceptor antagonists and possibly other compounds.

The effects of yohimbine on presynaptic and postsynaptic events during sympathetic nerve activation in cattle iris: a critique of presynaptic receptor theory.

1 The effects of presynaptic alpha-adrenoceptor blockade on both the efflux of 3H-transmitter and on the magnitude of the effector response were measured simultaneously in a smooth muscle preparation which responds to field stimulation with noradrenergic beta-receptor-mediated relaxation. 2 In the presence of atropine, the circular muscle of cattle iris relaxes in response to noradrenaline and to field stimulation at 2 Hz with 10, 20, 50 and 100 pulses. 3 Yohimbine (3 x 10(-6) M), a potent presynaptic alpha-adrenoceptor antagonist, increased the stimulation-induced efflux of tritium to about 2.0 times control values and, contrary to theory, did so to a similar extent regardless of pulse number and with apparent indifference to the synaptic concentration of transmitter, as confirmed by the varying size of the postsynaptic response. 4 In most cases, yohimbine had no significant effect on the magnitude of the relaxations to nerve stimulation. 5 It is concluded that negative feedback regulation of transmitter release, if it functions at all, and this itself seems doubtful, would not have a substantial impact on the size of the effector response.

Clonidine and presynaptic adrenoceptor theory.

The effects of clonidine, a presumed selective presynaptic alpha 2-adrenoceptor agonist or partial agonist, were examined in guinea-pig atria. Split left atrial preparations were stimulated transmurally at 2 Hz with 100 pulses of 0.5 ms duration and the efflux of 3H-transmitter determined. Clonidine inhibited efflux at 3 X 10(-8)M to 3 X 10(-7)M by about 30%. Yohimbine, at a concentration (10(-6)M) which caused a 3 fold increase in the release of 3H-transmitter during field stimulation, did not alter the ability of clonidine to inhibit transmitter efflux. At 10(-6)M clonidine alone had no significant effect on the stimulation-induced efflux of 3H-transmitter, but in the presence of yohimbine (10(-6)M), inhibited efflux by over 50%. The inhibitory effect of noradrenaline (10(-6)M) on 3H-transmitter efflux was antagonized by clonidine at 10(-6)M but not at 10(-8)M, although neither concentration of clonidine alone inhibited transmitter efflux. The present findings indicate that the effects of clonidine on the efflux of noradrenaline from sympathetic nerves cannot be accommodated within the currently held view that the compound is an agonist or partial agonist on presynaptic alpha 2-adrenoceptors. It appears that clonidine has multiple sites of action few of which are antagonized by a concentration of the prototypical presynaptic antagonist yohimbine, which enhances efflux 3 fold.

The effect of magnesium deficiency and excess on bovine coronary artery tone and responses to agonists.

1 The hypothesis that magnesium deficiency, linked to the magnesium content of drinking water, induces major tone increases in coronary arteries and enhances their responses to vasoactive agents to an extent sufficient to explain sudden death associated with ischaemic heart disease was examined in an in vitro preparation. 2 The spontaneous tone of cattle coronary arteries was not increased during a 30 min exposure to Mg2+-deficient Krebs until the mineral was omitted entirely from the bathing medium, and even then the observed increase was small. Only in strips maintained under extremely deficient conditions for a prolonged period, namely Mg2+ concentration of 0.2 mM and 0.0 mM for 3 h, was tone substantially greater than in controls in standard (1.2 mM) Mg2+-Krebs. 3 Responses to acetylcholine and to noradrenaline were not increased in Mg2+-free Krebs but those to potassium and to 5-hydroxytryptamine were enlarged over the lower parts of their concentration-response curves. Responses to potassium and to 5-hydroxytryptamine were also examined in Krebs containing very low concentration of Mg2+ (0.4 and 0.2 mM) and only modest increases in contraction size were detected. Increases in the Mg2+ concentration of the Krebs (to 4.8 mM) depressed responses to potassium and 5-hydroxytryptamine. 4 It is concluded that Mg2+ deficiency must be nearly complete (0.4-0.0 mM) to induce even moderate tone increases in coronary vessels, or to sensitize them to agonist responses, and that there is no reason to link marginally subnormal Mg2+ levels, occasionally reported in humans with heart disease, to marked changes in coronary dynamics.

Sensitization of noradrenaline responses by inhibitors of extraneuronal uptake in a coronary artery preparation.

Inhibition of neuronal uptake with cocaine had no significant effect on the dose-response curve to noradrenaline in strips of beef coronary artery. However, responses were enhanced about 3-fold by 17beta-oestradiol and the haloalkylamine GD-131, two known inhibitors of extraneuronal uptake.

A new approach to the measurement and classification of forms of supersensitivity of autonomic effector responses.

1 It is proposed that sensitizations of autonomic effectors to agonists by drugs or procedures be considered in two main categories: those involving changes in the effective concentration of agonist at receptors (type I) and those involving changes in the responding tissue beyond the initial combination of agonist and receptors (type II). Type I sensitizations are appropriately described by determining the dose-ratio (horizontal shift of the dose-response curve) and type II sensitizations by assessing the change in the magnitude of the response.2 The inadequacy of the dose-ratio in assessing sensitizations related to an altered physiology of the responding tissue is illustrated by means of hypothetical examples with particular reference to the slopes of dose-response curves and altered maximal responses.3 An evaluation of the enhancement of responses of rabbit aortic strips to agonists by reserpine indicates that it is a type II sensitization. The shifts of dose-response curves to noradrenaline, isoprenaline, normetanephrine and 5-hydroxytryptamine after reserpine-treatment, were described both by the dose-ratio and by the increment in the magnitude of the response at various contraction amplitudes. The dose-ratio varied unpredictably for each agonist depending on the response level selected for comparison and also varied between agonists. However, the mm increment in response magnitude after reserpine approximated a constant value. Responses to potassium which by horizontal procedures were assessed among the least increased, were found to be enhanced the most when considered as a type II sensitization.4 It is concluded that both type I and type II procedures should be applied when dealing with an unidentified sensitization and that the data be critically assessed. The appropriate use of these procedures can aid in identifying and clarifying sensitizations, as well as in elucidating the sequence of steps between receptor activation and response in an effector.

A vasodilator innervation to the central artery of the rabbit ear.

1 The possibility of a vasodilator innervation to the isolated and perfused central artery of the rabbit ear was examined.2 Stimulation of the periarterial nerves in the presence of noradrenaline or other agonist used to maintain a partial constriction of the ear artery, led to a decrease in intraluminal flow followed after the cessation of stimulation by an increase in flow beyond the pre-stimulation level.3 After blockade of adrenergic transmission with bretylium or guanethidine or of the alpha- and beta-adrenoceptors with phentolamine and propranolol, stimulation of the periarterial nerves in the presence of a background tone, led to a clearly detectable vasodilation. This dilatation was not blocked by treatment with atropine or mepyramine; nor was it enhanced by physostigmine.4 Pretreatment of rabbits with reserpine (2 mg/kg) to deplete catecholamine stores, eliminated both the vasoconstrictor and vasodilator responses to nerve stimulation. However, a lower dose of reserpine (0.2 to 0.5 mg/kg) selectively eliminated the vasoconstrictor component of periarterial nerve activation.5 The ear artery dilated in response to low concentrations of prostaglandin E(1), and E(2), in the presence of noradrenaline, but treatment with inhibitors of prostaglandin synthesis, indomethacin, aspirin or eicosa-5,8,11,14-tetraynoic acid did not reduce the vasodilator response. Attempts to extract a prostaglandin in the bathing medium were unsuccessful.6 The involvement of a purine nucleotide appeared unlikely since the ear artery dilated only in response to fairly high concentrations of adenosine 5'-triphosphate (ATP), adenosine 5'-diphosphate (ADP) and adenosine 5'-monophosphate (AMP). Furthermore, dipyridamole, an inhibitor of adenosine uptake, enhanced dilation due to exogenous ATP but not to periarterial nerve stimulation.7 It is concluded that the central artery of the rabbit ear has a vasodilator innervation but the identity of the transmitter remains to be established.

Steroid potentiation of responses to sympathomimetic amines in aortic strips.

1. Responses to catecholamines (adrenaline, noradrenaline, nordefrine) were enhanced by 17beta-oestradiol, progesterone and desoxycorticosterone in untreated and reserpine pretreated aortic strips. Responses to tyramine, believed mediated via endogenous catecholamines, were enhanced only in untreated strips.2. Responses to sympathomimetic amines lacking the catechol nucleus (phenylephrine, synephrine, methoxamine) were potentiated inconsistently by the steroids and reserpine pretreatment reduced markedly the frequency of potentiated responses.3. Known inhibitors of catechol-O-methyl transferase (tropolone, U-0521, pyrogallol) potentiated responses to catecholamines and abolished the enhancing effects of the steroids-when the steroids were given first, there was no further increase in response to catecholamines on adding inhibitors of catechol-O-methyl transferase.4. Experiments with the oil-immersion technique, to eliminate diffusion of drug from the tissue, indicated that 17beta-oestradiol, progesterone and desoxycorticosterone decreased the rate at which aortic strips inactivated adrenaline by O-methylation.5. It is concluded that 17beta-oestradiol, progesterone and desoxycorticosterone potentiate responses to catecholamines in aortic strips by inhibiting a major mechanism for their inactivation.

Evidence against the unitary hypothesis of agonist and antagonist action at presynaptic adrenoceptors.

1 The concept that presynaptic receptors regulate noradrenergic transmitter release via a system of inhibitory receptors mediating negative feedback relies on a supposed association between increases in stimulation-induced efflux of [3H]-noradrenaline by antagonists and blockade by them of the inhibitory effects of exogenous noradrenaline. 2 It was shown in guinea-pig ureter, that yohimbine (3 X 10(-7)M), a presumed selective presynaptic antagonist, increased transmitter efflux substantially at 1 Hz and 5 Hz with 100 pulses, purportedly representing antagonism of the inhibitory effect of locally released noradrenaline but did not reduce the inhibitory effect of exogenous noradrenaline (1.8 X 10(-6)M or 1.8 X 10(-7)M) except in one case. 3 Additionally, the inhibitory effect of oxymetazoline (1.0 X 10(-7)M or 1.0 X 10(-8)M) on stimulation-induced efflux was in no way antagonized by yohimbine (3 X 10(-7)M). 4 It is concluded that the increased efflux of [3H]-noradrenaline produced by antagonists and the decreased efflux produced by exogenous agonists may represent actions at different loci and that the hypothesis of presynaptic feedback regulatory sites is still not substantiated.

The effects of (+)- and (-)-propranolol on 3H-transmitter efflux in guinea-pig atria and the presynaptic beta-adrenoceptor hypothesis.

The effects of isoprenaline and of the (+)- and (-)-isomers of propranolol on the stimulation-induced overflow of 3H-transmitter was assessed in guinea-pig atria to evaluate the hypothesis of presynaptic beta-adrenoceptors. 2 Isoprenaline (1.2 x 10(-8) M) enhanced the efflux of tritium at 2 and 5 Hz with 100 pulses and did so to a similar extent at both frequencies. 3 The (-)-isomer of propranolol (1.0 x 10(-7) M) blocked the enhancing effect of isoprenaline but did not by itself modify transmitter efflux. 4 The (+)-isomer of propranolol, almost devoid of beta-adrenoceptor blocking properties, was also effective at 1.0 x 10(-7) M in blocking the enhancement of tritium efflux by isoprenaline. 5 The (-)-isomer of propranolol (1.0 x 10(-7) M) blocked almost entirely the inotropic response to isoprenaline (3 x 10(-7) M) but even 3.0 x 10(-6) M (+)-propranolol was ineffective in antagonizing the beta-adrenoceptor-mediated contractile responses to the catecholamine. 6 It is concluded that the presynaptic site of isoprenaline action does not show the requisite stereo-specificity of beta-adrenoceptors and that a 'non-specific' action of the antagonist probably accounts for its reduction of the effect of isoprenaline.

The lack of effect of oxytetracycline on responses to sympathetic nerve stimulation and catecholamines in vascular tissue.

The effects of oxytetracycline, an inhibitor of amine binding in connective tissue, on the responses of perfused rabbit ear arteries to sympathetic nerve stimulation and to intraluminally administered noradrenaline were examined. The contractions of aortic strips to catecholamines in the presence of oxytetracycline were also examined. Oxytetracycline (0.1 mM) had no discernable effect on the magnitude of constrictions, measured as reductions in flow, produced by either nerve stimulation (0.5-10 Hz) or noradrenaline (0.5-50 ng) in the ear artery. In addition, the time taken for vessels to recover towards control flow values after endogenously released or exogenously applied noradrenaline had acted was not increased by oxytetracycline. Oxytetracycline (0.1 mM) did not alter the position or shape of the concentration-response curve to noradrenaline nor did it enhance the amplitude of individual responses to catecholamines in aortic strips. It is concluded, contrary to the observations of Powis (1973), that oxytetracycline does not increase the magnitude or duration of responses to sympathetic nerve activation or to catecholamines and that binding to connective tissue is of no material consequence in terminating their action in vascular tissue.

Role of extraneuronal mechanisms in the termination of contractile responses to amines in vascular tissue.

1 The role of the uptake and release of agonist from extraneuronal sites in the termination of responses of rabbit aortic strips to amines was studied. 2 Strips were contracted with adrenaline or noradrenaline and after response plateau was reached, the muscle chambers were washed free of agonist and the relaxation in Krebs solution recorded. After inhibition of catechol-O-methyl-transferase, monoamine oxidase and neuronal uptake the relaxation rate was greatly prolonged. Evidence is provided that this very slow relaxation resulted from the accumulation of intact amine at extraneuronal sites during exposure to the agonist and its subsequent release past receptors due to a reversal of the concentration gradient after washout. 3 Pretreatment with the haloalkylamine, GD-131 (N-cyclohexylmethyl-N-ethyl-beta-chloroethylamine), an inhibitor of extraneuronal uptake, returned the slow relaxation rate after enzyme inhibition towards that of control strips. By blocking the extraneuronal transport of amines their accumulation at intracellular loci after enzyme inhibition was prevented. 4 The effects of GD-131 and 17beta-oestradiol on the relaxation rate of untreated strips contracted by adrenaline and noradrenaline confirmed that extraneuronal uptake to sites of enzymatic activity is the major mechanism terminating their action. 5 Inactivation of extraneuronal transport sites by GD-131 was prevented by protecting them with 17beta-oestradiol or normetanephrine during exposure to the haloalkylamine, pointing to a common site of action of these agents on a specific carrier system for amines. 6 Evidence is presented that the relaxation from contractions induced by histamine and 5-hydroxytryptamine also involves extraneuronal accumulation and release, probably by an uptake process which is identical to the one for catecholamines.

Adenosine and dipyridamole actions and interactions on isolated coronary artery strips of cattle.

1 The actions and interactions of adenosine and dipyridamole were investigated on isolated strips of coronary arteries of beef cattle. It was found that small diameter arteries (about 0.5-1.0 mm o9d.), raised to a moderate level of tone with potassium, responded with relaxation to low concentrations of adenosine. 2 Dipyridamole, over a broad concentration range (6.0 X 10(-8)-2.0 X 10(-5)M), enhanced these responses, shifting the adenosine concentration-response curve (3.7 X 10(-8)-1.1 X 10(-4)M) considerably to the left. In contrast, inhibitory concentrations-response curves to sodium nitrite and to noradrenaline were not materially altered by dipyridamole. 3 Studies of the uptake of [3H]-adenosine revealed a rapid uptake of the nucleoside by coronary artery strips, which was inhibited by dipyridamole (6.0X 10(-8)-2.0X10(-5)M); but this may not be sufficient to account fully for the observed sensitization. 4 It is concluded that the regulation of adenosine responses and the action of dipyridamole in the heart involve a more direct association with coronary vascular tissue than has been previously appreciated.

Effects of the inhibition of noradrenaline uptake and synthesis on the maintenance of the response to continuous nerve stimulation in the central artery of the rabbit ear.

1. The central artery of the rabbit ear was perfused through its lumen in vitro, with a constant pressure technique, and stimulated continuously via its periarterial sympathetic nerves at the physiological frequency of 5 Hz.2. The vasoconstrictor response, which led initially to an almost complete cessation of intraluminal flow, deteriorated steadily over a period of hours. The involvement of presynaptic mechanisms in this effect was indicated by the finding that noradrenaline, administered extraluminally, produced a similar response before the onset of continuous stimulation and at a late stage when the constriction had decreased markedly. In addition, the noradrenaline precursor DOPA, restored the depressed responses towards their original values, indicating that failure involved depletion of mediator for release.3. Responses to continuous stimulation declined significantly faster after inhibition of tyrosine hydroxylase with alpha-methyl-p-tyrosine. However, inhibition of the uptake of noradrenaline with cocaine did not enhance the decline of the response, even when the sensitization produced by the compound was taken into account.4. It is concluded that synthesis, along with the mobilization of stored mediator, rather than uptake and re-use of noradrenaline maintain the effector response in the central artery of the rabbit ear stimulated continuously at a frequency within the physiological range.

Mechanism of potentiation by amines of non-equilibrium blockade of the alpha-adrenoceptor.

1. The mechanism by which sympathomimetic and certain other amines enhance blockade of the alpha-adrenoceptors by the non-equilibrium antagonist N-ethoxycarbonyl-2-ethoxy-1,2-dihydroquinoline (EEDQ) in strips of rabbit aorta was examined.2. Non-equilibrium blockade of the 5-hydroxytryptamine receptors by EEDQ was not increased by sympathomimetic amines and was decreased by 5-hydroxytryptamine.3. Low concentrations of reversible competitive antagonists appeared to protect selectively against the additional blockade by EEDQ which develops in the presence of an amine.4. Phenoxybenzamine potentiated EEDQ blockade of the alpha-receptors but not of the 5-hydroxytryptamine receptors.5. Augmentation of EEDQ blockade was also detected in a variety of other tissues, but not in segments of rabbit intestine where alpha-adrenoceptors mediate an inhibitory response.6. It was concluded that EEDQ acts at two sites in antagonizing alpha-receptor mediated responses, and that one of these sites (site II) is separate from the site of action of agonists and phenoxybenzamine (site I). Amines which enhance blockade appear to exert their action by combining with a third site (site III), which may induce a conformational alteration at site II.7. It appears that the alpha-adrenoceptor may have multiple sites for drug interaction.