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To clarify the effects of hypoxia on adrenergic transmission, we examined the contractile responses of isolated rabbit aortic strips to electrical stimulation, the concentration-response relationships for noradrenaline and KCl, and the electrical stimulation-evoked overflows of total [H] and [H]noradrenaline from strips preloaded with [H]noradrenaline in media equilibrated with gas mixtures containing various concentrations of 0. Contractile responses to electrical stimulation were completely inhibited by tetrodotoxin and α-adrenoceptor antagonists such as phentolamine and phenoxybenzamine, but were not affected by indomethacin. When the concentration of O in the gas mixture was decreased from 95% to 20%, the contractile responses to electrical stimulation remained unchanged, but as the concentration of OZ was further decreased, the responses were inhibited concentration-dependently. At 0% O, the response was inhibited by about 80% when compared with control values obtained at 95% O, and the electrical stimulation-evoked overflows of total [H] and [H]noradrenaline into the superfusates were decreased by about 55%. At 0% 02, the concentration-response curve for exogenous noradrenaline was shifted to the right about 50-fold and the maximum response was decreased by 25%. The maximum contractile responses of aortic strips from animals pretreated with reserpine or 6-hydroxydopamine to high KCl were decreased slightly (about 15%). These results suggest that inhibition of adrenergic transmission under hypoxic conditions is mainly the result of a decrease in the stimulus-evoked release of noradrenaline and of a decrease in the affinity of α-adrenoceptor for noradrenaline and/ or inhibition of signal transduction mechanisms, although hypoxia also causes a slight decrease in the contractility of vascular smooth muscle. [ABSTRACT FROM AUTHOR]

The effect of N-ethylmaleimide (NEM), which has been shown to abolish rather selectively inhibition of adenylate cyclase, on the α-adrenoceptor modulation of noradrenaline release was studied. Slices of the rabbit hippocampus were loaded with H-noradrenaline, superfused continuously and stimulated twice electrically. NEM (30 μmol/l) applied for 30 min enhanced both basal and stimulation-evoked tritium overflow significantly. Occupation of the receptor by the α-adrenoceptor agonist clonidine prior to and during NEM treatment did not protect the α-adrenoceptor-mediated autoinhibitory feedback system from being affected by NEM. Preincubation of the hippocampal slices with NEM was without any influence on H-noradrenaline uptake. The inhibitory effect of clonidine on H-noradrenaline release was attenuated in a non-competitive manner. In addition, the facilitatory effect of the α-adrenoceptor antagonist yohimbine on the stimulusevoked tritium overflow was reduced. The facilitation of the evoked noradrenaline release by yohimbine or yohimbine and NEM converged with increasing concentrations of yohimbine, suggesting that yohimbine and NEM were acting at the same signal-transduction system. These results are compatible with the idea that NEM, by alkylating the N-unit of a presynaptically located adenylate cyclase, prevents the α-adrenoceptor-mediated modulation of noradrenaline release. [ABSTRACT FROM AUTHOR]

The influence of clonidine on the stimulation-evoked overflow of tritium was studied in brain slices preincubated withH-noradrenaline. The slices were prepared from parietal cortex (Cx), nucleus anterior hypothalami (nah) and nucleus tractus solitarii (nts). After preincubation, the tissues were superfused at 23°C or 37°C with a medium containing the noradrenaline uptake inhibitor desipramine. Electrical field stimulation was applied using stimulation frequencies of 0.3-10 Hz. [ABSTRACT FROM AUTHOR]

The effect of islet-activating protein (IAP) on α-adrenoceptor mediated modulation of noradrenaline release in the rabbit hippocampus was studied. Slices of the hippocampus were incubated for 6 h with IAP, subsequently loaded withH-noradrenaline and superfused continuously. IAP-pretreatment significantly enhanced the electrically evoked transmitter release and diminished the facilitatory effect of the α-adrenoceptor antagonist yohimbine. In addition, the inhibitory effect of the α-adrenoceptor agonist clonidine was reduced. These results provide circumstantial evidence that an inhibitory guanine-nucleotide-binding protein, most probably N of a presynaptically located adenylate cyclase, is involved in the α-autoreceptor mediated modulation of noradrenaline release. [ABSTRACT FROM AUTHOR]

Brain cortex slices from rabbits were preincubated with [H]noradrenaline and then superfused and stimulated electrically at 3Hz. In the presence of cocaine 30 μM, unlabelled noradrenaline, α-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 α-adrenoceptors which are exclusively of the α-type. Addition of various concentrations of cocaine, addition of pargyline, or stimulation at different current strengths was used to obtain either a high or a low stimulation-evoked overflow of tritium. Independently of the method used, a low evoked overflow coincided with a large percentage inhibition produced by 0.1μM 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 α-adrenergic agonists by increasing the biophase concentration of released noradrenaline. [ABSTRACT FROM AUTHOR]

Presynaptic α-autoreceptors in mouse atria were characterized in terms of the α, α, α and α subtypes. Segments of the atria were preincubated with H-noradrenaline and then superfused and stimulated electrically. The affinity of up to 16 antagonists for the autoreceptors was assessed as (1) pEC values, i.e. concentrations that increased previously autoinhibited release of H-noradrenaline (120 pulses, 3 Hz) by 30%, and (2) pK values against the release-inhibiting effect of 5-bromo-6-(2-imidazolin-2-ylamino)-quinoxaline (UK 14,304) under conditions of no or little autoinhibition (2 trains of 20 pulses, 50 Hz, train interval 120 s). The pK values correlated well with the pEC values ( r = 0.98; P<0.001; slope of regression line 0.93), indicating that UK 14,304 and released noradrenaline modulated the release of noradrenaline through pharmacologically identical receptors. Comparison with antagonist affinities for (1) prototypic native α radioligand binding sites, (2) radioligand binding sites in COS cells transfected with α subtype genes, and (3) previously classified presynaptic α-autoreceptors - all taken from the literature - indicated that the mouse atrial autoreceptors corresponded to the α subtype. For example, the pK values at mouse atrial autoreceptors correlated closely with pK values at native α binding sites in the bovine pineal gland ( r = 0.96; P<0.001); with pK values at α binding sites in COS cells transfected with the rat α gene ( r= 0.85; P<0.01); and with pK values at guinea-pig cerebral and atrial and mouse cerebral α-autoreceptors ( r=0.96-0.98; P<0.001). The antagonist pK values at mouse atrial autoreceptors correlated less with pK values at α, α, and α sites. It is concluded that the presynaptic α-autoreceptors in mouse atria are α This identification supports the hypothesis that at least the majority of α-autoreceptors belong to the α pair of orthologous α-adrenoceptors. [ABSTRACT FROM AUTHOR]

The study was devised to classify, by means of antagonist affinities, the presynaptic α-autoreceptors of rabbit atria and kidney in terms of α, α, α and α-A set of antagonists was chosen that was able to discriminate between the four subtypes. Small pieces of the left atrium and slices of the kidney cortex were preincubated with H-Noadrenaline and then superfused and stimulated electrically. In one series of experiments, tissue pieces were stimulated by relatively long pulse trains (1 or 2 min) leading to α-autoinhibition. All 11 (atria) or 10 (kidney) antagonists increased the evoked overflow of tritium. pEC values (concentrations causing 30% increase) were interpolated from concentration-response curves. In a second series of experiments, tissue pieces were stimulated by brief pulse trains (0.4 s) that did not lead to α-autoinhibition, and concentration-inhibition curves of the α-selective agonist 5-bromo-6-(2-imidazolin-2-ylamino)-quinoxaline (UK 14,304) were determined. Most of the 11 (atria) or 8 (kidney) antagonists shifted the concentration-inhibition curve of UK 14,304 to the right. pK values of the antagonists were calculated from the shifts. pEC values correlated with pK values, both in atria ( r = 0.728) and in the kidney ( r = 0.930). pEC values in atria correlated with pEC values in the kidney ( r = 0.988) and pK values in atria correlated with pK values in kidney ( r = 0.923). It is concluded that the α-autoreceptors in atria and the kidney are the same. Comparison with antagonist affinities for prototypic native α binding sites, α binding sites in cells transfected with α subtype genes, and previously classified presynaptic α-adrenoceptors - all taken from the literature - indicates that both autoreceptors are α. This conclusion is reached with either of the two independent estimates of autoreceptor affinity, pEC and pK. The results are compatible with the hypothesis that at least the majority of α-autoreceptors belong to the α branch of the α-adrenoceptor tree, across mammalian or at least rodent and lagomorph species. [ABSTRACT FROM AUTHOR]

The study was devised to classify, by means of antagonist affinities, the presynaptic α-autoreceptors in mouse cerebral cortex in terms of α, α, α and α. A set of antagonists was chosen that was able to discriminate between the four subtypes. Slices of the cortex were preincubated with H-noradrenaline and then superfused and stimulated electrically. The stimulation periods used (4 pulses, 100 Hz) did not lead to α-autoinhibition as shown by the lack of an increase by rauwolscine of the evoked overflow of tritium. The α-selective agonists 5-bromo-6-(2-imidazolin-2-ylamino)-quinoxaline (UK 14,304) and α-methylnoradrenaline reduced the evoked overflow. All 10 antagonists shifted the concentration-inhibition curve of UK 14,304 to the right. Rauwolscine also shifted the concentration-inhibition curve of α-methylnoradrenaline to the right. pK values of the antagonists were calculated from the shifts. The pK values of rauwolscine against UK 14,304 and α-methylnoradrenaline were very similar (8.0 and 7.9, respectively). Comparison with antagonist affinities for prototypic native α binding sites, α binding sites in cells transfected with α subtype genes, and previously classified presynaptic α-adrenoceptors - all taken from the literature - indicates that the α-autoreceptors in mouse brain cortex are α. This is the first subtype determination of α-autoreceptors in the mouse. It supports the hypothesis that at least the majority of α-autoreceptors belong to the α branch of the α-adrenoceptor tree. [ABSTRACT FROM AUTHOR]

The study was devised to classify, by means of antagonist and agonist affinities, the presynaptic α-autoreceptors in guinea-pig heart atria and brain cortex in terms of α, α, α and α. A set of antagonists and agonists was chosen that was able to discriminate between the four subtypes. Small pieces of the atria and slices of the brain cortex were preincubated with H-noradrenaline and then superfused and stimulated electrically. In one series of experiments (atria only), tissue pieces were stimulated by relatively long pulse trains (1 min) leading to marked α-autoinhibition. All 10 antagonists increased the evoked overflow of tritium. pEC values (concentrations causing 30% increase) were interpolated from concentration-response curves. In a second series of experiments (atria and brain slices), tissue pieces were stimulated by brief pulse trains (0.4 s or 40 ms) that led to little (atria) or no (brain slices) α-autoinhibition, and antagonist effects against the α-selective agonist 5-bromo-6(2-imidazolin-2-ylamino)-quinoxaline UK 14,304 were examined. All 10 (atria) or 8 (brain) antagonists shifted the concentration-inhibition curve of UK 14,304 to the right. pK values of the antagonists were calculated from the shifts. In a third series of experiments (brain slices only), also with brief pulse trains (40 ms), pK values (negative logarithms of dissociation constants of agonist-α-adrenoceptor complexes) were determined by comparison of concentration-inhibition curves of UK 14,304, guanoxabenz and oxymetazoline in normal tissue and in tissue in which a fraction of the receptors had been blocked by phenoxybenzamine. pEC values in atria correlated with pK values ( r = 0.942). pK values in atria correlated with pK values in the brain cortex ( r = 0.970). It is concluded that the α-autoreceptors in atria and the brain cortex are the same. Comparison with antagonist affinities for prototypic native α binding sites, binding sites in cells transfected with α subtype genes, and previously classified presynaptic α-adrenoceptors - all taken from the literature - indicates that both autoreceptors are α. In atria, this identification is reached with either of the two independent estimates of autoreceptor affinity, pEC and pK, and in the brain cortex it is supported by the agonist pK values. The results are compatible with the hypothesis that at least the majority of α-autoreceptors belong to the α branch of the α-adrenoceptor tree. [ABSTRACT FROM AUTHOR]

The study was designed to classify in terms of α, α, α and α the presynaptic α-autoreceptors, as well as the α-receptors modulating the release of acetylcholine, in the myenteric plexus-longitudinal muscle (MPLM) preparation of the guinea-pig ileum. A set of antagonists was chosen that was able to discriminate between the four subtypes. Small pieces of the MPLM preparation were preincubated with H-noradrenaline or H-choline and then superfused and stimulated electrically. The stimulation periods used (3H-noradrenaline: 3 trains of 20 pulses, 50 Hz, train interval 60 s; H-choline: single trains of 30 pulses, 0.2 Hz) did not lead to α-autoinhibition or inhibition of H-acetylcholine release by endogenous noradrenaline. The α-selective agonist 5-bromo6-(2-imidazolin-2-ylamino)-quinoxaline (UK 14,304) reduced the evoked overflow of tritium in both H-noradrenaline and H-choline experiments. Most (H-noradrenaline) or all (H-choline) of the 10 antagonists shifted the concentration-inhibition curves of UK 14,304 to the right. pK values of the antagonists were calculated from the shifts. pK values from H-noradrenaline experiments correlated with pK values from H-choline experiments ( r = 0.981). It is concluded that α-autoreceptors and α-heteroreceptors modulating the release of acetylcholine in the MPLM preparation are of the same subtype. Comparison with antagonist affinities for prototypic native α binding sites, binding sites in cells transfected with α subtype genes, and previously classified presynaptic α-adrenoceptors - all taken from the literature - indicates that both are α. The results are consonant with the hypothesis that at least the majority of α-autoreceptors belong to the α branch of the α-adrenoceptor tree, across mammalian or at least across rodent and lagomorph species. The same may hold true for α-adrenoceptors on non-noradrenergic neurones. [ABSTRACT FROM AUTHOR]

An involvement of imidazoline recognition sites in the modulation of transmitter release was investigated in the rabbit pulmonary artery and aorta preincubated with [H]noradrenaline and superfused with physiological salt solution containing cocaine, corticosterone and propranolol. Electrical impulses were applied transmurally at 0.66 or 2 Hz. In the absence of further drugs, rauwolscine as well as the imidazoline derivatives BDF 6143 [4-chloro-2-(2-imidazoline-2-ylamino)-isoindoline], idazoxan and phentolamine increased the H overflow from the pulmonary artery, evoked by electrical stimulation at 2 Hz; the effect was due to the α-autoreceptor blocking property of these drugs. The maximum increase in overflow obtainable with the imidazolines was considerably lower than with rauwolscine. The concentration-response curves of the imidazolines were bell-shaped. At 0.66 Hz, BDF 6143 did not facilitate, but concentration-dependently inhibited, whereas idazoxan failed to change the evoked H overflow. When, at the stimulation frequency of 2 Hz, presynaptic α- adrenoceptors were blocked by rauwolscine and/or pre-exposure to phenoxybenzamine, the electrically evoked H overflow from the pulmonary artery and/or aorta was inhibited by the following imidazoline derivatives: the α-adrenoceptor antagonists BDF 6143, idazoxan and phentolamine, the α-adrenoceptor agonist with α-blocking property cirazoline as well as the α-adrenoceptor agonists clonidine and moxonidine. The maximum inhibition caused by BDF 6143 was greater than that due to clonidine and moxonidine; the latter two, hence, behaved as partial agonists. At the stimulation frequency of 0.66 Hz, the imidazolines exhibited a higher potency than, but a similar intrinsic activity to that at 2 Hz. Noradrenaline did not affect the evoked H overflow. The BDF 6143-induced inhibition of evoked H overflow was not modified by metitepine, atropine, theophylline, dipyridamole and indometacin, but was counteracted by the partial agonists clonidine and moxonidine. The results exclude the possibility that α- and α-adrenoceptors, 5-HT receptors, muscarine receptors, P purinoceptors and prostaglandin receptors are involved in the imidazoline-induced inhibition of noradrenaline release. They provide evidence indicating that the inhibitory effect is mediated by imidazoline receptors on the postganglionic sympathetic nerve terminals of the rabbit pulmonary artery and aorta. [ABSTRACT FROM AUTHOR]

Segments of the rat vena cava preincubated with [H]noradrenaline and superfused with [H]noradrenaline-free solution containing desipramine and corticosterone were stimulated electrically (standard parameters: 150 mA, 0.3 ms, 0.66 Hz; duration 6 min). In some experiments the stimulation parameters were modified in order to obtain similar absolute release values despite the presence of an α-adrenoceptor agonist or antagonist or of 5-hydroxytryptamine (5-HT). In a first set of experiments, the vascular segments were first exposed to an α-adrenoceptor ligand, which was kept present throughout the remainder of superfusion, and then to 5-HT. The release-inhibiting effect of 5-HT was attenuated by the α-adrenoceptor agonists clonidine and B-HT 920 whereas it was enhanced by the α-adrenoceptor antagonists phentolamine and idazoxan. The α-adrenoceptor antagonist prazosin did not change the 5-HT-induced inhibition of noradrenaline (NA) release. In a second set of experiments, 5-HT was administered first and kept present in the superfusion fluid for the remainder of the experiment. In the presence of 5-HT, the overflow-inhibiting effects of B-HT 920 and clonidine and the overflow-enhancing effect of idazoxan were reduced. The results demonstrate that activation of one kind of receptor decreased the inhibition of noradrenaline release produced by activation of the other. These effects were not the consequence of the change of release per se induced by the interacting drugs, since they also occurred when the release was adjusted to similar levels by modification of the stimulation parameters. Therefore, the effects are probably due to mutual interaction between presynaptic α-adrenoceptors and 5-HT receptors. The level (receptors themselves, G proteins and/or ion channels) at which this interaction occurs is unknown. [ABSTRACT FROM AUTHOR]