Pharmacological characterization of two distinct alpha 1-adrenoceptor subtypes in rabbit thoracic aorta.

1. alpha 1-Adrenoceptor subtypes in rabbit thoracic aorta have been examined in binding and functional experiments. 2. [3H]-prazosin bound to two distinct populations of alpha 1-adrenoceptors (pKD,high = 9.94, Rhigh = 79.2 fmol mg-1 protein; pKD,low = 8.59, Rlow = 215 fmol mg-1 protein). Pretreatment with chloroethylclonidine (CEC, 10 microM) almost inactivated the prazosin-high affinity sites and reduced the number of the low affinity sites without changing the pKD value. 3. In the displacement experiments with CEC-untreated membranes, unlabelled prazosin, WB4101 and HV723 displaced the binding of 200 pM [3H]-prazosin monophasically; the affinities for WB4101 (pK1 = 8.88) and HV723 (8.49) were about 10 times lower than that for prazosin (9.99). In the CEC-pretreated membranes also, the antagonists inhibited the binding of 1000 pM [3H]-prazosin monophasically; the pK1 values for prazosin, WB4101 and HV723 were 9.09, 8.97 and 8.17, respectively. These results suggest that the prazosin-high and low affinity sites can be independently appraised in the former and latter experimental conditions. Noradrenaline, but not methoxamine, showed slightly higher affinity for the prazosin-high affinity site than for the low affinity site. 4. In the functional experiments, noradrenaline (0.001-100 microM) and methoxamine (0.1-100 microM) produced concentration-dependent contractions. Pretreatment with CEC inhibited the contractions induced by low concentrations of noradrenaline but without effect on the responses to methoxamine.(ABSTRACT TRUNCATED AT 250 WORDS)     

Heterogeneity of alpha 1-adrenoceptor subtypes involved in adrenergic contractions of dog blood vessels.

1. We determined the alpha 1-adrenoceptor subtypes involved in adrenergic contractions of eight different blood vessels isolated from the dog. 2. Noradrenaline produced concentration-dependent contractions in all the blood vessels tested, which were competitively inhibited by prazosin, WB4101, HV723 and 5-methylurapidil. However, there was considerable difference between the vessels with regard to the pKB values for all the antagonists. The alpha 1-adrenoceptors of dog vertebral and carotid arteries had high affinity for prazosin (pKB > 9.0) but low affinity for WB4101 (< 8.5), 5-methylurapidil (< 7.5) and HV723 (< or = 8.5). By contrast, HV723 had higher affinity (> 9.0) than prazosin (< 8.3), WB4101 (< 8.7) and 5-methylurapidil (< 8.2) in the portal vein, mesenteric artery and vein, and renal artery. In the femoral artery and vein, however, the four antagonists showed pKB values in the range 8.0-8.7. 3. Chloroethylclonidine (10 microM) produced a remarkable reduction of the contractile responses to noradrenaline in the vertebral and carotid arteries as compared with those in the other vessels. Nifedipine inhibited the responses to noradrenaline in all the tissues tested, and had marked effects in the portal vein. 4. Sympathetic adrenergic contractions induced by transmural electrical stimulation were also inhibited by prazosin and HV723 at different potencies among tissues. The relative potencies of both the antagonists paralleled the relationship in inhibiting the responses to exogenous noradrenaline in each vessel.(ABSTRACT TRUNCATED AT 250 WORDS)     

Pharmacological profiles of a novel a1-adrenoceptor agonist, PNO-49B, at a1-adrenoceptor subtypes

The effects of a newly synthesized compound, PNO-49B, (R)-(–)-3'-(2-amino-1-hydroxyethyl)-4'-fluoromethanesulfonanilide hydrochloride, on a ₁-adrenoceptor subtypes were examined in various tissues in which the following distribution of a ₁-adrenoceptor subtypes has been suggested: dog carotid artery (a _1B), dog mesenteric artery (a _1N), rabbit thoracic aorta (a _1B+a _1L), rat liver (a _1B), rat vas deferens (a _1A+a _1L), rat cerebral cortex (a _1A+a _1B) and rat thoracic aorta (controversial subtype). PNO-49B (0.1–100?µM) produced concentration- dependent contractions in dog mesenteric artery, rabbit thoracic aorta, rat thoracic aorta and rat vas deferens; and the maximal amplitudes of contraction were almost the same as or slighly less than those of noradrenaline. By contrast, the maximal response to PNO-49B in dog carotid artery was markedly smaller than the response to noradrenaline. In rabbit thoracic aorta, the contractile response to PNO-49B was not affected by inactivation of the a _1B subtype with chloroethylclonidine (CEC), although the response to noradrenaline was attenuated by that treatment. The dissociation constants (K _A) of PNO-49B were not different among the rat thoracic aorta, dog carotid and mesenteric arteries and rabbit thoracic aorta (CEC-pretreated). The contractile responses to PNO-49B were inhibited competitively by prazosin, HV723 (a-ethyl-3,4,5-trimethoxy-a-(3-((2-(2-methoxyphenoxy)-ethyl)-amino(propyl)benzeneacetonitrile fumarate) and by WB4101 (2-(2,6-dimethoxyphenoxyethyl)-aminomethyl-1,4-benzodioxane). The estimated pA ₂ values were high for prazosin and WB4101 in rat thoracic aorta and for HV723 in dog mesenteric artery, whereas the pA ₂ values for these three antagonists in rabbit thoracic aorta were low and were not altered by pretreatment with CEC. The binding of [ ³H]-prazosin to membranes prepared from rat vas deferens and liver was inhibited by PNO-49B in a concentration-dependent manner. The resulting pK ₁ value for the liver was approximately 1.5?log units lower (one thirtieth in affinity) than the values for the epididymal and prostatic portions of the vas deferens. PNO-49B also inhibited biphasically [ ³H]prazosin binding to prazosin-high affinity sites of rat cerebral cortex membranes, and the low but high affinity sites for PNO-49B was abolished by CEC-pretreatment. PNO-49B had no effect on the prejunctional a ₂-adrenoceptors in rat vas deferens (prostatic portion) nor on the ß-adrenoceptors in rat atria. The contractile response to PNO-49B in rat thoracic aorta was not inhibited by cimetidine, pyrilamine or ketanserin. These results indicate that PNO-49B is an a ₁-adrenoceptor agonist with a lower affinity and/or efficacy at the a _1B subtype as compared with other a ₁-subtypes.     

Subtypes of alpha 1-adrenoceptors involved in noradrenaline-induced contractions of rat thoracic aorta and dog carotid artery.

We tried to determine alpha 1-adrenoceptor subtypes involved in noradrenaline-induced contractions of rat thoracic aorta and dog carotid artery. Prazosin competitively antagonized the contractions induced by noradrenaline in both the arteries with a high pKB value (approximately 9.7). WB4101, benoxathian, phentolamine, HV723 and 5-methylurapidil also competitively antagonized the responses to noradrenaline in both the arteries: however, the affinities for the antagonists were significantly higher in the rat thoracic aorta than in the dog carotid artery. The affinities for the competitive antagonists were not changed by treatment with nifedipine. In the rat thoracic aorta, chlorethylclonidine (CEC) elicited either a persistent contraction with rhythmic activities before treatment with nifedipine or partial inactivation of alpha 1-adrenoceptors in the presence of nifedipine. On the other hand, CEC produced only inactivation of alpha 1-adrenoceptors in the dog carotid artery. These results suggest that noradrenaline-induced contractions of the rat thoracic aorta and dog carotid artery are respectively mediated through distinct alpha 1-adrenoceptor subtypes. According to the alpha 1A, alpha 1B subclassification, the alpha 1-adrenoceptor of dog carotid artery is like the alpha 1B subtype, while that of rat thoracic aorta is atypical. Both subtypes are also identified as a high affinity site for prazosin (alpha 1H subtype) in the alpha 1H, alpha 1L and alpha 1N subclassification.     

Investigation of the subtypes of alpha 1-adrenoceptor mediating contractions of rat aorta, vas deferens and spleen.

1. The subtypes of alpha 1-adrenoceptor mediating contractions to exogenous noradrenaline (NA) or phenylephrine in rat vas deferens, spleen and aorta, and mediating contractions to endogenous NA in rat vas deferens have been examined. 2. In rat vas deferens, the competitive antagonists prazosin, WB 4101, benoxathian and 5-methyl-urapidil inhibited contractions to NA with pA2 values of 9.26, 9.54, 9.02 and 8.43, respectively. The irreversible antagonist chloroethylclonidine (CEC) (100 microM) failed to affect contractions to NA. 3. In rat vas deferens in the presence of nifedipine (10 microM), contractions to NA were significantly attenuated and under these conditions, CEC (100 microM) significantly reduced the maximum response to NA. 4. In rat spleen, the competitive antagonists prazosin, WB 4101 and benoxathian inhibited contractions to phenylephrine with pA2 values of 9.56, 8.85 and 7.60, respectively, and 5-methyl-urapidil had a KB of 6.62. CEC (100 microM) significantly reduced the maximum contraction to phenylephrine. 5. In rat aorta, the competitive antagonists, prazosin, WB 4101, benoxathian and 5-methyl-urapidil inhibited contractions to NA with pA2 values of 9.45, 9.21, 8.55 and 8.12, respectively. CEC (100 microM) produced an approximately parallel shift in the potency of NA, without significantly reducing the maximum response. 6. In epididymal portions of rat vas deferens in the presence of nifedipine (10 microM), the isometric contraction to a single electrical pulse was significantly reduced by CEC (100 microM), and by the competitive antagonists prazosin, WB 4101, benoxathian and 5-methyl-urapidil at concentrations of 1 nM.(ABSTRACT TRUNCATED AT 250 WORDS)     

Three distinct binding sites for [3H]-prazosin in the rat cerebral cortex.

1. The putative alpha 1-adrenoceptor subtypes of rat cerebral cortex membranes were characterized in binding. 2. Specific binding of [3H]-prazosin was saturable between 20-5000 pm. Scatchard plots of the binding data were non-linear, indicating the presence of two distinct affinity sites for prazosin (pKD, high = 10.18, Rhigh = 308 fmol mg-1 protein; pKD, low = 8.96, Rlow = 221 fmol mg-1 protein). 3. In the membranes pretreated with chlorethylclonidine (CEC) two affinity sites for prazosin were also observed: the affinities were similar to those without CEC pretreatment, but the maximum numbers of binding sites were reduced by CEC pretreatment to 23 and 62% for prazosin-high (Rhigh) and low affinity sites (Rlow), respectively. 4. The prazosin-high affinity sites were further subdivided into two subclasses by WB4101(2-(2,6-dimethoxyphenoxyethyl)aminomethyl-1,4-benzodioxane) and phentolamine; the low affinity sites for WB4101 and phentolamine were more potently inactivated by CEC as compared with the high affinity sites. On the other hand, prazosin, HV723 (alpha-ethyl-3,4,5-trimethoxy-alpha-(3-((2-(2-methoxyphenoxy)ethyl)- amino )-propyl)benzeneacetonitrile fumarate) and yohimbine inhibited [3H]-prazosin binding to prazosin-high affinity sites monophasically. 5. In addition to the high affinity sites, the prazosin-low affinity sites were labelled at high concentrations of [3H]-prazosin. Thus, prazosin and WB4101 showed shallow displacement curves. On the other hand, HV723 and yohimbine did not discriminate between prazosin-high and low affinity sites. 6. Two distinct alpha 1-adrenoceptor subclassifications have been recently proposed (alpha 1A, alpha 1B subtypes and alpha 1H, alpha 1L, alpha 1N subtypes).(ABSTRACT TRUNCATED AT 250 WORDS)     

Studies on RX 781094: a selective, potent and specific antagonist of alpha 2-adrenoceptors.

1 The selectivity and specificity of RX 781094 [2-(2-(1,4 benzodioxanyl))2-imidazoline HCl] for alpha-adrenoceptors have been examined in peripheral tissues. 2 In isolated tissue experiments RX 781094 was a competitive antagonist at prejunctional alpha 2-adrenoceptors situated on the sympathetic nerve terminals of the rat (pA2 = 8.56) and mouse (pA2 = 7.93) vas deferens and on the parasympathetic nerve terminals of the guinea-pig ileum (pA2 = 8.55). 3 Although RX 781094 was also a competitive antagonist at the postjunctional alpha 1-adrenoceptors of the rat anococcygeus muscle (pA2 = 6.10) its affinity for these receptors was markedly less than that displayed for prejunctional sites. From pA2 values obtained in the rat vas deferens and anococcygeus muscle the calculated alpha 2/alpha 1-adrenoceptor selectivity ratio for RX 781094 was 288. 4 The rank order of alpha 2/alpha 1-adrenoceptor selectivities for the antagonists studied was RX 781094 greater than RS 21361 greater than yohimbine greater than piperoxan greater than phentolamine greater than WB 4101 greater than prazosin. 5 RX 781094 had extremely low affinity for beta-adrenoceptors, histamine receptors, cholinoceptors, 5-hydroxytryptamine and opiate receptors in vitro. 6 In pithed rats, intravenous administration of RX 781094 antagonized the prejunctional alpha 2-adrenoceptor agonist effects of clonidine and guanabenz on electrically-induced contractions of the vas deferens and anococcygeus muscle respectively. 7 In the vas deferens the rank order of alpha 2-adrenoceptor antagonist potencies was RX 781094 greater than phentolamine greater than piperoxan greater than yohimbine greater than RS 21361 greater than WB 4101. Only RX 781094, yohimbine and RS 21361 were active against guanabenz in the anococcygeus muscle. 8 In the pithed rat, RX 781094 preferentially antagonized the pressor responses evoked by postjunctional alpha 2-adrenoceptor activation by UK 14,304 although higher doses also inhibited the effects of phenylephrine and cirazoline at postjunctional alpha 1-adrenoceptors. 9 RX 781094 had little effect on the cardiovascular responses to 5-hydroxytryptramine, angiotensin II, histamine, acetylcholine and isoprenaline in pithed rats and rats anaesthetized with pentobarbitone. 10 These results demonstrate that RX 781094 is a potent and selective alpha 2-adrenoceptor antagonist with a high degree of specificity for these receptors.     

Characterization of alpha1-adrenoceptor-mediated contraction in the mouse thoracic aorta

In the mouse thoracic aorta, noradrenaline, adrenaline, phenylephrine and methoxamine behaved as full agonists. The pA(2) values for 8-[2-[4-(2-methoxyphenyl)-1-piperazinyl]ethyl]-8-azaspiro[4,5]decane-7,9-dione dihydrochloride (BMY 7378) against each agonist were in good agreement with the generally accepted affinity value of alpha(1D)-adrenoceptors. 5-Methylurapidil, 2-[2,6-dimethoxyphenoxyethyl]aminomethyl-1,4-benzodioxane hydrochloride (WB 4101) and prazosin inhibited the contraction in response to noradrenaline. A significant correlation was obtained between the antagonist affinities in mouse thoracic aorta and those of native alpha(1D)-adrenoceptors in rat thoracic aorta or with those of cloned alpha(1d)-adrenoceptors, but not with those for either alpha(1a)- or alpha(1b)-adrenoceptors. Buspirone behaved as a partial agonist in mouse thoracic aorta, the contraction of which was antagonized by BMY 7378 with a pA(2) value (8.49) consistent with that found against noradrenaline (8.43). Clonidine acted as a partial agonist (pD(2)=5.94). The pK(p) value for clonidine against noradrenaline was similar to the pD(2) value for clonidine. The apparent pK(B) value for BMY 7378 against clonidine was similar to the pA(2) value against other full agonists used in the present study. These results suggest that the alpha(1D)-adrenoceptor subtype exists, and that the full agonists and the partial agonists evoke the contraction mediated through the alpha(1D)-adrenoceptor in mouse thoracic aorta.     

Does segmental difference in alpha 1-adrenoceptor subtype explain contractile difference in rat abdominal and thoracic aortae?

The cyclooxygenase inhibitor, indomethacin, depresses adrenergic agonist constriction of endothelium-denuded rat abdominal, but not thoracic, aorta. In order to explain this finding, we explored the possibility of segmental differences in the population of alpha 1-adrenoceptor (AR) subtypes. In endothelium-denuded tissues, phenylephrine elicited concentration-dependent contractions in the thoracic and abdominal aortic rings with potencies and maximal effects that, respectively, did not differ significantly (P > .05). Indomethacin (1 x 10(-5) M) inhibited phenylephrine-induced contractions only in abdominal aorta. The subtype-selective alpha 1D-AR antagonist, BMY 7378, was found to antagonize contractions to phenylephrine competitively in abdominal (pA2 8.44) and thoracic (pA2 8.56) aortic rings. These data are consistent with published alpha 1D-AR functional potency and clonal alpha 1D-AR binding affinity. In addition, cumulative concentration-contraction curves for phenylephrine were competitively antagonized in the rat abdominal and thoracic aortae by prazosin, 5-methylurapidil and WB 4101, with pA2 values of 9.39 and 9.61, 7.64 and 7.85, and 9.43 and 9.58, respectively. These compounds with varying degrees of subtype selectivity inhibited contractions of the thoracic and abdominal aortae with affinities consistent with those determined at the alpha 1D-AR subtype. The results of this study suggest that the contraction to phenylephrine of the rat abdominal and thoracic aorta is mediated via the same alpha 1D-AR subtype.     

Pharmacological characterization of alpha 1-adrenoceptor subtypes in rat heart: a binding study.

1. The alpha 1-adrenoceptor subtypes of rat heart were characterized in binding experiments performed with [3H]-prazosin as the radiolabel. The specific binding to the alpha 1-adrenoceptors was determined with 0.3 microM prazosin, because phentolamine (10 microM) was insufficient to inhibit completely the specific binding of high concentrations of [3H]-prazosin. 2. In saturation experiments, [3H]-prazosin bound to two distinct affinity sites (pKD = 10.39 and 8.19). The proportion of the low affinity sites was approximately 84% of total specific binding. Membranes pretreated with chloroethylclonidine (CEC, 10 microM) also showed two distinct affinity sites for [3H]-prazosin, although the maximum numbers of high and low affinity sites were reduced by 86 and 64%, respectively. 3. In competition experiments, [3H]-prazosin (100 pM) binding was inhibited by WB4101 (2-(2,6-dimethoxy-phenoxyethyl)aminomethyl-1,4-benzodioxane) and 5-methylurapidil. The inhibition curves displayed shallow slopes which could be subdivided into high and low affinity components (pKi = 10.43 and 8.36 for WB4101, 8.62 and 6.61 for 5-methylurapidil). However, unlabelled prazosin or HV723 (alpha-ethyl-3,4,5-trimethoxy-alpha-(3-((2-(2-methoxyphenoxy)-ethyl)amin o) propyl)benzeneacetonitrile fumarate) competed for [3H]-prazosin binding monophasically (pKi = 10.34 and 8.28, respectively). In CEC-pretreated membranes, prazosin, WB4101, 5-methylurapidil and HV723 antagonized the [3H]-prazosin (100 pM) binding monophasically (pKi = 9.70, 9.56, 8.60 and 8.82, for each antagonist). 4. On the other hand, 1000 pM [3H]-prazosin binding was inhibited by unlabelled prazosin biphasically (pKi = 10.49 and 8.49). HV723 did not discriminate both prazosin-high and low affinity sites (pKi = 8.18).(ABSTRACT TRUNCATED AT 250 WORDS)     

The modulatory role of vascular endothelium in the interaction of agonists and antagonists with alpha-adrenoceptors in the rat aorta.

1. We have examined the effect of endothelium on the antagonistic action of prazosin, doxazosin, yohimbine and phentolamine against phenylephrine, clonidine and noradrenaline. 2. The action of prazosin against phenylephrine was similar to that earlier reported against noradrenaline, acting as a non-competitive antagonist in the presence of endothelium and as a competitive antagonist in the absence of endothelium. Prazosin also acted as a non-competitive antagonist against clonidine in the absence of endothelium. 3. Doxazosin behaved in a similar way to prazosin against noradrenaline, phenylephrine and clonidine acting as a non-competitive antagonist in the presence of endothelium and as competitive antagonist after removal of endothelium. In contrast, yohimbine and phentolamine acted as competitive antagonists both in the presence and in the absence of endothelium. 4. Analysis of the concentration-response curves for noradrenaline, phenylephrine and clonidine in the presence and in the absence of endothelium showed that the affinity for all three agonists was the same but not the efficacy and the receptor reserve, both of which were lower in the presence than in the absence of endothelium. 5. The rank order of agonist potency in the absence of endothelium was noradrenaline greater than phenylephrine greater than clonidine. The rank order of antagonist potency was prazosin greater than or equal to doxazosin greater than phentolamine greater than yohimbine. 6. The results show that vascular endothelium modulates the contractile response to alpha-adrenoceptor agonists and also modifies the action of the antagonists prazosin and doxazosin but not that of yohimbine and phentolamine. This effect of endothelium was related to a change in agonist efficacy and receptor reserve. These results also suggest that the alpha-adrenoceptors of the isolated aorta of the rat are predominantly, if not exclusively of the alpha 1-subtype.     

alpha(1)-Adrenoceptor subtypes in the mouse mesenteric artery and abdominal aorta.

1. Subtypes of alpha(1)-adrenoceptor-mediated contractions to noradrenaline in mouse mesenteric artery and abdominal aorta were examined. 2. In mesenteric artery, BMY7378, 5-methylurapidil, WB4101 and prazosin were inhibited contraction to noradrenaline The good correlation for pA(2) values of antagonists in native alpha(1D)- (rat thoracic aorta) adrenoceptor subtype and pK(i) values in rat cloned alpha(1d)-adrenoceptor with the pA(2) values estimated in the mouse mesenteric artery was obtained. However, the pA(2) value for BMY7378 is significantly lower than the accepted value against the alpha(1D)-adrenoceptor subtype. 3. In the abdominal aorta, it was obtained the regional difference for the sensitivity for noradrenaline. 4. In the upper abdominal aorta, the good correlation for the pA(2) values of the antagonists in the native alpha(1D)-adrenoceptor subtype and pK(i) values in the cloned alpha(1d)-adrenoceptor with the pA(2) values estimated in the upper abdominal aorta was obtained, and regression line was close to the line of identity. 5. In the lower abdominal aorta, the good correlation for the reported pK(i) values in the cloned alpha(1a)-adrenoceptor subtype with the pA(2) values estimated in the mouse lower abdominal aorta was obtained, and regression line was close to the line of identity. 6. In conclusion, the present functional data in the mouse suggest that (1) alpha(1D)-like adrenoceptors are present in the mesenteric artery, (2) there is the regional difference for the sensitivity for noradrenaline in the abdominal aorta and (3) noradrenaline evokes the contraction mediated through alpha(1D)-adrenoceptor in the upper abdominal aorta, whereas there is alpha(1A)-adrenoceptor-mediated contraction in the lower abdominal aorta.     

Relation between adrenergic neurogenic contraction and alpha 1-adrenoceptor subtypes in dog mesenteric and carotid arteries and rabbit carotid arteries.

1. We examined the distribution of alpha 1-adrenoceptor subtypes and their relation to adrenergic neurogenic contraction induced by electrical transmural stimulation in the dog mesenteric and carotid arteries and the rabbit carotid artery. 2. In the dog mesenteric artery, contraction to noradrenaline was competitively inhibited by HV723 (pKB = 9.37) and prazosin (pKB = 8.40). Pretreatment with chlorethylclonidine (CEC) slightly attenuated only the contractions induced by low concentrations of noradrenaline. Contraction induced by electrical transmural stimulation was inhibited at lower concentrations of HV723 than of prazosin. 3. In the dog carotid artery, contraction to noradrenaline was inhibited with higher affinity by prazosin (pKB = 9.82) than by HV723 (pKB = 8.47). Prazosin was also more potent than HV723 in inhibiting the contraction to electrical stimulation. Pretreatment with CEC markedly attenuated or abolished contraction to noradrenaline and electrical stimulation. 4. In the rabbit carotid artery, prazosin inhibited noradrenaline-induced contraction biphasically (pKB = 9.91 and 8.60). After CEC pretreatment, contraction to noradrenaline was attenuated moderately and the high affinity site for prazosin was abolished. HV723 competitively inhibited the noradrenaline response with a similar pKB value (approximately 8.5) regardless of CEC treatment. Contraction to electrical stimulation was inhibited by prazosin more effectively than by HV723 in preparations not treated with CEC, while it was equipotently inhibited by both antagonists in CEC-treated preparations.(ABSTRACT TRUNCATED AT 250 WORDS)     

Analysis of the effects of alpha 1-adrenoceptor antagonists on noradrenaline-mediated contraction of rat small mesenteric artery.

1. In this study, we examined the interaction between noradrenaline (NA) and phenylephrine (PE) with seven antagonists (prazosin, tamsulosin, phentolamine, WB-4101, 5-methylurapidil, spiperone and HV 723) in an attempt to characterize the alpha 1-adrenoceptor population of the rat isolated small mesenteric artery (SMA) preparation. 2. Six of the seven antagonists investigated produced concentration-dependent, parallel, rightward shift of the NA concentration-effect (E/[A]) curves. The exception was tamsulosin, which produced significant decrease of the upper asymptote. In the case of 5-methylurapidil and HV723, the Schild plot slope parameters were not significantly different from unity over the range of concentrations used. However, the Schild plot slopes obtained for the other antagonists were all significantly greater than unity, inconsistent with expectations for simple competitive antagonism. 3. HV723, prazosin and tamsulosin were also tested using PE as an agonist. All three antagonists produced concentration-dependent, parallel, rightward shifts of the PE curves and Schild analysis yielded slope parameters not significantly different from unity. The pKB estimates obtained for tamsulosin and prazosin were not significantly different from the pA2 values obtained when NA was used as agonist. In the case of HV723, the 95% confidence intervals for the pKB values yielded with NA and PE did not overlap (pKB = 8.80-9.13 and 8.15-8.77 for NA and PE, respectively). 4. In the absence of evidence to indicate that the steep Schild plots were due to failure to satisfy the basic criteria for quantitative analysis in a one-receptor system, we considered the possibility that the complexity was caused by an action of NA at inhibitory D1 receptors. The selective D1 receptor antagonists, SCH-23390 (10 nM), had no significant effect on the NA E/[A] control curve, but the apparent potency of 100 nM prazosin was reduced by approximately 3.5 fold. 5. This study indicates that the steep Schild plots obtained from the interaction between NA and alpha 1-adrenoceptor antagonists were due to the simultaneous activation of inhibitory D1 receptors by NA. Notwithstanding this complexity, our explanatory model of the system (see Appendix) suggests that the antagonist affinity values estimated in the absence of D1 receptor block were not significantly affected by this other action of NA. The low affinity estimate obtained for prazosin suggests that the pharmacologically-defined alpha IL-subtype operates in the SMA.     

Two distinct alpha 1-adrenoceptor subtypes involved in noradrenaline contraction of the rabbit thoracic aorta

1. Recently, alpha 1-adrenoceptors in blood vessels have been classified into three subtypes (alpha 1H, alpha 1L and alpha 1N). We examined which subtype (or subtypes) is involved in the noradrenaline-induced contraction of rabbit thoracic aorta. 2. Noradrenaline produced a concentration-dependent contraction in the rabbit isolated thoracic aorta. Prazosin antagonized the contractions to noradrenaline, resulting in a rightward displacement of the concentration-response curve. However, the shift was not proportional to the concentration of prazosin; Schild plots showed that the inhibition by prazosin was biphasic, implying that noradrenaline acted through two receptor populations. Two affinity constants (pKB values of 10.02 and 8.83) were determined for prazosin at these sites. 3. However, under continuous treatment with 1 nM prazosin, or in strips pretreated with chlorethylclonidine (CEC; an alpha 1H inactivating agent) to remove the contribution of one receptor population, prazosin showed a single pKB or pA2 value of approximately 8.3. 4. Yohimbine also produced biphasic antagonism of noradrenaline-induced contractions, resulting in two affinity constants (pKB = 6.52 and 6.17). However, a monophasic Schild plot was obtained for yohimbine either in the presence of 1 nM prazosin (pA2 = 6.08) or in strips pretreated with CEC (pA2 = 6.03). 5. The Schild plot for HV723 (a selective alpha 1N-antagonist) yielded a monophasic slope (pKB = 8.47) and the inhibition was not affected by 1 nM prazosin or CEC-pretreatment. 6. [3H]-prazosin bound to alpha 1-adrenoceptors of the aortic membrane preparations with two different affinities (pKD = 9.94 and 8.37).(ABSTRACT TRUNCATED AT 250 WORDS)     

Comparison of pre- and postsynaptic alpha-adrenoceptor blocking effects of E-643 in the isolated vas deferens of the rat

Effectiveness of E-643, a newly developed alpha-blocker, and four alpha-antagonists in blocking pre- and postsynaptic alpha-adrenoceptors were compared in the isolated rat vas deferens. The inhibitory effect of clonidine on the field-stimulated twitch response was antagonized in the presence of the alpha-antagonist. The order of affinity (pA2) for presynaptic alpha-receptors, as assessed from parallel shift of the dose-response curve to clonidine, was: phentolamine greater than yohimbine greater than tolazoline greater than E-643 greater than or equal to prazosin. At concentrations from 10(-8) to 10(-6) M, neither E-643 nor prazosin had any effect on the twitch which had been depressed by the treatment with clonidine, whereas phentolamine, yohimbine and tolazoline partially reversed it. Contractile effects of cumulative concentrations of noradrenaline were also antagonized by alpha-antagonists. The order of affinity (pA2) for postsynaptic alpha-receptors was: E-643 greater than or equal to prazosin greater than phentolamine greater than yohimbine greater than tolazoline. Selectivity for pre- versus postsynaptic alpha-receptors was assessed by comparing KB values for pre- and postsynaptic alpha-receptors. The order of selectivity for the presynaptic alpha-receptors was : yohimbine greater than tolazoline greater than phentolamine much greater than prazosin greater than or equal to E-643. It is concluded that E-643 is a potent and highly selective postsynaptic alpha-blocker.     

Pharmacological activity of (-)-discretamine, a novel vascular alpha-adrenoceptor and 5-hydroxytryptamine receptor antagonist, isolated from Fissistigma glaucescens.

1. The pharmacological activity of (-)-discretamine, isolated from Fissistigma glaucescens, was determined in rat isolated thoracic aorta, cardiac tissues and ventricular myocytes and guinea-pig isolated trachea. 2. (-)-Discretamine was found to be an alpha 1-adrenoceptor blocking agent in rat thoracic aorta as revealed by its competitive antagonism of noradrenaline (pA2 = 7.20 +/- 0.10)- or phenylephrine (pA2 = 7.60 +/- 0.09)-induced vasoconstriction. It was as potent as phentolamine (pA2 = 7.51 +/- 0.10), but was more potent than yohimbine (pA2 = 6.18 +/- 0.06). Removal of endothelium significantly increased the antagonistic potency of (-)-discretamine on noradrenaline (pA2 = 7.52 +/- 0.09)- or phenylephrine (pA2 = 7.90 +/- 0.09)-induced vasoconstriction. 3. (-)-Discretamine was also an alpha 2-adrenoceptor blocking agent (pA2 = 6.30 +/- 0.15) and a 5-hydroxytryptamine antagonist (pA2 = 6.87 +/- 0.06), both in rat aorta denuded of endothelium. 4. (-)-Discretamine protected alpha-adrenoceptors from alkylation by the irreversible blocking agent, phenoxybenzamine. 5. [3H]-inositol monophosphate formation caused by noradrenaline (3 microM) in rat thoracic aorta was suppressed by (-)-discretamine (10 and 30 microM) and prazosin (3 microM). 6. A high concentration of (-)-discretamine (30 microM) did not affect the contraction induced by the thromboxane receptor agonist U-46619, prostaglandin F2 alpha (PGF2 alpha), angiotensin II, high K+ or endothelin in rat aorta denuded of endothelium. Neither cyclic AMP nor cyclic GMP content of rat thoracic aorta was changed by (-)-discretamine.(ABSTRACT TRUNCATED AT 250 WORDS)     

1-Adrenoceptor subtype involved in the positive inotropic response to phenylephrine in rat atria

Phenylephrine produced positive inotropic responses in isolated rat right and left atria. The responses were competitively inhibited by ₁-adrenoceptor antagonists (prazosin, WB4101 and HV723) with relatively low affinities (pA₂ values close to 8.0). Chloroethylclonidine had no significant effect on the responses to phenylephrine. These results suggest that the positive inotropic response to phenylephrine in rat atria is mediated through ₁-adrenoceptors which cannot be defined by the _1A, _1B subclassification.     

Studies on YM-12617: A selective and potent antagonist of postsynaptic α1-adrenoceptors

YM-12617, 5-[2-[[2-(2-ethoxyphenoxy)ethyl]-amino]propyl]-2 -methoxybenzenesulfonamide HCl is a structurally new type of extremely potent alpha 1-adrenoceptor antagonist. Its alpha-adrenoceptor blocking properties have been compared with those of prazosin, phentolamine and yohimbine using both pharmacological and 3H-ligand binding techniques in vitro and in vivo. In the isolated rabbit aorta, a tissue known to contain mainly alpha 1-adrenoceptors at postjunctional sites, YM-12617 competitively antagonized noradrenaline-induced contraction with a pA2 value of 10.11. Although YM-12617 was also a competitive antagonist toward clonidine at prejunctional alpha 2-adrenoceptors in the isolated rat vas deferens, its affinity for these receptors (pA2 = 6.41) was 5,000 times lower than that displayed for the postjunctional alpha 1-adrenoceptors in the isolated rabbit aorta. YM-12617 displaced both 3H-WB 4101 and 3H-clonidine binding to rat brain membranes; however, the affinity of YM-12617 for alpha 1-adrenoceptors (pKi = 9.64) was 3800 times higher than that for alpha 2-adrenoceptors (pKi = 6.06). Based on pA2 values obtained in the isolated tissues and pKi values in the binding assays, YM-12617 was 2-18, 36-117 and 1,740-5,750 times more potent than prazosin, phentolamine and yohimbine in antagonizing alpha 1-adrenoceptors, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)     

Different alpha-adrenoceptors modulate the release of 5-hydroxytryptamine and noradrenaline in rat cortex.

1 The potassium-evoked release of [3H]-noradrenaline from slices of rat occipital cortex and the potassium-evoked release of [3H]-5-hydroxytryptamine from slices of rat frontal cortex were measured using a superfusion system. 2 The rank order of potency for a number of alpha-adrenoceptor agonists was different for the two neuronal systems, clonidine and azepexole being the most potent inhibitors of noradrenaline release and methoxamine and phenylephrine being the most potent against 5-hydroxytryptamine release. 3 The rank order of potency for a series of alpha-adrenoceptor antagonists in reversing the inhibition of noradrenaline release produced by clonidine was: phentolamine greater than rauwolscine = yohimbine = corynanthine much greater than WB4101, whereas against methoxamine-inhibition of 5-hydroxytryptamine release the rank order of potency was: WB4101 greater than phentolamine greater than corynanthine greater than yohimbine greater than rauwolscine. 4 The results suggest that the alpha-adrenoceptors which modulate potassium-evoked 5-hydroxytryptamine release are not identical with the alpha 2-adrenoceptors which modulate potassium-evoked 5-hydroxytryptamine release are not identical with the alpha 2-adrenoceptors located on noradrenergic nerve terminals and may more closely resemble alpha 1-than alpha 2-adrenoceptors.