Differential effect of tetrandrine on aortic relaxation and chronotropic activity in rat isolated aorta and atria

Investigation of the pathogenesis of arteriosclerosis and/or atherosclerosis has been progressed using molecular biology. New concepts have been developed and, receptors and substances have been found clinically and experimentally, which have led us to create new methods of evaluating or diagnosing the grade of atherosclerosis lesion. Dealing with the new concepts or knowledge in this symposium, this introductory paper describes an overview of pathogenesis of atherosclerosis, from which the new methods of evaluating or diagnosing lesion has been exploited. The injury to the endothelium leads to endothelial cell dysfunction, which initiates the acceleration of LDL oxidation and increases adherence of monocytes, macrophages and T lymphocytes, migrating subendothelially and causing large foam cells to develop because of lipid accumulation. Macrophages and platelets release many growth factors, which accelerate the growth of vascular smooth muscle cells, forming fibrous plaque. In these pathogenic processes of atherosclerosis, angiotensin II participates in releasing growth factor for cell proliferation and hepatocyte growth factor (HGF) participates in revascularization of the sclerotic lesion, suggesting a candidate marker for atherosclerosis. Hyperlipidemia and hypercoagulation are the major factors in advanced atherosclerosis. Using new methods to evaluate or diagnose lesions, further therapy and prevention for atherosclerosis will progress in future.     

beta-Glucosidase activity in the rat small intestine toward quercetin monoglucosides

In order to evaluate the positional specificity for a glucoside group in the hydrolysis of flavonoid glucosides in the rat small intestine, beta-glucosidase activity was measured with the quercetin monoglucosides, quercetin-3-O-beta-D-glucopyranoside (Q3G), quercetin-4'-O-beta-D-glucopyranoside (Q4'G) and quercetin-7-O-beta-D-glucopyranoside (Q7G), as well as with quercetin-3-O-rutinoside (rutin) and p-nitrophenyl-beta-D-glucopyranoside (NPG) by using the HPLC technique. Enzymes were prepared from rat small intestinal mucosa of the duodenum, jejunum and ileum, among which the enzyme activity of the jejunum was highest for all the glycosides tested. Q4'G was the richest substrate for a beta-glucosidase solution among these glycosides, while rutin and NPG were both poor substrates. This suggests that dietary flavonoid glucosides are primarily hydrolyzed and liberated aglycones in the jejunum.     

Activation of multiple sites by adenosine analogues in the rat isolated aorta

1. The presence of A2 receptors mediating relaxation in the rat isolated aorta has been previously demonstrated. However, agonist dependency of the degree of rightward shift elicited by 8-sulphophenyltheophylline (8-SPT) led to the suggestion that the population of receptors in this tissue is not a homogeneous one. In this study we have re-examined the effects of 8-SPT in the absence and presence of the NO synthase inhibitor L-NAME (NG-nitro-L-arginine methyl ester) and investigated antagonism of responses by the potent A2a receptor ligands PD 115,199 (N-[2-dimethylamino)ethyl]-N-methyl-4-(2,3,6,7-tetrahydro-2,6-dioxo-1,3 dipropyl-1H-purin-8-yl)) benzene sulphonamidexanthine), ZM 241385 (4-(2-[7-amino-2-(2-furyl) [1,2,4]-triazolo[2,3-a][1,3,5]triazin-5-yl amino]ethyl)phenol), and CGS 21680 (2-[p-(2-carboxyethyl)phenylamino]-5'-N-ethylcarboxamidoadenosine). We have also investigated the antagonist effects of BWA1433 (1,3-dipropyl-8-(4-acrylate)phenylxanthine) which has been shown to have affinity at rat A3 receptors. 2. Adenosine, R-PIA (N6-R-phenylisopropyl adenosine), CPA (N6-cyclopentyladenosine) and NECA (5'-N-ethylcarboxamidoadenosine) all elicited relaxant responses in the phenylephrine pre-contracted rat isolated aorta with the following potency order (p[A50] values in parentheses): NECA (7.07 +/- 0.11) > R-PIA (5.65 +/- 0.10) > CPA (5.05 +/- 0.12) > adenosine (4.44 +/- 0.12). 3. 8-SPT (10-100 microM) caused parallel rightward shifts of the E/[A] curves to NECA (pKB = 5.23 +/- 0.16). A smaller rightward shift of E/[A] curves to CPA was observed (pA2 = 4.85 +/- 0.17). However, no significant shifts of E/[A] curves to either adenosine or R-PIA were observed. 4. In the absence of endothelium E/[A] curves to NECA and CPA were right-shifted compared to controls. However, removal of the endothelium did not produce a substantial shift of adenosine E/[A] curves, and E/[A] curves to R-PIA were unaffected by removal of the endothelium. 5. In the presence of L-NAME (100 microM) E/[A] curves to NECA and CPA were right-shifted. However, no further shift of the CPA E/[A] curve was obtained when 8-SPT (50 microM) was administered concomitantly. The locations of curves to R-PIA and adenosine were unaffected by L-NAME (100 microM). 6. In the presence of PD 115,199 (0.1 microM) a parallel rightward shift of NECA E/[A] curves was observed (pA2 = 7.50 +/- 0.19). PD 115,199 (0.1 and 1 microM) gave smaller rightward shifts of E/[A] curves to R-PIA and CPA, but E/[A] curves to adenosine were not significantly shifted in the presence of PD 115,199 (0.1 or 1 microM). 7. The presence of ZM 241385 (3 nM-0.3 microM) caused parallel rightwad shifts of NECA E/[A] curves (pKB = 8.73 +/- 0.11). No significant shifts of E/[A] curves to adenosine, CPA or R-PIA were observed in the presence of 0.1 microM ZM 241385. 8. CGS 21680 (1 microM) elicited a relaxant response equivalent to approximately 40% of the NECA maximum response. In the presence of this concentration of CGS 21680, E/[A] curves to NECA were right-shifted in excess of 2-log units, whereas E/[A] curves to R-PIA were not significantly shifted. 9. BWA1433 (100 microM) caused a small but significant right-shift of the E/[A] curve to R-PIA yielding a pA2 estimate of 4.1 IB-MECA (N6-(3-iodo-benzyl)adenosine-5(1)-N-methyl uronamide) elicited relaxant responses which were resistant to blockade by 8-SPT (p[A]50 = 5.26 +/- 0.13). 10. The results suggest that whereas relaxations to NECA (10 nM-1 microM) are mediated via adenosine A2a receptors, which are located at least in part on the endothelium, R-PIA and CPA may activate A2b receptors on the endothelium and an additional, as yet undefined site, which is likely to be located on the smooth muscle and which is not susceptible to blockade by 8-SPT, PD 115,199 or ZM 241385. This site is unlikely to be an A3 receptor since the very small shift obtained in the presence of BWA1433 (100 microM), and the low potency of IB-MECA is not consistent with the affin     

Comparison of the attenuating effects of four beta-adrenoceptor agonists on rat isolated uterus and aorta

1. The ability of four beta-adrenoceptor agonists to attenuate oxytocin (0.2, 2 and 20 nmol/L) or KCl (20, 40 and 80 mmol/L)-induced contractions of the uterus (n = 5-8 for each agonist) and the KCl (18 mmol/L)-induced contractions of the aorta (n = 9 for each agonist) from rats, pretreated with oestradiol has been compared. 2. Isoprenaline, salbutamol, terbutaline and procaterol (0.1-10 mumol/L) attenuated the contractions of the uterus and the aorta. All four agonists had similar attenuating potencies on the uterus. 3. Procaterol caused the same maximal attenuation (33%) on the aorta as the other beta-adrenoceptor agonists and is thus acting as a full beta 2-adrenoceptor agonist under these experimental conditions. Isoprenaline and procaterol were much more potent than salbutamol and terbutaline in attenuating the aorta responses. 4. This study showed that isoprenaline and procaterol were potent attenuants on both the uterus and aorta whereas salbutamol and terbutaline were potent uterine but only modest aorta attenuants. This preliminary study indicates that the responsiveness of uterine and vascular tissue to certain beta 2-adrenoceptors differs.     

Identification of elastin peptides with vasorelaxant activity on rat thoracic aorta

Different Epstein-Barr-virus(EBV) variants were found to be associated with nasopharyngeal carcinoma (NPC). The type-C variant lacks the BamHI site between the BamHI W1* and I* regions and the type-f variant has an extra BamHI site in the BamHI F fragment. The BNLF1 gene (which encodes the LMP1 protein) from a nude-mouse-passaged CAO strain and from NPC biopsies from Taiwanese patients also exhibits variations resulting in structural and functional differences in the protein. The BZLF1 gene encodes the ZEBRA protein which triggers the EBV lytic cycle. A difference has been observed in 8 amino acids in the ZEBRA sequence in B95-8 (Z95) and P3HR1 (ZP3) cell lines. EBV found in NPC biopsies and peripheral-blood cells from Asians was predominantly of the ZP3 type (72%), while 81% of samples from different EBV-associated diseases and peripheral-blood cells from North Africa or Europe were of the Z95 type. We found that an alanine 206 had been replaced by a serine in the Z95 sequence in 72% of the NPC biopsies from European and North African patients. The Zser206 variant is found in a significantly lower percentage (p < 0.001) of other EBV-positive tissues from individuals in the same region (10-33%). In contrast, a 30-bp deletion is observed near the 3' end of the LMP1 gene in the majority of EBV (86%) from NPC and peripheral-blood cells from Asians, whereas a significantly lower percentage (p < 0.001) of NPC biopsies from European and North African patients (56%) have this deletion, as do lymphocytes from control individuals from the same region (36 and 55% respectively).     

Hydrogen peroxide-induced impairment of reactivity in rat isolated aorta: potentiation by 3-amino-1,2,4-triazole

1. In this study the impairment induced by hydrogen peroxide of vascular reactivity and the role of endogenous catalase in protection against this impairment was assessed in isolated rings of rat aorta. 2. Incubation with hydrogen peroxide at 1 mM, but not at 0.1 mM, for 15, 30 or 60 min followed by washout depressed, in a time-dependent manner, the subsequent ability of endothelium-containing and endothelium-denuded rings to contract to phenylephrine. 3. Incubation with 3-amino-1,2,4-triazole (50 mM, 90 min, followed by washout) to inhibit endogenous catalase had no effect by itself on subsequent phenylephrine-induced contraction. However, pretreatment with 3-amino-1,2,4-triazole did lead to a profound enhancement of the ability of hydrogen peroxide (1 mM, present for the final 30 min of the 90 min incubation, followed by washout) to depress phenylephrine-induced contraction in both endothelium-containing and endothelium-denuded rings. 4. Incubation with hydrogen peroxide at 1 mM, but not at 0.1 mM, for 15, 30 or 60 min followed by washout inhibited, in a time-dependent manner, the subsequent ability of acetylcholine (10 nM-3 microM) to induce endothelium-dependent relaxation. Furthermore, incubation with hydrogen peroxide 1 mM (30 min, followed by washout) also inhibited relaxation induced by glyceryl trinitrate (1-100 nM) or isoprenaline (10 nM-3 microM) in endothelium-denuded rings. 5. Incubation with 3-amino-1,2,4-triazole (50 mM, 90 min, followed by washout) had no effect by itself on relaxation induced by acetylcholine, glyceryl trinitrate or isoprenaline. In contrast, pretreatment with 3-amino-1,2,4-triazole led to profound enhancement of the ability of hydrogen peroxide (1 mM, present for final 30 min of the 90 min incubation) to block relaxation to acetylcholine, glyceryl trinitrate or isoprenaline. 6. On the basis of the actions of 3-amino-1,2,4-triazole, it is likely that endogenous catalase plays an important role in the protection of vascular reactivity of rat aorta against oxidant damage by high (1 mM) but not lower (0.1 mM) concentrations of hydrogen peroxide. The data are consistent with the promotion of non-selective damage to the vascular smooth muscle cells by hydrogen peroxide, but endothelial damage may also be sustained.     

Drug induced contraction and relaxation in mouse isolated aorta

Possibility of using mouse isolated aorta to evaluate the effect of vasoactive agents was demonstrated. The results suggested that aortic contraction induced by the alpha adrenoceptor agonist norepinephrine was sensitive to prazosin, and the contraction induced by the membrane depolarization agent KCl was sensitive to verapamil. Clonidine acting as a partial agonist attenuated the norepinephrine or methoxamine induced contraction. Both IBMX and nitroprusside relaxed the aortic contraction. These vascular changes induced by the above vasoactive agents in mouse isolated aorta were similar to those of rat described elsewhere. The present findings suggested that mouse isolated aorta can be used as a tool to test the effect of vasoactive agents.     

Effects of vitamin E deficiency on vasomotor activity and ultrastructural organisation of rat thoracic aorta

1. The effects of vitamin E deficiency were evaluated in aortic rings isolated from rats maintained on a diet deficient in vitamin E. 2. Endothelium-dependent vasodilator responses to acetylcholine (ACh) and calcium ionophore, A23187, were reduced in preparations from treated animals, compared to the age-matched controls. The maximal vasodilation to ACh was 66.4 +/- 9 (n = 4) and 38.8 +/- 7 (n = 4) % in control and 10 month-treated preparations, respectively. 3. The endothelium-independent vasodilator responses to sodium nitroprusside as well as the concentration-dependent contractile responses to noradrenaline, did not differ between treated and control preparations. 4. Electron microscopic examination of vascular segments and revealed that, following vitamin E deficiency, normal tissue organisation was disrupted, the endothelial monolayer either not being in contact with the underlying tissue or being absent in most of the areas analysed. 5. It is concluded that during vitamin E deficiency both morphological disruption and functional impairment of endothelium occur without observable modification of muscle cell function and morphology.     

Stereoselective differences in the vasorelaxing effects of S(+) and R(-) ketamine on rat isolated aorta

BACKGROUND: S(+) ketamine, because of its higher anesthetic potency and lower risk of psychotomimetic reactions, has been suggested to be superior to presently available racemic ketamine. The racemate is a direct vasodilator in vivo, and thus the authors investigated the vasorelaxing effects of ketamine enantiomers on rat aorta. METHODS: Rat isolated aortic rings with and without endothelium were contracted with 3 x 10(-7) M norepinephrine. Then 10(-5) to 3 x 10(-3) M S(+), R(-), or racemic ketamine were added cumulatively. Vascular responses to ketamine were further studied in rings pretreated with the nitric oxide synthase inhibitor N(omega)-nitro-L-arginine (NNLA), the adenosine triphosphate-sensitive K+ channel antagonist glibenclamide, and the L-type calcium channel blocking agent D888. RESULTS: Ketamine enantiomers and the racemate produced concentration-dependent vasorelaxation. The relaxing effect of S(+) ketamine was significantly weaker compared with R(-) ketamine and the racemate reflected by the half-maximum effective concentration (EC50) values of 11.6 x 10(-4), 4.8 x 10(-4), and 6 x 10(-4) M, respectively. Removal of the endothelium and NNLA or glibenclamide pretreatment did not significantly alter the vasorelaxing effect of ketamine. In contrast, D888 pretreatment significantly shifted the concentration-effect curves of both S(+) and R(-) ketamine rightward (EC50 values of 18.9 x 10(-4) and 8.5 x 10(-4) M, respectively), whereas the difference between the isomers was not affected. CONCLUSIONS: Vasorelaxation by ketamine enantiomers is quantitatively stereoselective: The effect of S(+)ketamine is significantly weaker compared with that of the racemate and R(-) ketamine. This stereoselective difference is not due to nitric oxide release, activation of adenosine triphosphate-sensitive potassium channels, or differential inhibition of L-type calcium channels.     

Inhibition of alpha1-adrenoceptor-mediated contractions in isolated tail arteries and aorta of the rat by alpha2-adrenoceptor agonists

The effects of alpha2-adrenoceptor agonists, clonidine, tizanidine and UK-14304 on alpha1-adrenoceptor-mediated contractile responses were studied in isolated tail arteries and thoracic aorta of the rat. When applied during sustained contractile responses to almost maximum concentration (10 microM) of phenylephrine, clonidine (0.3 microM to 100 microM) produced concentration-dependent relaxations in both tissues. The maximum relaxation was smaller in tail arteries than in thoracic aorta. Clonidine up to 100 microM failed to relax both tissues precontracted with KCl (60 microM) or U-46619 (1 microM), a thromboxane mimetic. The clonidine-induced relaxation in tail arteries, was reversed by alpha2-adrenoceptor antagonists, yohimbine and idazoxane. Effects of the alpha2-adrenoceptor antagonists were concentration-dependent (0.1 microM to 1 microM), but not in a competitive manner. On the other hand, the relaxation in thoracic aorta was not significantly antagonized by these alpha2-adrenoceptor antagonists. Tizanidine and UK-14304 also relaxed both tail arteries and thoracic aorta precontracted with phenylephrine. The characteristics of the relaxation and their antagonism by yohimbine in both arteries were similar to those induce by clonidine. In tail arteries, NG-nitro-L-arginine, a nitric oxide synthase inhibitor, at a concentration that completely inhibited acetylcholine-induced relaxations did not significantly affect the relaxation induced by clonidine. In contrast, the relaxation of thoracic aorta in response to clonidine was partly reduced in the presence of NG-nitro-L-arginine. These results indicate that the alpha2-adrenoceptor agonists selectively inhibit the contractions induced by phenylephrine in both tissues. Regional differences in the modes of the inhibition by the alpha2-adrenoceptor agonists exist.     

Evidence that angiotensin II, endothelins and nitric oxide regulate mitogen-activated protein kinase activity in rat aorta

We measured the activity of mitogen-activated protein (MAP) kinases, enzymes believed to be involved in the pathway for cell proliferation, in rat aortic strips with or without endothelium, and examined effects of angiotensin receptor antagonists, endothelin receptor antagonists and nitric oxide (NO)-related agents. Endothelium removal produced an activation of MAP kinase activity in the strips, whereas the enzyme activity was not affected in the adventitia. The MAP kinase activation was inhibited by either the angiotensin AT1 receptor antagonist losartan or the endothelin ETA receptor antagonist BQ 123. The combination of both antagonists caused an additive inhibition. The angiotensin AT2 receptor antagonist PD 123,319 and the endothelin ETB receptor antagonist BQ 788 did not affect the MAP kinase activation. The NO synthase inhibitor NG-nitro-L-arginine methyl ester (L-NAME) caused an activation of MAP kinase in the endothelium-intact aorta and the MAP kinase activation was inhibited by losartan or BQ123. The NO releaser nitroprusside inhibited the MAP kinase activation induced by endothelium removal or angiotensin II. These results suggest that even in isolated arteries, NO of endothelial origin tonically exert MAP kinase-inhibiting effects and endogenous angiotensin II and endothelins in the media are tonically released to cause MAP kinase-stimulating effects in medial smooth muscle.     

Interaction between thiol-modifying agents and P1075, a K(ATP) channel opener, in rat isolated aorta

In the past year progress in the study of cationic species has been made, particularly in our understanding of the factors which control the selective recognition of biologically important cations such as ammonium, alkali and alkaline earth metal ions, and of metal ions used in biomedicine such as lanthanides and iron(III). Based on this knowledge, several new hosts with improved transport, photophysical and biological properties have been designed.     

Endothelium-dependent effect of perindopril and enalaprilat in rat thoracic aorta

AIM: To study the effect of the angiotensin-converting enzyme (ACE) inhibitors perindopril (Per) and enalaprilat (Ena) on the reactivity of the endothelium in normal rats. METHODS: Male rats were treated intragastrically with Per (2 mg.kg-1.d-1) or placebo (n = 18) for 6 wk. Aorta was isolated for experiment. Another set of isolated aortic rings with and without endothelium were incubated with Ena (0.1 mumol.L-1) for 30 min. Responses to acetylcholine, serotonin, phenylephrine, sodium nitroprusside (SN), and nitroglycerin (Nit) were observed. RESULTS: Endothelium-dependent relaxation to acetylcholine was augmented in aortic rings from rats treated with Per in comparison with control. The IC50 value (95% confidence limits) decreased from 3.8 (0.56-26.1) mumol.L-1 (control group) to 0.98 (0.28-3.41) mumol.L-1 (Per-treated group). The maximal relaxation was augmented from 62 +/- 9% to 78 +/- 10% (P < 0.01). However, the responses to the endothelium-independent vasodilators, SN and Nit, were similar. Serotonin- and phenylephrine-induced contractions were decreased, which were influenced by basal release of endothelium-derived relaxing factor (EDRF). EC50 values was 6.1 (2.6-14.4) nmol.L-1 vs 8.3 (3.6-18.8) nmol.L-1 in comparison with control group and Per-treated group. The maximal contraction was decreased from 2.42 +/- 0.29 g (control group) to 1.96 +/- 0.25 g (treated group) (P < 0.01). Similar results were found in incubation with Ena. CONCLUSION: Ena and Per enhanced the basic release of EDRF from vascular endothelium.     

Metal ion stimulation of phospholipase D-like activity of isolated rat intestinal mitochondria

OBJECTIVE: To investigate the role of phospholipase during the activation and priming of neutrophil nicotinamide adenine dinucleotide phosphate (NADPH) oxidase by peritoneal dialysis effluent (PDE). DESIGN: Examine the action of 4-hour dwell PDE upon phospholipase activation in the circulating neutrophils obtained from healthy individuals. RESULTS: We have previously reported that PDE stimulated superoxide release by the NADPH oxidase of human neutrophils and primed the response to the bacterial peptide, fMLP (fMetLeuPhe). To elucidate the biochemical mechanisms underlying these observations, we have examined the roles of phospholipases (PL) C, D, and A2, whose activation causes the release of a range of intracellular secondary messengers. Following fMLP stimulation, we observed a rapid activation of both PLC and PLD as well as a small but nonsignificant increase in PLA2 activity. Peritoneal dialysis effluent alone failed to stimulate either PLC or PLD, while pre-incubation with PDE had no affect upon fMLP-induced PLC and PLD activation. However, PDE caused a small but nonsignificant increase in PLA2 activity (which was comparable to that observed with fMLP) and primed the fMLP-induced response. In common with a role for PLA2 and the subsequent release of arachidonic acid (AA), we have demonstrated dose-dependent inhibition of PDE-induced superoxide release by the PLA2 inhibitor mepacrine, as well as activation and priming of the fMLP-induced superoxide generation by AA. CONCLUSIONS: These results imply that PDE-induced NADPH-oxidase activation and priming in human neutrophils is mediated via a PLA2-dependent but PLC- and PLD-independent mechanism.     

Possible mechanism of captopril induced endothelium-dependent relaxation in isolated rabbit aorta

Natural resistance-associated macrophage protein 1 (NRAMP1) is a putative membrane protein that dominates natural resistance to infection. An NRAMP1-glutathione S-transferase fusion protein was used to test the ability of the NRAMP1 NH2-terminal domain to bind to taxol-stabilized microtubules. Co-sedimentation analysis showed that the fusion protein binds to microtubules. Although the NH2-terminal domain of the NRAMP1 molecule has structural homology with the Pro-rich region of microtubule-associated protein 4 (MAP4), the presence of the MAP4 microtubule-binding domain fragment had little effect on the binding of the fusion protein to microtubules.     

Relaxation of isolated aorta of the rabbit by picolinic acids

1 When the isolated thoracic aorta of the rabbit was contracted with prostaglandin F2alpha, 5-alkylpicolinic acids produced dose-dependent relaxations. 2 Picolinic acid, 2,5-pyridinedicarboxylic acid and 5-acetylpicolinic acid which do not have the 5-alkyl residue failed to relax blood vessels. 3 The vascular relaxation was dependent on the number of carbon atoms in the 5-alkyl compounds. 4 Relaxations which occurred with 5-alkylpicolinic acids were not affected by pretreatment with either propranolol or atropine. 5 It is concluded that the 5-alkyl residue is necessary for the vascular relaxation with 5-alkylpicolinic acid and that it was not produced by stimulation of beta-adrenoceptors or cholinoceptors but rather through an activation of the basic process.     

Buthus martensi karsch venom: prejunctional adrenergic activity in the rat isolated anococcygeus muscle

The effect of Buthus martensi Karsch venom (MKV) on adrenergic responses was investigated using the rat isolated anococcygeus muscle (Acm), since several scorpion venoms can cause peripheral sympathetic nerve stimulation with enhanced adrenergic responses. The effects of phentolamine (5 microM), guanethidine (5 microM), tetrodotoxin (2 microM), desipramine (1.5 microM) and reserpine pretreatment in vivo (5 mg/kg s.c. x 24 hr and 5 mg/kg i.p. x 3 hr) on contractile responses of the rat Acm to field stimulation, noradrenaline (3 microM), tyramine (10 microM), crude MKV (2 micrograms/ml), carbachol (3 microM) and potassium chloride (50 mM) were compared. Phentolamine, guanethidine, tetrodotoxin and reserpine pretreatment completely blocked the contractile responses of the Acm to MKV and to field stimulation but desipramine potentiated the responses. The responses to NA were completely blocked by phentolamine, but were potentiated by guanethidine, desipramine and reserpine pretreatment. The contractile responses to tyramine were completely blocked by phentolamine, desipramine and reserpine pretreatment. The low doses (0.1 microgram/ml x 3) of MKV, which did not produce any observable increase in tone of the anococcygeus muscle, potentiated the contractile responses to field stimulation, but not the responses to exogenous NA. Thus, the adrenergic agonist action of MKV in the rat isolated anococcygeus muscle is mediated by some prejunctional mechanism(s) of action, presumably stimulating the release of the neurotransmitter noradrenaline.