Pertussis toxin differentiates between alpha 1- and alpha 2-adrenoceptor-mediated inhibition of noradrenaline release from rat kidney cortex

In slices of rat kidney cortex incubated in [3H]noradrenaline, the alpha 1-adrenoceptor agonist methoxamine (10 microM), the alpha 2-adrenoceptor agonist clonidine (0.1 microM), as well as adenosine (10 microM), inhibited the electrical stimulation-induced (S-I) outflow of radioactivity, at a stimulation frequency of 1 Hz. Prior treatment of rats with pertussis toxin (25 micrograms/kg i.v.), which abolished the negative inotropic effect of carbachol (10 microM) on isolated atria, prevented the inhibition caused by methoxamine, but not that caused by clonidine or adenosine. At a stimulation frequency of 5 Hz, the alpha 2-adrenoceptor antagonist idazoxan (0.1 microM) and the prostaglandin synthesis inhibitor indomethacin (10 microM) both facilitated the S-I outflow of radioactivity, and neither of these effects were altered by pertussis toxin. These results suggest that a pertussis toxin sensitive G-protein is involved in alpha 1-adrenoceptor inhibition of noradrenaline release, but not in alpha 2-adrenoceptor, adenosine or prostaglandin inhibition.     

Prejunctional alpha 2-adrenoceptors in mouse atria function through G-proteins which are sensitive to N-ethylmaleimide, but not pertussis toxin.

1. The identity of the G-proteins involved in prejunctional alpha 2-adrenoceptor signal transduction in mouse atria was examined by use of the G-protein inactivators N-ethylmaleimide and pertussis toxin. 2. The alpha 2-adrenoceptor partial agonist clonidine (0.03 microM) inhibited the electrical stimulation-induced (S-I) outflow of radioactivity from mouse atria which were incubated with [3H]-noradrenaline and stimulated at 5 Hz. The partial alpha 2-adrenoceptor agonist St 363 (10 microM) inhibited the S-I outflow of radioactivity at the lower stimulation frequency of 2.5 Hz. The inhibitory effects of these compounds were not altered in mice pretreated with pertussis toxin (1.5 micrograms, i.v.). 3. The alpha 2-adrenoceptor antagonist, idazoxan (0.1 microM), increased the S-I outflow of radioactivity from mouse atria stimulated at 5 Hz, and this effect was not altered in atria from mice pretreated with pertussis toxin. 4. The inhibitory effects of clonidine and St 363 and the facilitatory effect of idazoxan on the S-I outflow of radioactivity from mouse atria were significantly less in atria incubated with N-ethylmaleimide (NEM, 3 microM) for 60 min before the [3H]-noradrenaline incubation. 5. The results suggest that prejunctional alpha 2-adrenoceptors in mouse atria function through G-proteins which are NEM-sensitive, but pertussis toxin insensitive.     

The effect of pertussis toxin treatment on alpha-1-adrenoceptor-mediated pressor responses in the pithed rat: dependence on agonist efficacy but not chemical class.

The role of pertussis-toxin-sensitive guanine nucleotide regulatory proteins (G proteins) in the signal transduction processes involved in post-junctional vascular alpha 1-adrenoceptor-mediated vasoconstriction has been investigated in the pithed rat using two chemical classes of alpha-adrenoceptor agonists, the phenethylamines and imidazolines, in order to determine if they utilize different signal transduction mechanisms. Pertussis toxin pretreatment (50 micrograms/kg, i.v., 3 days prior to experimentation) slightly inhibited the pressor response to the full alpha 1-adrenoceptor agonist of the phenethylamine class (-)-norepinephrine (in the presence of rauwolscine, 1 mg/kg, i.v.), whereas it markedly inhibited the pressor response to the partial alpha 1-adrenoceptor agonist of the imidazoline class oxymetazoline (in the presence of rauwolscine, 1 mg/kg, i.v.). However, after elimination of the alpha 1-adrenoceptor reserve for (-)-norepinephrine with phenoxybenzamine (0.1 mg/kg, i.v.), the pressor response to this agonist became sensitive to inhibition by pertussis toxin treatment. The pattern of inhibition of alpha 1-adrenoceptor-mediated pressor responses produced by pertussis toxin was similar to that produced by the calcium channel antagonist nifedipine (1 mg/kg, i.a.). The results support the hypothesis that vascular alpha 1-adrenoceptors may be coupled to a G protein which is sensitive to pertussis toxin and which couples the alpha 1-adrenoceptor to the influx of extracellular calcium, which possibly another G protein that is insensitive to pertussis toxin that couples the alpha 1-adrenoceptor to the release of intracellular calcium. The intrinsic efficacy of the agonist, and not its chemical class, determines which signal transduction mechanisms will be utilized.     

Differential effect of pertussis toxin on pre- and postjunctional alpha 2-adrenoceptors in the cardiovascular system of the pithed rat

The role of the pertussis toxin-sensitive guanine nucleotide binding regulatory protein in pre- and postjunctional alpha 2-adrenoceptor-mediated responses has been investigated in the pithed rat. Pertussis toxin (50 micrograms/kg i.v., administered 3 days prior to experimentation) markedly inhibited the alpha 2-adrenoceptor-mediated vasopressor response to B-HT 933, but had no effect on the alpha 2-adrenoceptor-mediated cardiac neuroinhibitory effect of B-HT 933. Thus, postjunctional alpha 2-adrenoceptor activation in vascular smooth muscle, but not prejunctional alpha 2-adrenoceptor activation on sympathetic neurons, utilizes a pertussis toxin-sensitive guanine nucleotide binding regulatory protein.     

Effects of Bordetella pertussis toxin pretreatment on the antiarrhythmic action of ischaemic preconditioning in anaesthetized rats.

1. Bordetella pertussis toxin, which catalyses the ADP-ribosylation of certain guanine nucleotide binding proteins (G proteins), thus functionally uncoupling them from associated receptors, was examined to determine whether it modified the antiarrhythmic effect of ischaemic preconditioning in anaesthetized rats. 2. Pertussis toxin (25 micrograms kg-1, i.p., 48 h prior to heart isolation) attenuated the negative chronotropic effect of acetylcholine (ACh) in rat isolated Langendorff perfused hearts. ACh (10 microM) reduced heart rate by 4% in hearts taken from pertussis toxin-treated animals, compared to a reduction of 57% in hearts taken from animals treated only with vehicle. 3. In anaesthetized rats, ischaemic preconditioning (a single 3 min occlusion of the left main coronary artery followed by 10 min reperfusion) had a pronounced antiarrhythmic effect during a subsequent 30 min period of regional myocardial ischaemia. Compared to hearts receiving only a 30 min period of left coronary occlusion, there was a reduced mortality (67% and 0% for control and preconditioned groups, respectively; P < 0.01) and decreased incidences of ventricular tachycardia (VT) and ventricular fibrillation (VF). Pretreatment with pertussis toxin (25 micrograms kg-1, i.p., 48 h previously) did not modify the arrhythmias associated with a 30 min period of regional myocardial ischaemia, neither did it modify the reduction in mortality (from 56% to 0%; P < 0.05) associated with preconditioning. Furthermore, the decrease in total ventricular premature beat count induced by preconditioning seen in controls (from 427 +/- 130 to 95 +/- 45) was also seen in pertussis toxin-treated rats (from 252 +/- 190 to 57 +/- 25).(ABSTRACT TRUNCATED AT 250 WORDS)     

The effects of aldose reductase inhibition with ponalrestat on changes in vascular function in streptozotocin diabetic rats.

1. The responses of rat isolated aortae to vasoconstrictor and vasodilator agents have been studied in 14-day streptozotocin-diabetic rats. The effects of treatment with the aldose reductase inhibitor, ponalrestat, on these responses have also been investigated. 2. Maximum contractile responses and aortic sensitivity to phenylephrine were significantly enhanced in 14-day diabetic aortae. 3. In contrast, endothelium-dependent relaxations to carbachol were depressed in diabetic rats, whilst endothelium-independent relaxations to forskolin and sodium nitroprusside were unchanged. 4. Pretreatment with ponalrestat (25 mg kg-1, daily) prevented both the enhanced maximum contractile responses to phenylephrine and the depressed endothelium-dependent relaxations to carbachol in aortae from 14-day diabetic rats. Ponalrestat however, had no effect on the reduced phenylephrine EC50 values observed in tissues from diabetic animals. 5. It is concluded that ponalrestat prevents the depression of endothelium-dependent aortic relaxations induced by diabetes of 14 days duration, suggesting that the polyol pathway is involved in these vascular changes. Ponalrestat does not prevent the increase in aortic sensitivity to alpha 1-adrenoceptor agonists.     

Pertussis toxin-sensitive and -insensitive mechanisms of alpha1-adrenoceptor-mediated inotropic responses in rat heart.

In rat left ventricular papillary muscle, phenylephrine, an alpha1-adrenoceptor agonist, induced a triphasic inotropic response; an initial transient, small, positive inotropic effect followed by a transient chloroethylclonidine-sensitive negative inotropic effect and a sustained 2-(2,6-dimethoxyphenoxyethyl)aminomethyl-1,4-benzodioxane (WB4101)-sensitive positive inotropic effect. Treatment with pertussis toxin for 2 days significantly inhibited only the transient negative inotropic effect without changing the sustained positive inotropic effect. This treatment also prevented the acetylcholine (1 microM)-induced negative inotropic effect. Further, phenylephrine-induced transient negative inotropic effect was attenuated in the presence of ouabain. These results suggest that pertussis toxin-sensitive or -insensitive G-protein may be responsible for alpha1-adrenoceptor subtype-mediated negative inotropic effect or positive inotropic effect, respectively, in which the transient negative inotropic effect was produced via the stimulation of Na+, K+ pump, presumably through pertussis toxin-sensitive G-protein-dependent pathway.     

The involvement of endothelium-derived relaxing factor in the regulation of renal cortical blood flow in the rat.

1. In the present study the role of endogenous nitric oxide (NO) was investigated, in the regulation of renal cortical blood flow (RCBF) in vivo in anaesthetized rats under conditions in which prostacyclin involvement had been eliminated. 2. Infusions of the NO synthesis inhibitor NG-monomethyl-L-arginine (MeArg) at 1 or 3 mg kg-1 min-1, i.v., produced significant decreases in RCBF of 29 +/- 7% and 35 +/- 5%, respectively. These effects were reversed by co-infusion of a 3 fold excess of L-arginine (L-Arg). 3. Similarly, intravenous infusion of N omega-nitro-L-arginine methyl ester (NO2Arg) at 30 or 300 micrograms kg-1 min-1 attenuated RCBF by 21 +/- 4% or 53 +/- 4%, respectively, and these effects were reversed by L-Arg (3 or 10 mg kg-1 min-1, i.v.). Most importantly, a low dose of NO2Arg (30 micrograms kg-1 min-1, i.v.), while having no pressor effect, considerably reduced RCBF, indicating that basal release of NO is important for the maintenance of renal cortical blood flow. 4. MeArg (3 mg kg-1 min-1, i.v.) or NO2Arg (300 micrograms kg-1 min-1, i.v.) inhibited endothelium-dependent acetylcholine (ACh, 10 micrograms kg-1 min-1, i.v. for 3 min) increases in RCBF in an L-Arg reversible manner, but did not affect endothelium-independent (dopamine 10 micrograms kg-1 min-1, i.v., for 3 min) increases.(ABSTRACT TRUNCATED AT 250 WORDS)     

Modulation by central postsynaptic alpha 2-adrenoceptors of the jaw-opening reflex induced by orofacial stimulation in rats.

1. The modulation by alpha 2-adrenoceptors of the jaw-opening reflex (digastric electromyographic responses) elicited by orofacial electrical stimulation (OF-JOR) in pentobarbitone anaesthetized rats was investigated. 2. Increasing doses of clonidine (0.1-1000 micrograms kg-1, i.v.) reduced, in a dose-dependent manner until abolition, the amplitude and duration of the OF-JOR and increased the latency to onset. The sum of amplitudes of the reflex was the most sensitive parameter to the inhibitory effects of clonidine (ED50 = 13.9 micrograms kg-1). 3. Pretreatment with the alpha 2-adrenoceptor antagonist, idazoxan (0.03-1 mg kg-1, i.v.), caused a dose-dependent shift (1.5 to 37 fold) to the right of the dose-response curve for clonidine without significant change of maximum inhibitory effect, in a manner compatible with competitive antagonism (ED50B = 29.0 micrograms kg-1). Pretreatment with yohimbine (0.3 mg kg-1, i.v.) also antagonized the inhibitory effect of clonidine on the OF-JOR. In contrast, the alpha 2-adrenoceptor antagonist ARC-239 (0.3 mg kg-1, i.v.) did not antagonize the effect of clonidine on the reflex. 4. In rats pretreated with reserpine (5 mg kg-1, s.c., 18 h) the OF-JOR was not modified, but the potency of clonidine in inhibiting the reflex was potentiated (ED50 value decreased to 6.8 micrograms kg-1) without a significant change of maximum inhibitory effect. 5. Increasing doses of amphetamine (0.1-3000 micrograms kg-1, i.v.) caused a dose-related, but partial, inhibition of the OF-JOR (ED50 = 135 micrograms kg-1; Emax = 67%).(ABSTRACT TRUNCATED AT 250 WORDS)     

Differential effects of cyclooxygenase inhibitors on the cardiovascular response to hyperosmotic mannitol

The intravenous infusion of mannitol (1.37 mosm kg-1 min-1 during 4 min) to anesthetized, open-chest, intact dogs produced a hemodynamic response characterized by a positive inotropic effect and a decrease in total peripheral resistance. This response is attributable to a direct effect of hypertonicity (plasma osmolality raised from 309 +/- 4 to 332 +/- 5 mosm kg-1 H2O). Pretreatment with indomethacin (5 mg kg-1 i.v.) or meclofenamic acid (2 mg kg-1 i.v.) attenuated the mannitol-induced response but acetylsalycilic acid (10 mg kg-1 i.v.) was without effect. Indomethacin (5 mg kg-1 i.v.) did not change the decrease in canine hindlimb perfusion pressure produced by intra-arterial mannitol 5 mosm kg-1. Indomethacin (3 microM) did not alter the chronotropic and inotropic responses to mannitol (150 mosm above normal) in isolated rabbit atria. These results suggest that the cardiovascular response to hyperosmotic mannitol may be partly mediated in intact animals by prostaglandin release.     

PERTUSSIS TOXIN-SENSITIVE G-PROTEINS AND REGULATION OF BLOOD PRESSURE IN THE SPONTANEOUSLY HYPERTENSIVE RAT

1. Increased Gi-protein-mediated receptor-effector coupling in the vasculature of the spontaneously hypertensive rat (SHR) has been proposed as a contributing factor in the maintenance of elevated blood pressure. If increased Gi-protein-mediated activity plays an important role in hypertension in SHR, then inhibition of Gi-proteins by pertussis toxin would be expected to decrease blood pressure in this genetic hypertensive model. To address this hypothesis, studies were undertaken comparing the cardiovascular effects of pertussis toxin in SHR and normotensive Wistar-Kyoto (WKY) rats. 2. Spontaneously hypertensive and WKY rats were instrumented with radiotelemetry devices and blood pressure measurements were recorded in conscious rats. Following a single injection of pertussis toxin (10 micrograms/kg, i.v.), mean arterial blood pressure fell from 161 +/- 3 to 146 +/- 1 mmHg in the SHR and the effect was sustained for more than 2 weeks. In contrast, 10 micrograms/kg, i.v., pertussis toxin produced no significant effect on blood pressure in WKY rats (103 +/- 4 vs 101 +/- 5 mmHg). 3. In a separate study, SHR and WKY rats were administered 30 micrograms/kg, i.v., pertussis toxin or 150 microL/kg, i.v., saline and, 3-5 days later, rats were anaesthetized and instrumented to permit measurement of blood pressure and renal function. At this higher dose, pertussis toxin reduced blood pressure in both strains of rat, although the effect was markedly greater in SHR (approximately 40 mmHg decrease) compared with WKY rats (approximately 15 mmHg decrease). In SHR, pertussis toxin increased renal blood flow (from 5.7 +/- 0.3 to 7.5 +/- 0.8 mL/min per g kidney) and decreased renal vascular resistance (from 31 +/- 2 to 19 +/- 2 mmHg/mL per min per g kidney). In WKY rats, pertussis toxin had no significant effect on renal parameters. 4. Results from these studies indicate that a pertussis toxin-sensitive Gi-protein-mediated pathway contributes to the maintenance of hypertension and elevated renal vascular tone in the SHR.     

Pertussis toxin suppresses carbachol-evoked cardiodepression but does not modify cardiostimulation mediated through β1- and putative β4-adrenoceptors in mouse left atria: no evidence for β2- and β3-adrenoceptor function

Activation of beta1-, beta2-, beta 3- and putative beta4-adrenoceptors modifies cardiac function. These receptors are usually coupled to Gs protein, but beta2- and beta3-adrenoceptors could also couple to Gi/o proteins. The mouse heart is used increasingly for studies of genetically disrupted or overexpressed proteins, including beta-adrenoceptor subtypes. We therefore investigated in contracting mouse left atria (2 Hz, 37 degrees C) if inactivation of Gi/o proteins with pertussis toxin modifies or uncovers effects mediated through beta-adrenoceptor subtypes. The negative inotropic effects of carbachol in atria exposed to catecholamine or high calcium (6.8 mmol/l) were assumed to be mediated through activation of muscarinic receptors coupled to Gi/o. We report conditions under which incubation of left atria with 200 ng/ml pertussis toxin for 24 h nearly abolished the carbachol responses. Although it has been reported that muscarinic receptor-mediated cardiodepression has an obligatory contribution of nitric oxide, the nitric oxide synthase inhibitor N(G)-monomethyl-L-arginine (0.1-1 mmol/l) did not modify the negative inotropic effects of carbachol, inconsistent with an involvement of nitric oxide. The positive inotropic effects of (-)-noradrenaline and (-)-adrenaline, mediated through beta1-adrenoceptors, were not affected by pertussis toxin. (-)-Adrenaline did not cause positive inotropic effects attributable to beta2-adrenoceptor-mediation, in the presence of CGP 20712A (300 nmol/l) to block beta1-adrenoceptors, in control atria or atria pretreated with pertussis toxin. The positive inotropic effects of (-)-CGP 12177 (1 micromol/l), a compound with agonist activity at the putative beta4-adrenoceptor, were unaffected by pertussis toxin. The beta3-adrenoceptor-selective agonist BRL 37344 (1 micromol/l), in the presence of (-)-propranolol (200 nmol/l), did not cause positive or negative inotropic effects in control and pertussis toxin-treated atria. In left atria obtained from mice injected with 150 microg/kg i.p. pertussis toxin which abolished carbachol-evoked cardiode-pression, the positive inotropic effects of (-)-adrenaline were antagonised by CGP 20712A. The beta2-adrenoceptor-selective antagonist ICI 118551 (50 nmol/l) did not cause additional blockade of the effects of high (-)-adrenaline concentrations in the presence of CGP 20712A, ruling out the involvement of beta2-adrenoceptors. The results with intraparenteral PTX validate our in vitro PTX method. We conclude that inhibition of murine Gi/o proteins does not alter atrial positive inotropic effects mediated through beta1- and putative beta4-adrenoceptors and does not reveal functional beta2- and beta3-adrenoceptors.     

Supersensitivity of isolated atria from diabetic rats to adenosine and methacholine: modulation by pertussis toxin.

The chronotropic response of isolated right atria obtained from rats made diabetic 14-15 weeks previously by streptozotocin, was compared with age-matched controls. Diabetic rat atria are significantly more sensitive to the negative chronotropic actions of adenosine and of methacholine. Pretreating both control and diabetic rats with 2.5 mg kg-1 pertussis toxin attenuated the negative chronotropic effects of methacholine and adenosine on isolated atria, although diabetic atria still displayed a significantly greater sensitivity to these agonists (P less than 0.05-0.001). The negative chronotropic effects of methacholine and adenosine on both control and diabetic atria were abolished following pretreatment with higher doses of pertussis toxin (10 mg kg-1). These results suggest that pertussis toxin-sensitive G proteins may be involved in the supersensitivity of diabetic hearts to methacholine and adenosine.     

Allopurinol and amlodipine improve coronary vasodilatation after myocardial ischaemia and reperfusion in anaesthetized dogs.

1. We have assessed the effect of allopurinol, amlodipine and propranolol pretreatment on both endothelium-dependent and endothelium-independent coronary vasodilatation in vivo, by comparing pre-ischaemic responses with those measured after 60 min of coronary artery occlusion and 30 min of reperfusion in anaesthetized dogs. 2. In 15 untreated dogs ischaemia and reperfusion attenuated the increases in coronary blood flow produced by either acetylcholine (0.01-0.05 micrograms kg-1, i.a.) or glyceryl trinitrate (0.05-0.2 micrograms kg-1, i.a.), to an average of 39 +/- 4% and 42 +/- 5% of the pre-ischaemic control response, respectively (both P < 0.05). 3. In 5 dogs treated with allopurinol (25 mg kg-1, orally, 24 h previously, plus 50 mg kg-1, i.v., 5 min before occlusion), the increases in coronary blood flow after ischaemia and reperfusion (acetylcholine: 78 +/- 12%, glyceryl trinitrate: 60 +/- 3% of pre-ischaemic response) were significantly larger than post-ischaemic responses in untreated dogs (both P < 0.05). 4. Similarly, amlodipine treatment (3 micrograms kg-1 min-1, i.v., starting 90 min before occlusion) in 5 dogs improved post-ischaemic increases in blood flow (acetylcholine: 58.5%, glyceryl trinitrate: 66 +/- 6% of pre-ischaemic response, significantly greater than post-ischaemic responses in untreated dogs, P < 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)     

Impaired endothelium-dependent relaxation of dog coronary arteries after myocardial ischaemia and reperfusion: prevention by amlodipine, propranolol and allopurinol.

1. Anaesthetized, open-chest dogs were subjected to 60 min of left circumflex coronary artery occlusion followed by 90 min of reperfusion. Endothelium-dependent and -independent relaxant responses of the isolated coronary arterial rings were then investigated. 2. The endothelium-dependent, acetylcholine-induced relaxation of ischaemic/reperfused arterial rings was significantly attenuated in comparison to control rings (1.9 fold rightward shift, ischaemic/reperfused maximum relaxation = 57 +/- 13% of control maximum relaxation; P less than 0.05). In contrast, glyceryl trinitrate produced similar relaxant responses in control and ischaemic rings. 3. Pretreatment of dogs with either amlodipine (3 micrograms kg-1 min-1, i.v.) or propranolol (1 mg kg-1, i.v.) completely prevented the postischaemic impairment of endothelium-dependent relaxant responses (100 +/- 3% and 90 +/- 5% of control maximum relaxation, respectively). 4. Allopurinol pretreatment (25 mg kg-1, p.o. 24 h previously, plus 50 mg kg-1 i.v. 5 min before arterial occlusion) partially protected against endothelial dysfunction by preventing the ischaemia-induced rightward shift of the acetylcholine relaxation curve and increasing the maximum relaxation response (83 +/- 7% of control rings). 5. These results confirm that endothelium-dependent coronary vascular relaxation is impaired by ischaemia and reperfusion, and that the ischaemia-induced impairment is reduced by pretreatment with amlodipine, propranolol or allopurinol.     

Pharmacological differentiation by pertussis toxin of the in vivo acute responses to fMLP and PAF in guinea-pig lungs.

1. The effects of pertussis toxin on the N-formyl-L-methionyl-L-leucyl-L-phenylalanine (fMLP) and platelet-activating factor (PAF)-induced variations in pulmonary capillary albumin exchanges, blood volume, leucocyte or platelet sequestration were studied in the guinea-pig, by use of radioactive tracers. The effects of pertussis toxin on pulmonary insufflation pressure were studied in parallel. 2. The i.v. administration of fMLP and PAF to the guinea-pig was followed by bronchoconstriction, increased lung capillary albumin exchanges (vasopermeability) sequestration of leucocytes, leucopenia and reduction of blood volume (vasoconstriction). PAF also induced platelet sequestration in lungs and thrombocytopenia. 3. Pertussis toxin (10 micrograms kg-1, i.v., 72 h before the experiment) prevented all the studied fMLP-induced effects, but failed to modify PAF-induced bronchoconstriction, lung vasoconstriction, platelet sequestration, thrombocytopenia and the increased capillary vasopermeability. In the same conditions the lung leucocyte sequestration was not significantly affected when leucopenia was partially reduced. 4. It is suggested that the effects of fMLP, but not those of PAF, involve a Gi-like protein.     

Study of the in vivo and in vitro cardiovascular effects of (+)-glaucine and N-carbethoxysecoglaucine in rats.

1. The cardiovascular and vasorelaxant effects of (+)-glaucine and of a semisynthetic derivative (N-carbethoxysecoglaucine) were studied in rats. 2. N-carbethoxysecoglaucine did not modify either systolic arterial pressure or heart rate values in conscious (25 mg kg-1, p.o.) and anaesthetized normotensive rats (5 mg kg-1, i.v.). Furthermore, this compound showed no activity in the experiments carried out on rat isolated aorta [contractility and 45Ca2+ influx assays (5 microM)] and did not modify the rate and force of contraction in rat isolated atria (5 microM). 3. In conscious normotensive rats, oral administration of (+)-glaucine (25 mg kg-1) did not modify either systolic arterial pressure or heart rate. 4. In anaesthetized normotensive rats, (+)-glaucine (5 mg kg-1, i.v.) produced a remarkable fall in mean arterial pressure (MAP) accompanied by a significant decrease in heart rate. In the same preparation, (+)-glaucine (5 mg kg-1, i.v.) did not modify the cardiovascular effects induced by noradrenaline (NA) (5 micrograms kg-1) and 5-hydroxytryptamine (5-HT) (300 micrograms kg-1) but markedly inhibited those induced by nicotine (200 micrograms kg-1). 5. In isolated intact aorta of rat, (+)-glaucine (0.15-5 microM) competitively inhibited the contractions induced by NA (with a pA2 value of 7.14) and non-competitively those induced by 5-HT (in normal Krebs solution) and Ca2+ (in depolarizing Ca(2+)-free high-K+ 50 mM solution), with depression of the maximal response and with pD2 values of 5.56 and 5.26, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of glyceryl trinitrate on endothelium-dependent and -independent relaxation and cyclic GMP levels in rat aorta and human coronary artery

The effects of glyceryl trinitrate-induced desensitization on relaxations and/or elevated cyclic GMP levels due to the nitrogen oxide-containing vasodilators (glyceryl trinitrate and sodium nitroprusside), the endothelium-dependent vasodilators (acetylcholine and the Ca2+ ionophore A23187), and the atrial peptides (atriopeptin II) were investigated in the rat thoracic aorta and human coronary artery. Prior exposure of rat thoracic aorta to glyceryl trinitrate decreased relaxations to glyceryl trinitrate, sodium nitroprusside, and acetylcholine, whereas relaxations to atriopeptin II and 8-bromo cyclic GMP remained unaltered. In human coronary artery, glyceryl trinitrate pretreatment inhibited relaxations to glyceryl trinitrate, sodium nitroprusside, and the Ca2+ ionophore A23187. Relaxation to glyceryl trinitrate was inhibited more than that to sodium nitroprusside in both tissues. Acetylcholine-induced relaxation in rat thoracic aorta was slightly inhibited, whereas relaxation to the Ca2+ ionophore A23187 in human coronary artery was markedly depressed. Pretreatment with glyceryl trinitrate decreased the elevated cyclic GMP levels due to glyceryl trinitrate and acetylcholine in rat thoracic aorta and to glyceryl trinitrate and the Ca2+ ionophore A23187 in human coronary artery. Removal of the endothelium abolished the increased cyclic GMP levels and relaxation due to the Ca2+ ionophore A23187 and decreased basal cyclic GMP levels in the human coronary artery. In contrast, atriopeptin II-induced increased cyclic GMP levels were unaltered by glyceryl trinitrate pretreatment in rat thoracic aorta. The present results suggest that: glyceryl trinitrate-induced desensitization inhibits relaxation to the nitrogen oxide-containing vasodilators and endothelium-dependent vasodilators in both the rat thoracic aorta and human coronary artery: the inhibition of relaxation is associated with decreased formation of cyclic GMP;(ABSTRACT TRUNCATED AT 250 WORDS)     

Adrenergic-cholinergic interactions in left atria: a study using K+ channel agonists, antagonist and pertussis toxin.

1. The role of activation of potassium conductance in the antagonism by the muscarinic agonist carbachol of positive inotropic responses to alpha- and beta-adrenoceptor stimulation was studied in electrically driven left atrial strips of the rabbit. 2. The potassium channel antagonist, 4-aminopyridine, attenuated the direct negative inotropic response to carbachol and the reversal by carbachol of positive inotropic responses to the alpha-adrenoceptor agonist phenylephrine (in the presence of timolol). The inhibitory effect of carbachol on positive inotropic responses to the beta-adrenoceptor agonist isoprenaline was much less affected by 4-aminopyridine. 3. Pretreatment of rabbits with pertussis toxin also attenuated the direct negative inotropic response to carbachol and the inhibitory effect of carbachol on positive inotropic responses to phenylephrine. 4. Neither carbachol nor phenylephrine, alone or in combination, had any effect on left atrial adenosine 3':5'-cyclic monophosphate (cyclic AMP) levels. 5. The potassium channel agonist, pinacidil, exerted a dose-dependent negative inotropic response in rabbit left atria and reversed positive inotropic responses to phenylephrine and isoprenaline. In the dose-range tested, pinacidil had a greater inhibitory effect on positive inotropic responses to phenylephrine than on positive inotropic responses to isoprenaline. 6. Pretreatment of left atria with pinacidil or cromakalim, another potassium channel agonist, antagonized positive inotropic responses to phenylephrine but not to isoprenaline. 7. These results suggest that activation of potassium conductance plays an important role in the inhibition by carbachol of positive inotropic responses of rabbit left atria to phenylephrine but not to isoprenaline.     

Selective inhibition of basal but not agonist-stimulated activity of nitric oxide in rat aorta by NG-monomethyl-L-arginine.

1. Two inhibitors of nitric oxide synthase, NG-monomethyl-L-arginine (L-NMMA, 1-100 microM) and NG-nitro-L-arginine (L-NOARG, 3-300 microM), each produced a concentration-dependent augmentation of phenylephrine-induced tone in endothelium-containing but not endothelium-denuded rings of rat aorta. Pretreatment with L-arginine (10 mM) prevented the augmentation of tone induced by L-NOARG and L-NMMA. 2. Following induction of sub-maximal tone with phenylephrine in endothelium-containing rings, acetylcholine (1 nM-3 microM) induced relaxations which were inhibited in a concentration-dependent manner by L-NOARG (10-100 microM). 3. In contrast to the action of L-NOARG, L-NMMA (100-1000 microM) had no effect on acetylcholine-induced relaxations. L-NMMA (100-300 microM) also had no effect on the endothelium-dependent relaxant actions of ATP (0.1-100 microM), whereas L-NOARG (100 microM) produced powerful blockade. 4. Unexpectedly, pretreatment with L-NMMA (30-300 microM), as with the endogenous substrate L-arginine (10 microM-10 mM), inhibited in a concentration-dependent manner the ability of L-NOARG (30 microM) to block acetylcholine-induced relaxation. 5. The ability of L-NOARG to augment phenylephrine-induced tone and inhibit relaxation by acetylcholine and ATP in endothelium-containing rings is consistent with blockade of basal and agonist-stimulated production of nitric oxide, respectively. 6. The ability of L-NMMA to augment phenylephrine-induced tone without affecting relaxation to acetylcholine or ATP in endothelium-containing rings suggests a selective ability to block basal but not agonist-stimulated production of nitric oxide in rat aorta.