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Many women with typical anginal chest pain have normal coronary angiograms, which may be due to altered endothelial function. We evaluated the endothelial markers cyclic GMP (cGMP) and immunoreactive endothelin (ir-ET) regarding presence of coronary atherosclerosis in women with clinical signs of unstable coronary artery disease (CAD). Plasma levels of cGMP and ir-ET were determined in 118 patients and 84 controls. Ischaemia was evaluated at an exercise test. Of the patients 20% had normal vessels, 14% insignificant CAD and 66%, significant stenosis at coronary angiography. Mean (95% CI) concentration of cGMP (nmol/l) was higher in patients than in controls (5.05 (4.53; 5.58) vs. 3.79 (3.34; 4.23)). Separating patients according to daily intake of nitroglycerin, only patients with this medication had significantly higher cGMP level (5.73 (4.88; 6.58)), whereas the difference between those without (4.35 (3.76; 4.94)) and controls disappeared. Patients with ischaemia at exercise test had higher cGMP level than those without (6.01 (5.13; 6.88) vs. 4.30 (3.66; 4.94)), even after adjusting for nitroglycerin treatment. ir-ET (pmol/l) was lower in patients with normal vessels than patients with coronary atherosclerosis (0.83 (0.78; 0.88) vs. 0.98 (0.92; 1.04)) and than the control group (0.91 (0.87; 0.94)). The difference between the control group and patients with atherosclerosis was also significant. Patients with unstable CAD and long-term nitroglycerin treatment have increased cGMP level. Patients with exercise-induced ischaemia have higher cGMP level than those without, irrespective of nitroglycerin treatment, which may reflect a general compensatory mechanism. Patients with normal vessels have low level of ir-ET, indicating different mechanisms for ischaemia/angina in these patients compared with patients with atherosclerosis.

In order to study the relations between plasma cyclic guanosine monophosphate (cGMP) and anthropometric variables, plasma atrial natriuretic factor (ANP) and serum angiotensin-converting enzyme activity (ACE), a population study with random selection of 217 subjects (49% men) 18-69 years old was performed. Venous blood was drawn at 08.00 h and ambulatory blood pressure (ABP) recording was then performed. There was no gender-difference in cGMP levels (men 2.2 +/- 1.1 nmol L-1, women 2.2 +/- 1.1 nmol L-1). A weak negative correlation was seen between cGMP and systolic ABP in both genders 20-44 years of age (r = -0.3, p = 0.05 for both), but this correlation did not persist in multivariate analysis with correction for ANP, age and ACE. In women, cGMP correlated positively with ANP (r = 0.27, p = 0.005) independently of ANP, ACE, ABP and age. ACE correlated negatively with cGMP in men (all; r = -0.22, p = 0.03, 45-69 years of age r = -0.49, p = 0.0002) and this correlation was independent of ANP, ACE, ABP and age. ACE catalyses the breakdown of bradykinin, which stimulates the release of NO. As the second messenger of NO is cGMP, the negative correlation between ACE and cGMP in men might be a reflection of reduced production of NO. Our results also suggest that there are gender differences in the stimuli for basal cGMP production.

Objective: To investigate influence of whole-body cooling on cyclic GMP (cGMP) in women with Raynaud's phenomenon and in healthy women., Design: The study was performed as an open, parallel-group comparison between women with Raynaud's phenomenon and healthy women during the winter month of February., Setting: The municipality of Västerås (Sweden)., Participants: The Raynaud group comprised 24 female patients. The control group consisted of 21 healthy females., Main Outcome Measure: The venous levels of cGMP were measured on three different occasions: just before and after 40 min of whole-body cooling and after 20 min rest at room temperature (21 degrees C)., Results: Venous cGMP increased significantly in the control group after cold exposure (mean difference 0.43 pmol mL-1; 95% CI, 0.018-0.848; t = 2.18; df = 20; P = 0.02) and remained at a high level after 20 min rest (mean difference 0.58 pmol mL-1; 95% CI, 0.063-1.108; t = 2.34; df = 20; P = 0.015). In contrast, the levels of venous cGMP in the Raynaud group did not change significantly. The difference in increase between the two groups was significant (P < 0.02). The diastolic blood pressure in the Raynaud group increased after 40 min of whole-body cooling and was still significantly increased (P < 0.001) after 20 min rest at room temperature (21 degrees C)., Conclusion: These results indicate that women suffering from Raynaud's phenomenon lack the physiological response of cGMP to cold exposure, which may explain their increased vasospastic response.

The present study compares the tissue distribution of glyceryl trinitrate (GTN) in plasma, heart, brain, aortic tissue, and adipose tissue from GTN tolerant and GTN nontolerant rats at various time points. Furthermore, the cGMP levels in brain, heart, and aortic tissue were studied at various time points as well as the concentration-effect relationship for GTN in aorta isolated at different time points after the last exposure to GTN. Concentrations of GTN were found to be higher in all tissues studied as compared with plasma, and the concentrations of GTN were higher in tissues from tolerant rats as compared with nontolerant rats, except for aortic tissue. Concentration-effect curves obtained in vitro showed that aortic smooth muscle was still tolerant 24 h after the last dose of GTN. The cGMP level in brain was significantly increased by 40% 2 h after a single dose of GTN (50 mg/kg) and in aortic tissue by 50% at 15 min and at 2 h after a single dose of GTN (50 mg/kg). There was no effect on cGMP in brain, while an increase was seen in aortic tissue 15 min after the last dose in tolerant animals. No change in cGMP level was seen in heart neither in nontolerant nor in tolerant animals at 15 min and at 2 h. No effect on cGMP levels in brain, heart, and aortic tissue was seen 8, 16, and 24 h after exposure to GTN in either tolerant or nontolerant rats. In conclusion, GTN does not involve the cGMP system in heart, and tolerance development caused a less pronounced GTN-induced cGMP increase in aortic tissue.(ABSTRACT TRUNCATED AT 250 WORDS)

Electrical field stimulation (EFS) of phenylephrine-contracted bovine mesenteric arteries pretreated with guanethidine elicited a relaxation that amounted to roughly 40%. This relaxation was sensitive to tetrodotoxin pretreatment, suggesting a neurogenic origin. The EFS-induced relaxation was correlated to an increase in cGMP level, from 14.2 +/- 2.5 pmol/g wet wt in nonstimulated arteries to 31.6 +/- 3.4 pmol/g wet wt after 1 minute of EFS. cAMP values were not affected by EFS. Methylene blue (5 microM) and the compound LY 83583 (10 microM), inhibitors of soluble guanylate cyclase, inhibited the EFS-induced relaxation by 60% and 50%, respectively. Zaprinast (1 microM), a selective inhibitor of cGMP degradation, significantly (p = 0.005) potentiated the EFS-induced relaxation. The relaxation induced by EFS in bovine mesenteric arteries exhibits characteristics similar to the relaxations evoked by organic nitroesters and endothelium-dependent vasodilators, both of which are suggested to be mediated by cGMP and probably with nitric oxide as the common activator of the cGMP system. The possible involvement of nitric oxide as a mediator of EFS-induced relaxations was investigated with the use of known modulators of endogenous nitric oxide production. Preincubation of the arteries with 1 mM arginine or 1 mM N-alpha-benzoyl-L-arginine, both reported to potentiate endogenous nitric oxide production, or 5 mM L-canavanine, 0.25 mM NG-monomethyl-L-arginine, or 0.1 mM NG-nitro-L-arginine, alleged inhibitors of endogenous nitric oxide production, were without effect on the relaxation induced by EFS. However, pyrogallol, a generator of superoxide anions, was a potent inhibitor of relaxations induced by EFS in bovine mesenteric arteries.(ABSTRACT TRUNCATED AT 250 WORDS)

It was recently observed that the relaxation induced by glyceryl trinitrate (GTN) showed a biphasic concentration-response curve; a high-sensitivity component represented by concentrations less than 1 nM and a low-sensitivity component represented by concentrations greater than 1 nM. The effect of two glyceryl trinitrate concentrations (0.1 nM and 1 microM) were tested on the uptake of 45Ca2+ to tissue pieces of bovine mesenteric arteries (BMA) as well as on the uptake of 45Ca2+ to a microsomal preparation of BMA. The effect of GTN and 8-Br-cGMP was also studied on the IP3-induced release of Ca2+ from the microsomal preparation preloaded with 45Ca2+. The influence of IP3 and GTN on the activity of Ca2(+)-ATPase in the microsomal preparation was tested as well. The phenylephrine-stimulated uptake of Ca2+ to tissue pieces of BMA was significantly reduced by the high GTN-concentration (1 microM) but not by the lower concentration. The uptake of Ca2+ to the microsomal preparation was significantly stimulated by the two GTN-concentrations tested, as well as by 8-Br-cGMP (0.1 mM). The calcium release induced by IP3 (1 microM) from the microsomal preparation was inhibited by both the low and the high GTN-concentration and by 8-Br-cGMP (0.1 mM). The Ca2(+)-ATPase activity was stimulated by both GTN-concentrations tested while it was inhibited by IP3. It is concluded that GTN is able to induce a reduction of the free intracellular Ca2+ by several mechanisms, which are of importance for the relaxation represented by the high-affinity component. The low-affinity component in addition reduces the inflow of Ca2+ over the plasma membrane.

The inhibitory nucleotide-regulatory protein (Gl) has been shown to lose its adenylate cyclase inhibitory effect upon treatment with pertussis toxin. To find out whether a pertussis sensitive mechanism is involved in the regulation of the cGMP-system, bovine mesenteric arteries were incubated in buffer containing pertussis toxin, and the relaxation and intracellular cGMP accumulation induced by different groups of vasodilating agents were studied. The present results show a pertussis toxin induced decrease in relaxation as well as a decrease in the cGMP-elevation induced by the endothelium dependent vasodilators acetylcholine and calcium ionophore A 23187. Arteries treated with atrial natriuretic peptide showed no alterations in relaxation or cGMP content after incubation with pertussis toxin. A 40 kD soluble ribosylation substrate for pertussis toxin was identified in bovine mesenteric artery. These results suggest that a pertussis toxin sensitive mechanism is involved in the vasodilating mechanism of acetylcholine and calcium ionophore A 23187, while no evidence for such a mechanism could be found regarding the vasodilatory action of atrial natriuretic peptide.

Human peripheral vein (v. saphena magna) was exposed in vitro to glyceryl trinitrate (GTN) 0.1 mM for 1 h. The subsequent relaxant effect of GTN (10nM-0.1 mM) on vessels contracted by serotonin (0.25 microM) was significantly reduced on vessels preexposed to GTN as compared to control vessels, indicating a development of partial tolerance as far as the vascular smooth muscle relaxant effect is concerned. The impaired relaxant effect was paralleled by a reduced increment of intracellular cyclic GMP. This is, in turn, probably a consequence of both a diminished guanylate cyclase activity and an elevated phosphodiesterase activity in the GTN-tolerant vessels. The relaxant activity as well as the level of intracellular cyclic GMP were restored in GTN-tolerant vessels by dipyridamole (5 microM), an agent with phosphodiesterase inhibiting properties.

The effect of pertussis toxin (PTX) and the cyclic GMP-lowering agent LY83583 on the relaxatory response induced by glyceryl trinitrate (GTN), isosorbide dinitrate (ISDN) and isosorbide-5-mononitrate (ISMN) in bovine mesenteric artery was investigated. Pretreatment with PTX (100 ng/ml; 2h) induced a 100-fold right shift of the concentration-effect curve for GTN, while no effect on the relaxatory response elicited by ISDN and ISMN was seen 10 microM LY83583 markedly reduced the relaxatory effect of all the organic nitroesters. Based on the different sensitivity towards PTX it is suggested that GTN induces vascular smooth muscle relaxation by a partly different mechanism than ISDN and ISMN. However, cGMP seems to play a crucial role in mediating the relaxatory response of all the tested organic nitroesters since LY83583 profoundly suppressed the relaxatory response.

In the present study the possible existence of cross-tolerance between GTN, atrial natriuretic peptide and the endothelium-dependent vasodilators acetylcholine and calcium ionophore A23187 was investigated. Pretreatment of bovine mesenteric arteries (BMA) with GTN (0.44 mM) for 2 hrs caused a significant (P less than 0.001) right-shift of the concentration-effect curve for GTN as compared to controls (pD2 = 6.96 +/- 0.208 and 4.57 +/- 0.148 in controls and GTN treated vessels, respectively), thus indicating the development of tolerance. Cross-tolerance between GTN and the endothelium-dependent vasodilators acetylcholine and calcium ionophore A23187 was found as judged from cumulative concentration-effect curves. The cGMP response evoked by 1 microM GTN was markedly blunted in the GTN-pretreated vessels (P = 0.0056). Similarly the cGMP elevation induced by 10 microM acetylcholine and 0.1 microM calcium ionophore A 23187 was significantly reduced in GTN-pretreated muscle specimens (P = 0.0475 and P = 0.0103, respectively). No cross-tolerance between GTN and atrial natriuretic peptide (ANP) could be established from tension studies. Furthermore, the cGMP response induced by atrial natriuretic peptide was not significantly different in GTN-tolerant vessels as compared to in controls (P = 0.2097). BMA specimens were also incubated with acetylcholine and ANP (2 hrs and 6 hrs, respectively) and the vessels were thereafter tested for their responsiveness to acetylcholine, ANP and GTN. Preincubation with acetylcholine (10 microM) for 2 hrs did not affect the relaxant response to a subsequent challenge with acetylcholine or GTN. Preincubation of BMA with ANP (0.1 microM) for 6 hrs was also without effect on the responsiveness to ANP or GTN.(ABSTRACT TRUNCATED AT 250 WORDS)

Plasma levels of cyclic nucleotides were determined by radioimmunoassay in patients with (1) angina-like chest pain and normal coronary arteries (suspected spasm angina), (2) exercise-induced angina, and (3) heart diseases other than angina pectoris, as well as in (4) normal subjects. The concentration of cyclic GMP in plasma was significantly lower (by at least three-fold) in patients with suspected spasm angina, as compared with the other groups. No statistically significant difference in the plasma levels of cAMP was observed between the different patient groups. The low cGMP levels in plasma from patients with angina-like chest pain and normal coronary arteries might be an indication of a defect in the vasculature, making it more sensitive to contractile stimuli.

Strips of bovine mesenteric artery mounted in disposable organ baths made of polyethylene showed a biphasic relaxation pattern when exposed to glyceryl trinitrate (GTN). The concentration response curve could be resolved into a high affinity component (pD2 11.9) and a low affinity component (pD2 7.5) by means of non-linear regression analysis. The relaxation induced by both low (0.01 nM - 0.1 nM) and high (1 microM) concentrations of GTN seemed to be mediated by cyclic GMP. We found a 2-3-fold increase in cGMP at 0.01 - 0.1 nM GTN and a 5-fold increase at 1 microM GTN. Cyclic AMP levels were unchanged. We also found that GTN-induced relaxation was increased, for a given GTN concentration, when the endothelium was removed, especially in the low concentration range.

Bovine mesenteric arteries were exposed to glyceryl trinitrate (GTN) in vitro for 2 and 8 h in a low concentration (0.1 nmol/l), that equals the plasma concentrations reached by using GTN-discs in patients with angina pectoris. After reexposure to GTN a significantly reduced relaxant response was observed. In the concentration range 1 pmol/l to 10 nmol/l a 10-fold right shift of the concentration-effect curve was noted as compared to control vessels not previously exposed to GTN. The reduced relaxant response was accompanied by a significantly impaired ability to form cyclic guanosine-3',5'-monophosphate (cGMP). The results are interpreted as a development of tolerance towards GTN at the level of the vascular smooth muscle cell. A concentration of GTN (0.1 mmol/l) previously used to induce tolerance in vitro, produced a much more pronounced shift to the right of the concentration-response curve, especially in the concentration range 10 nmol/l to 0.1 mmol/l. This might indicate that tolerance to GTN can be induced by 2 different mechanisms, which in turn might be interpreted as the existence of 2 mechanisms of action by GTN in inducing vascular smooth muscle relaxation.

The relaxant effect of glyceryltrinitrate (GTN) on human vena saphena magna was studied in vitro. Vessels contracted by serotonin (0.25 microM) and phenylephrine (0.1 mM) were relaxed to the same extent (EC50 = 10 microM) by GTN, whereas in 100 mM K+-depolarized vessels the relaxation was significantly lower. The relaxant effect produced by GTN was preceded by an elevation of cyclic guanosine-3',5'-monophosphate (cGMP). For 0.1 mM GTN there was a 3-fold increase in cGMP after 3 min. A correlation between relaxation and increase in cGMP was established. When GTN was combined with dipyridamole (5 microM) the relaxant effect of GTN was significantly greater (EC50 = 0.1 microM). Phosphodiesterase inhibition, as a possible mechanism behind the observed better relaxation for the combination (GTN+dipyridamole), is briefly discussed. In conclusion, the relaxant effect of GTN on isolated human vena saphena magna seems to be dependent on the contractile stimuli used, increased by the addition of DIP and to be mediated via cGMP.

In order to study distribution in tissue, rats were treated subcutaneously with glyceryl trinitrate, GTN, (50 mg/kg). The concentrations of GTN were measured in plasma, brain, heart, adipose tissue and aortic tissue at different sampling times by a gas chromatographic method with electron-capture detection. The peak GTN-concentration was reached after 2 hours in all tissues examined. The highest concentration of GTN was found in adipose tissue, where the level was approximately forty times higher than in plasma. The concentration of GTN in brain, heart and aortic tissue was about 2-3 times as high as in plasma. The cGMP level was measured in heart and brain. An increase of the cGMP level was found in brain 2 hours after GTN administration. No cGMP increase was found in heart tissue. The results indicate a substantial distribution of GTN to tissue, and an increase of cGMP in brain. The distribution of the substance in the tissues as shown, might have both pharmacokinetic and pharmacodynamic implications.