Rolf G. G. Andersson

Tolerance towards nitroglycerin, induced in vivo, is correlated to a reduced cGMP response and an alteration in cGMP turnover.

Abstract Rats were made tolerant towards nitroglycerin (GTN) by subcutaneous injections of 50 mg/kg GTN, 3 times daily for 3 consecutive days. The effects of a test dose of GTN on tension and on the cGMP response were studied in aortic preparations. The activities of the enzymes regulating cGMP turnover were also investigated. In noradrenaline (2.5 μM)-contracted tissue from GTN-treated animals, the relaxant response towards the test dose of GTN (44 μM) was reduced by about 75% as compared to the ethanol-treated control rats. The cGMP-elevating action of GTN was reduced by 55%, while no significant effect on the cAMP level could be detected. The cyclic GMP-phosphodiesterase (G-PDE) activity was increased from 770 pmol/min × mg prot (ethanol-pretreated rats) to 1340 pmol/min × mg prot (GTN-pretreated animals). The guanylate cyclase activity, stimulated with 1 mM nitroprusside, was determined with both MnGTP and MgGTP as substrate and found to be reduced by about 75% in aortic homogenates from the GTN-pretreated animals. A slight depression in cGMP-dependent protein kinase activity was detected in aortas from GTN-treated animals. However, this depression seemed to be due to an increased breakdown of the activator (cGMP) since the inclusion of 2.5 mM theophylline in the assay solution abolished the difference. These results strongly support the suggestion that cGMP acts as a mediator of GTN-induced vascular smooth muscle relaxation. The tolerance towards the pharmacological action of GTN after repeated exposure could well be explained by the reduced formation and increased rate of breakdown of cGMP as demonstrated in this study.

Relationship between nitroglycerin, cyclic GMP and relaxation of vascular smooth muscle

Abstract Nitroglycerin (NG) caused a dose dependent-relaxation of the bovine mesenteric artery with an ED 50 -value of 2.7 × 10 −8 M. The relaxant effect of NG was significantly correlated to an increase in the cGMP content of the artery. There was a significant non-linear component in the data. At moderate cGMP levels relaxation and cGMP changes were correlated. At high levels of cGMP, however, the mechanism responsible for the nitroglycerin-mediated relaxation seemed to be completely activated and a further increase in cGMP did not induce additional relaxation. The cGMP content of the preparation was not significantly changed by nitroglycerin. The cGMP increase induced by nitroglycerin preceded the relaxation. A maximal increase of cGMP was observed after 2 min and the levels subsequently declined. This decline was not accompanied by an increase in the tissue tension. It is suggested from these experiments that cGMP might cause a relaxation of the vascular smooth muscle. Furthermore, if this suggestion is true, there seems to exist a “receptor reserve” for NG with respect to its relaxing action, since an over-capacity for cGMP production is present.

β-Adrenergic receptors in human myometrium during pregnancy: Changes in the number of receptors after β-mimetic treatment☆

Abstract The concentration of available β-adrenoceptors has been studied in the myometrium of women treated with terbutaline for premature uterine contractions and in an untreated control group. Myometrial strips were taken at cesarean section from the lower uterine segment and the uterine fundus. The concentration of β-adrenoceptors was determined with a radioligand binding assay. In untreated women we found no difference in the concentration of β-adrenoceptors in the uterine fundus compared to that in the lower uterine segment. The cyclic adenosine monophosphate production after β-adrenoceptor agonist stimulation in vitro was equal in both locations. In the terbutaline-treated women, the binding site concentrations in both the fundus and lower uterine segment were lower compared to those in the control group. The decrease was most pronounced in the fundus where receptor concentration was only half that found in the control group. The results suggest that treatment with β-mimetics causes a desensitization in the β-adrenoceptor system of human myometrium during pregnancy. This desensitization may partly explain the limited duration of the relaxant effect of β-mimetics often seen in the treatment of preterm labor

Prejunctional alpha2 adrenoceptors inhibit contraction of tracheal smooth muscle by inhibiting cholinergic neurotransmission

Abstract Ring preparations obtained from the guinea pig trachea contracted on short trains of electrical field stimulation. These contractions were mediated by activation of cholinergic nerves since they were abolished by atropine or tetrodotoxin. In the presence of beta blocking drugs noradrenaline and adrenaline dose-dependently inhibited contractions induced by field stimulation. By contrast, contractions on exogenous acetylcholine were left completely unaffected. It is concluded that the adrenergic agonists inhibited cholinergic neurotransmission by a prejunctional action. In order to characterize the noradrenaline receptor the effects of alpha 1 and alpha 2 blockers were evaluated using the Schild plot. For comparison experiments were also conducted on the guinea pig aorta and electrically stimulated guinea pig ileum. The results indicate that in guinea pig trachea and ileum noradrenaline inhibits cholinergic neurotransmission by acting on prejunctional alpha 2 receptors whereas in guinea pig aorta it induces contraction by stimulating alpha 1 receptors.

Tea flavanols inhibit angiotensin-converting enzyme activity and increase nitric oxide production in human endothelial cells

A diversity of pharmacological effects on the cardiovascular system have been reported for Camellia sinensis: antioxidative, antiproliferative and anti‐angiogenic activity, and nitric oxide synthase activation. The purpose of this study was to investigate if the connection between tea and angiotensin‐converting enzyme (ACE) and nitric oxide (NO) might be an explanation of the pharmacological effects of tea on the cardiovascular system. Cultured endothelial cells from human umbilical veins (HUVEC) were incubated with extracts of Japanese Sencha (green tea), Indian Assam Broken Orange Pekoe (black tea) and Rooibos tea, respectively. The main flavanols and purine alkaloids in green and black tea were examined for their effects on ACE and NO. After incubation with green tea, black tea and Rooibos tea for 10 min, a significant and dose‐dependent inhibition of ACE activity in HUVEC was seen with the green tea and the black tea. No significant effect on ACE was seen with the Rooibos tea. After 10‐min incubation with (–)‐epicatechin, (–)‐epigallocatechin, (–)‐epicatechingallate and (–)‐ epigallocatechingallate, a dose‐dependent inhibition of ACE activity in HUVEC was seen for all four tea catechins. After 24‐h incubation, a significantly increased dose‐dependent effect on NO production in HUVEC was seen for the green tea, the black tea and the Rooibos tea. After 24‐h incubation with (–)‐epicatechin, (–)‐epigallocatechin, (–)‐epicatechingallate and (–)‐epigallocatechingallate, a dose‐dependent increased NO production in HUVEC was seen. In conclusion, tea extracts from C. sinensis may have the potential to prevent and protect against cardiovascular disease.

In vivo Microdialysis Estimation of Histamine in Human Skin

Microdialysis, a new bioanalytical sampling technique, enables the measurement of substances in the extracellular space. This study investigates the use of the technique in the in vivo measurement of histamine levels in human skin. Microdialysis probes are equipped at the tip with a semipermeable polycarbonate membrane which permits the passive diffusion of substances. 16 probes were inserted, via a guide, into the skin of the ventral forearm of 8 patients or volunteers. The probe was perfused at a flow of 5 microliters/min, with samples being collected at intervals of 10 min and analysed by RIA technique. The mean histamine level in the first 10-min sample following probe insertion was 39.4 nM. The mean histamine value fell with successive 10-min samples (8.8, 4.6, 2.3 nM). An equilibration period of 40 min following probe insertion is suggested for histamine studies, where provocation of the skin is to be performed. Microdialysis appears to be a promising new tool for quantitative and chronological studies of cutaneous inflammatory mediators.

Suppression by neuropeptide Y of capsaicin-sensitive sensory nerve-mediated contraction in guinea-pig airways

1 In the present study we have examined whether neuropeptide Y (NPY) interferes with non‐adrenergic, non‐cholinergic nerve‐mediated contractions and relaxations in the guinea‐pig airways. In these experiments we have used ring preparations of bronchi and trachea, incubated in the presence of atropine, propranolol and indomethacin (each 1 μm). 2 The contractile response to electrical stimulation of non‐adrenergic, non‐cholinergic nerve fibres was suppressed by NPY and NPY 13–36 in a concentration‐dependent manner, these agents having similar inhibitory potencies. NPY caused a more complete inhibition than the C‐terminal fragment. 3 NPY affected neither the basal tension nor the substance P‐evoked contraction in the bronchi and trachea and did not interfere with nerve‐mediated, non‐adrenergic relaxation in the trachea. 4 On the basis of these results, it is suggested that NPY may act on the terminals of sensory neurones in the airways to prevent antidromic, excitatory neurotransmission by inhibiting transmitter release.

Effects of albuterol enantiomers on in vitro bronchial reactivity.

Asthma is one of the most common diseases in the industrialised countries. The underlying mechanisms are complex and still not fully understood although inflammation of the airways plays an important role. There are to day several types of drugs used in the treatment of asthma such as anti-inflammatory drugs, specific antagonists for inflammatory mediators and bronchodilators. Beta-agonists are the main choice for relaxing airway constriction, however unwanted effects of beta-agonists on patients with asthma has been reported. The betaantagonists that are used for treatment of hypertension and various other conditions also is shown to be deleterious in asthmatics. In the present study we have used guinea pig airways to examine the proposed deleterious effects of beta-agonists and antagonists. We have shown that the (S)-enantiomeric forms of salbutamol and formoterol are able to potentiate cholinergic stimuli and we have shown that the potentiation was indomethacin sensitive in airway preparations of sensitised guinea pigs. We also showed and confirmed that the (R)-enantiomeric forms of salbutamol and formoterol were more potent in relaxing airway smooth muscle contracted with different stimuli compared to the (S)-enantiomers. The betaantagonists propranolol and pindolol were shown to be able to contract tracheal preparations if they had been pre-treated with a beta-agonist and the contraction was not simply a blockade of the beta-adrenoceptor induced relaxation. Propranolol contraction was stereo-selective and (S)-propranolol was more effective in inducing contraction than (R)-propranolol. Moreover, atenolol a betacselective antagonist induced significantly smaller contractions compared to general beta-antagonists. This indicates that the beta2-adrenocepor probably is involved in the beta-antagonist induced contraction. The cyclooxygenase inhibitor indomethacin, the 5-lipoxygenase inhibitor MK886 and a thromboxane A2 antagonist as well as capsaicin reduced the beta-antagonist induced contraction. This indicates that several arachidonic acid products as well as neuropeptides may be involved in the beta-antagonistinduced contraction.The worsening of asthma by beta-antagonists is well known and the risks associated with beta-agonists are discussed, but the mechanisms behind these effects need further clarification. In this thesis some of the possible mechanism have been discussed, further studies are needed in order to get more safe and effective asthma treatment regime.

Antiatherosclerotic effects of the angiotensin-converting enzyme inhibitors captopril and fosinopril in hypercholesterolemic minipigs

We evaluated the two angiotensin-converting enzyme (ACE) inhibitors captopril and fosinopril with regard to possible antiatherosclerotic effects in minipigs. Experimental hypercholesterolemia and atherosclerosis was produced in 33 minipigs of the Gottingen strain by an egg yolk/cholesterol-enriched diet for 1 year. One group (n = 11) was fed the atherogenic diet alone and served as a control. A second group (n = 11) received captopril (80 mg/kg/day) added to the atherogenic diet, and a third group (n = 11) was treated in the same manner but with fosinopril (8 mg/kg/day). The drug treatments produced significant reduction in serum ACE activity associated with a reactive increase in plasma renin activity (PRA), but had only minor effects on plasma lipids and lipoproteins. At the end of the treatment period, all animals were killed and examined for degree of atherosclerosis. The percentage of atherosclerotic area in the abdominal aorta was significantly lower in both drug-treated groups as compared with controls. Furthermore, accumulation of cholesterol in the thoracic and abdominal aorta was inhibited by drug treatment. Finally, the percentage of intimal thickening in abdominal aorta was significantly reduced in the drug-treated groups. In conclusion, the ACE inhibitors captopril and fosinopril inhibited development of atherosclerosis in hypercholesterolemic minipigs.

Effect of Vaccinium myrtillus and its polyphenols on angiotensin-converting enzyme activity in human endothelial cells.

This study investigates if the connection between Vaccinium myrtillus and angiotensin-converting enzyme (ACE) might be an explanation of the pharmacological effects on circulation. Cultured endothelial cells from human umbilical veins were incubated with bilberry 25E extract. The main anthocyanidins combined in myrtillin chloride and separately in cyanidin, delphinidin, and malvidin, respectively, were examined concerning their effects on ACE. After 10 min of incubation with bilberry 25E, a significant, dose-dependent inhibition of ACE activity was seen, and after incubation with myrtillin chloride a significant inhibition was seen. No effect was seen with the anthocyanidins. The effect seems to be dependent on this specific mixture of anthocyanins in the bilberry. V. myrtillus may thus have the potential to prevent and protect against cardiovascular diseases.