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Biomedical subjects

E Bassenge

Publications and source records attributed to E Bassenge.

At least 37 records · Page 2Linked to original sources
PubMed1998

Intrinsic neural regulation of the heart in the chronic, conscious dog.

The present experiments were performed to examine the capability of the intrinsic cardiac nerves (ICN) to modify cardiac performance in the resting chronic, conscious dog. Control and cardiac-denervated dogs were instrumented for recording of left atrial (LA) and ventricular (LV) contractility, heart rate, and atrioventricular (AV) conduction time. Acetylcholine (ACh) and nicotine (Nic) were administered via an indwelling coronary artery catheter. Limited distribution from the injection site only allowed access to the LA, LV, and AV node. Both beta-blockade with timolol and cardiac denervation were used to separate direct effects of ICN stimulation from indirect (e.g., reflex) effects. ACh produced the expected negative inotropic and dromotropic changes. ICN stimulation with Nic caused large decreases in LA on but only trivial effects on the LV. We concluded that the ICN has limited effects on cardiac performance in the resting animal under minimal sympathetic drive. It is likely, however, that the ICN is capable of significantly depressing cardiac function under conditions of elevated sympathetic tone as would be encountered in exercise.

Acetylcholine↗
PubMed1998

Formation of Reactive Oxygen Species in Various Vascular Cells During Glyceryltrinitrate Metabolism.

BACKGROUND: Anti-ischemic therapy with organic nitrates as nitric oxide (NO) donors is complicated by the induction of tolerance. When nitrates are metabolized to release NO, there is a considerable coproduction of reactive oxygen species (superoxide radical and peroxynitrite) in vessels leading to inactivation of NO, to diminished cyclic quanosine monophosphate production in smooth muscle cells (SMC), to impaired vasomotor responses to the endothelium-derived relaxation factor (EDRF), and to formation of nitrotyrosine as a marker of glyceryltrinitrate (GTN)-induced formation of peroxynitrite. The aim of the study was to analyze in vitro the formation of superoxide radicals and of peroxynitrite in GTN-treated endothelial and smooth muscle cells and in washed ex vivo platelets using electron spin resonance and spin-trapping techniques. METHODS AND RESULTS: Using 5,5-dimethyl-1-pyrroline-N-oxide (DMPO) as a spin trap, it was shown that in platelets, smooth muscle, and endothelial cells incubated acutely for 15 minutes with 0.5 mM GTN, the rate of generation of reactive oxygen species (ROS) was twice as high as under control conditions. Using the new spin-trap 2H-imidazole-1-oxide (TMIO), a GTN-induced peroxynitrite formation was detected in SMC and in platelets incubated with 0.5 mM GTN for 15 minutes. Spin-trap 1-hydroxy-3-carboxy-pyrrolidine (CP-H) was used to estimate the rate of ROS formation in platelets incubated for 15 minutes with 0.5 mM GTN; the rate amounted to 14.6 +/- 1.1 nM/min/mg protein compared with 4.0 +/- 0.4 nM/min/mg protein in controls. The rate of ROS formation in SMCs was substantially increased (240 +/- 16%) after initiation of GTN tolerance by treatment of the cells in culture with 100 µM GTN for 24 hours. CONCLUSIONS: GTN increases the formation of superoxide radicals in endothelial cells, SMCs, and platelets. Peroxynitrite is formed during GTN metabolism in vascular cells and may contribute to the development of tolerance. A decrease in the nitrate-induced inhibition of platelet aggregation during GTN tolerance is associated with oxidative actions of ROS formed in platelets during GTN metabolism.

Journal Article↗
PubMed1998

Formation of reactive oxygen species by pentaerithrityltetranitrate and glyceryl trinitrate in vitro and development of nitrate tolerance.

Anti-ischemic therapy with organic nitrates is complicated by tolerance. Induction of tolerance is incompletely understood and likely multifactorial. Recently, increased production of reactive oxygen species (ROS) has been investigated, but it has not been clear if this is a direct consequence of the organic nitrate on the vessel or an in vivo adaptation to the drugs. To examine the possibility that nitrates could directly stimulate vascular ROS production, we compared the development of nitrate tolerance with the formation of ROS induced by pentaerithrityltetranitrate (PETN) or nitroglycerin (GTN) in vitro in porcine smooth muscle cells, endothelial cells, washed ex vivo platelets and whole blood. By examining cGMP formation, it was found that 24-hr treatment with GTN but not PETN induced significant nitrate tolerance, which was prevented by parallel treatment with Vit C. Incubation of vascular cells acutely with 0.5 mM GTN doubled the rate of ROS generation, whereas PETN had no such effect. The rate of ROS (peroxynitrite and O2) formation detected by specific spin traps in tolerant smooth muscle cells, treated for 24 hr with 0.01 mM GTN, was substantially higher (30.5 nM/min) than in control cells acutely treated with 0.5 mM GTN (25 nM/min). In contrast to PETN, GTN induces nitrate tolerance and also increases the formation of ROS both in vascular cells and in whole blood. ROS formation is minimally stimulated by PETN comparable to data obtained in Vit C-suppressed GTN tolerance. ROS formation induced by organic nitrates seems to be a key factor in the development of nitrate tolerance.

Animals↗
PubMed1997

Spin trapping of superoxide radicals and peroxynitrite by 1-hydroxy-3-carboxy-pyrrolidine and 1-hydroxy-2,2,6, 6-tetramethyl-4-oxo-piperidine and the stability of corresponding nitroxyl radicals towards biological reductants.

The reactions of new spin trap 1-hydroxy-3-carboxy-pyrrolidine (CP-H) with superoxide radicals and peroxynitrite were studied. The rate constants were determined as 3.2 x 10(3) and 4.5 x 10(9) M-1s-1, respectively. It was found that 2mM of spin trap CP-H or 1-hydroxy-2,2,6,6-tetramethyl-4-oxo-piperidine (TEMPONE-H) provide almost the same spin trapping efficacy. In contrast to TEMPONE-H the reaction of CP-H with peroxynitrite was inhibited by 20 mM DMSO. This simplifies the quantification of peroxynitrite formation. During the reaction of CP-H and TEMPONE-H with superoxide radicals or peroxynitrite the stable nitroxide radicals 3-carboxy-proxyl (CP) and 2,2,6,6-tetramethyl-4-oxo-piperidinoxyl (TEMPONE) are formed. It was found that the rate of reduction of CP by glutathione or by smooth muscle cells was two-fold slower and the reduction of CP by ascorbate was 66-fold slower than corresponding rates of reduction of TEMPONE. Therefore quantification of the formation of superoxide radicals and of peroxynitrite by CP-H is much less hindered by a variety of biological reductants than in case of TEMPONE-H. Thus, CP-H is more suitable for spin trapping of superoxide radicals and peroxynitrite in biological systems than the TEMPONE-H.

Animals↗
PubMed1997

Quantification of peroxynitrite, superoxide, and peroxyl radicals by a new spin trap hydroxylamine 1-hydroxy-2,2,6,6-tetramethyl-4-oxo-piperidine.

The reactions of hydroxylamine 1-hydroxy-2,2,6,6-tetramethyl-4-oxo-piperidine hydrochloride (TEMPONE-H) with peroxynitrite, superoxide and peroxyl radicals were studied. It was shown that under these reactions TEMPONE-H is oxidized into a stable nitroxide 1-hydroxy-2,2,6,6-tetramethyl-4-oxo-piperidi-noxyl (TEMPONE). The reactivity of TEMPONE-H towards reactive oxygen species was compared with the spin traps DMPO and TMIO as well as with DMSO and SOD. The rate constants of reactions of TEMPONE-H with peroxynitrite and superoxide radicals were 6 x 10(9) M(-1)s(-1) and 1.2x10(4) M(-1)s(-1), respectively. Using TEMPONE-H the sensitivity in the detection of peroxynitrite or superoxide radical was about 10-fold higher than using the spin traps DMPO or TMIO. Thus, TEMPONE-H may be used as a spin trap in chemical and biological systems to quantify peroxynitrite and superoxide radical formation.

Amidines↗
PubMed1997

Quantification of superoxide radicals and peroxynitrite in vascular cells using oxidation of sterically hindered hydroxylamines and electron spin resonance.

The reactions of two hydroxylamines, 1-hydroxy-3-carboxy-pyrrolidine (CP-H) and 1-hydroxy-2,2,6,6-tetramethyl-4-oxo-piperidine (TEMPONE-H), with superoxide radicals and peroxynitrite were studied. In these reactions corresponding stable nitroxyl radicals 3-carboxy-proxyl (CP) and 1-hydroxy-2,2,6,6-tetramethyl-4-oxopiperidinoxyl (TEMPONE) are formed and the amount of them can be quantified by electron spin resonance (ESR). It was found that CP-H and TEMPONE-H provide almost the same efficacy in assaying peroxynitrite by ESR in vitro at pH 7.4. The formation of superoxide radicals in suspensions of cells was discriminated from that of peroxynitrite using superoxide dismutase or dimethyl sulfoxide as competitive reagents. The stability of the radicals CP and TEMPONE in the presence of ascorbate or thiols was studied in vitro. The reduction rate of CP by ascorbate was 66-fold slower than the rate of reduction of TEMPONE. Therefore, the quantification of the formation of superoxide radicals and of peroxynitrite is much less affected by ascorbic acid when CP-H, but not TEMPONE-H, is used. Both TEMPONE-H and CP-H were used to determine the formation rates of superoxide radicals and peroxynitrite in suspensions of cultured aortic smooth muscle cells and endothelial cells, in washed ex vivo platelets, and in blood treated with glycerol trinitrate (GTN) as an NO donor. It was shown that both the acute addition of GTN (0.5 mM) to vascular cells and the incubation of smooth muscle or endothelial cells in culture with 0.1 mM GTN for 24 h enhance significantly the formation of reactive oxygen species in cells. The rates of of superoxide radical formation were increased at least in two times and peroxynitrite was detected. Hydroxylamines TEMPONE-H and CP-H can be used as nontoxic compounds in ESR assay capable of quantifying the formation of superoxide radicals and peroxynitrite in suspensions of cells and in the whole blood with high sensitivity.

Analysis of Variance↗
PubMed1997

Unexpected, tolerance-devoid vasomotor and platelet actions of pentaerythrityl tetranitrate.

Efficacy of nitrate therapy is limited by tolerance. A surprising upregulation of ex vivo platelet activity, a decrease in platelet thiol levels, and an enhanced release of vasoconstrictors from platelets is associated with enhanced superoxide-mediated oxidant stress leading to vascular tolerance to nitrates. We tested the NO-donor pentaerythrityl tetranitrate (PETN), which to date had not been precisely tested either with regard to the induction of tolerance or to a potential development of changes in platelet activity in comparison with glycerol trinitrate (GTN). Long-term instrumented dogs nonintermittently received: 1.5 microg/kg/min GTN, i.v., with or without vitamin C (55 microg/kg/min, i.v.) or PETN 4 x 60 mg/day orally for 5 days. Tested daily were (a) the dilation of the epicardial arteries, (b) thrombin-induced (0.5 U/ml) increases of the intracellular Ca2+ concentration and aggregability of platelets, (c) concentrations of reduced low-molecular-weight thiols (LMTs) in plasma and platelets, and (d) formation of reactive oxygen species (ROSs). During nonintermittent PETN and during GTN with additional vitamin C, a 9.8 +/- 0.4% coronary artery dilation was observed in contrast to that with GTN alone, which resulted in complete tolerance at day 4. This vascular tolerance was associated with enhanced platelet activity and formation of ROSs (incubated platelets) and a 38 +/- 3% reduction in LMT. These unfavorable changes were absent in the presence of PETN or with additional vitamin C as an antioxidant. Vascular tolerance associated with platelet upregulation is avoided either by nonintermittent nitroglycerin (5 days) when vitamin C is coadministered or by pentaerythrityl tetranitrate without the coadministration of vitamin C.

Animals↗
PubMed1997

Effect of carbon dioxide-enriched water and fresh water on the cutaneous microcirculation and oxygen tension in the skin of the foot.

The effects of immersion of the lower leg and foot in fresh water and in carbon dioxide (CO2)-enriched water (1200 mg CO2 per kg water; succinate+sodium bicarbonate: Actibath, KAO Tokyo) on cutaneous circulation, vasomotion and oxygen tension (PO2) were measured by laser Doppler flowmetry and transcutaneous oximetry. On the first of two consecutive days patients were randomly assigned to have the lower extremities immersed in either fresh water or CO2-enriched water under standardized conditions (temperature, 34 degrees C; depth, 35 cm; immersion time, twenty minutes) with concurrent measurement. On the second day patients were switched to the other bath type. For both sets of measurements probes were attached symmetrically to the dorsum of each foot. Included in the study were 18 patients with mild, bilateral, peripheral, occlusive arterial disease (intermittent claudication, femoral or iliac type). During immersion in CO2-enriched water the Doppler laser signal and vasomotion amplitude rose by 300%, while PO2 increased by 10%. These increases were still apparent during the latter part of the measurement period, following withdrawal of the limbs from the bath, while patients were seated and supine. During immersion in fresh water and thereafter the Doppler laser signal was unchanged and the PO2 increase was considerably less marked. The authors were thus able to demonstrate vasodilation and increased oxygen utilization (Bohr effect) resulting from topical CO2 application, and hence, that the use of topical CO2 has an objective basis.

Aged↗
PubMed1997

Effects of serial percutaneous application of carbon dioxide in intermittent claudication: results of a controlled trial.

In a prospective, controlled clinical trial, serial application of carbon-dioxide-enriched water was compared with fresh water. Twenty-four patients with peripheral arterial occlusive disease (stable claudication) were randomly allocated to one of two serial intervention groups, lower extremities immersed in either fresh water or in CO2-enriched water (1000 mg CO2/kg) water under standardized conditions (temperature, 33 degrees C; depth, 40 cm; immersion time, 30 min; five times a week over 4 weeks). The serial application of carbon-dioxide-enriched water increased arterial peak flow (reactive hyperemia), transcutaneous oxygen tension (basal value and half-recovery-time), and pain-free walking distance. The serial fresh water application did not change these values. The authors conclude that serial carbon dioxide application can be clinically effective in patients with arterial obstructions in the lower extremities.

Adult↗
PubMed1997

Urinary NItrotyrosine Content as a Marker of Peroxynitrite-induced Tolerance to Organic NItrates.

BACKGROUND: Anti-ischemic therapy with nitrovaasodilators as NO-donors is complicated by the induction of tolerance. When nitrovasodilators are metabolized to release NO there is a considerable coproduction of oxygen-derived radicals leading to a diminished cyclic GMP production and to impaired vasomotory responses. We analyzed in vivo the glyceroltrinitrate-induced generation of strong oxidative/nitrating compounds contributing to development of tolerance. METHODS AND RESULTS: In 16 patients we studied the urinary nitrotyrosine excretion during either (1) placebo control conditions, (2) 2-day nonintermittent transdermal nitroglycerin administration (0.4 mg/h), (3) 2-day nonintermittent glyceroltrinitrate administration (0.4 mg/h) along with a continuous infusion of vitamin C (55 µg/kg/min) as an antioxidant, or (4) with vitamin C but without glyceroltrinitrate (diminished urinary nitrotyrosine content of 34 +/- 18 µg/day observed). Glyceroltrinitrate administration augmented urinary nitrotyrosine from 56 +/- 24 (basal) to 186 +/- 32 µg/day (glyceroltrinitrate tolerance). Coadministration of vitamin C caused complete elimination of tolerance and a decrease in urinary nitrotyrosine to 130 +/- 28 µg/day. Glyceroltrinitrate-induced formation of oxidants was confirmed in vitro comparing glyceroltrinitrate-induced and peroxynitrite-induced tachyphylaxis in isolated perfused rabbit hearts and analyzing tolerance-induced inactivation of solbule guanylyl cyclase in cultured aortic smooth muscle cells. CONCLUSIONS: Augmented urinary nitrotyrosine excretion during glyceroltrinitrate administration reflects enhanced formation of peroxynitrite and of nitrotyrosine. Glyceroltrinitrate-induced tolerance is the result of oxidative stress and can be suppressed by additional antioxidant therapy aimed to prevent glyceroltrinitrate-induced formation and/or actions of peroxynitrite.

Journal Article↗
PubMed1996

EPR and laser flash photolysis studies of the reaction of nitric oxide with water soluble NO trap Fe(II)-proline-dithiocarbamate complex.

Diethyl-dithiocarbamate (DETC) chelating ferrous iron is used during EPR detection of nitric oxide (.NO) as a precursor of spin traps for the quantification of released .NO both in vitro and in vivo. This method is based on the high affinity of water insoluble iron-dithiocarbamate complexes towards .NO. The nitrosyl-iron-dithiocarbamate complex NOFe(DETC)2 is formed with characteristic triplet EPR spectrum. In the present paper we have studied the reaction of water soluble Fe(II)-proline-dithiocarbamate complex, Fe(PDTC)2, with nitric oxide using both EPR spectroscopy and laser flash photolysis. It has been found that the treatment of the Fe(PDTC)2 complex either by authentic .NO dissolved in aqua buffer solutions or by different .NO donors results in the quantitative appearance of an EPR signal of a stable mononitrosyl complex NOFe(PDTC)2 with a g-factor giso = 2.04 and a hyperfine splitting constant aN = 12.5 G. The laser flash photolysis method has been used to study the kinetics of .NO trapping by Fe(PDTC)2. Nitric oxide release from a light sensitive .NO donor, piridazinofuroxan, was induced by a short flash of UV light generated by a XeCl eximer laser. The rate constant of the reaction of .NO with Fe(PDTC)2 was found to be (1.1 +/- 0.3).10(8) M-1s-1, manifesting a high efficiency of .NO trapping by Fe(PDTC)2.

Electron Spin Resonance Spectroscopy↗
PubMed1996

Long-term angiotensin-converting enzyme inhibition with high-dose enalapril retards nitrate tolerance in large epicardial arteries and prevents rebound coronary vasoconstriction in vivo.

BACKGROUND: Rebound myocardial ischemia develops in patients with unstable or stable angina pectoris after sudden cessation of nitroglycerin therapy. Long-term nitroglycerin infusion is associated with increases in plasma renin activity and catecholamine release rates, both of which may lead to excess angiotensin II and alpha-adrenergic-mediated vasoconstriction, particularly on withdrawal of nitroglycerin. METHODS AND RESULTS: Chronically instrumented dogs were treated for 5 days with nitroglycerin (1.5 micrograms.kg-1.min-1 i.v.) alone or in combination with the angiotensin-converting enzyme (ACE) inhibitor enalapril (0.1 mg/kg two times daily or 1 mg/kg). With long-term nitroglycerin therapy, the left anterior circumflex artery was maximally dilated 4 hours after the start of nitroglycerin infusion (9.5 +/- 0.6%) and returned to baseline levels within the third day of treatment (baseline, 2.52 +/- 0.07 mm; day 3, 2.55 +/- 0.07 mm; P = NS), indicating a complete loss of nitroglycerin-induced coronary vasodilatation. Nitroglycerin infusion also was accompanied by a transient increase in plasma renin activity. Sudden withdrawal of nitroglycerin infusion caused a progressive constriction of the left anterior circumflex artery, which peaked 4 hours after nitroglycerin infusion cessation (-7.8 +/- 0.2%). This occurred in the absence of elevated plasma renin activity. Concomitant treatment with high-dose enalapril (1 mg.kg-1.d-1) markedly reduced the degree of tolerance and prevented the rebound constriction on cessation of nitroglycerin therapy. CONCLUSIONS: Long-term ACE inhibition with high-dose enalapril reduces nitroglycerin tolerance and prevents rebound vasoconstriction in coronary arteries. These phenomena were not associated with an activated circulating renin-angiotensin system. This observation suggests that during long-term nitroglycerin treatment, intrinsic abnormalities of the vascular smooth muscle may have developed that are suppressed by concomitant ACE inhibitor therapy. The present study also favors a combination of nitroglycerin and ACE inhibitors to maintain nitrate sensitivity of the vasculature during long-term nitroglycerin treatment.

Angiotensin-Converting Enzyme Inhibitors↗
PubMed1996

Tolerance to nitrates and simultaneous upregulation of platelet activity prevented by enhancing antioxidant state.

We analysed the induction of tolerance to nitrates both in the vasculature (in vivo) and platelets (ex vivo). Simultaneously, we tested mechanisms underlying the induction of tolerance and interventions to prevent or overcome this phenomenon. For this purpose nitroglycerin (GTN 1.5 micrograms/kg per min i.v.), alone or in combination with ascorbate (55 micrograms/kg per min i.v.) as antioxidant, was infused continuously for a period of 5 days into chronically instrumented dogs. Along with haemodynamic parameters, ex vivo platelet function was continuously monitored. Following the start of GTN infusions there was a maximal coronary dilator response (245 +/- 15 microm) and, as an index of venodilation, a fall of left ventricular end-diastolic pressure (by 2.3 +/- 0.4 mmHg). Both responses declined progressively and disappeared during the infusion period. However, in combination with ascorbate as antioxidant the dilator responses were maintained fully throughout the infusion period. With GTN alone there was a progressive, unexpected upregulation of platelet activity demonstrated by enhanced thrombin-stimulated intracellular Ca2+ levels and increases in the microviscosity of platelet membranes (indicating enhanced receptor expression) associated with a progressive impairment in basal, unstimulated cGMP levels. These changes could also be prevented completely by i.v. co-administration of ascorbate. From these results it is concluded that vascular tolerance is closely reflected by simultaneous changes in platelet function and further, that both can be prevented completely by appropriate antioxidants such as ascorbate.

Animals↗
PubMed1995

In vivo spin trapping of glyceryl trinitrate-derived nitric oxide in rabbit blood vessels and organs.

BACKGROUND: The objectives of this study were (1) to assess glyceryl trinitrate (GTN)-derived nitric oxide (NO) formation in vascular tissues and organs of anesthetized rabbits in vivo, (2) to establish a correlation between tissue NO levels and a biological response, and (3) to verify biotransformation of GTN to NO by cytochrome P-450. METHODS AND RESULTS: NO was trapped in tissues in vivo as a stable paramagnetic mononitrosyl-iron-diethyldithiocarbamate complex [NOFe(DETC)2]. After removal of the tissues, NO was determined by cryogenic electron spin resonance spectroscopy. NO formation in vitro was assessed by spin trapping and by activation of soluble guanylyl cyclase. The GTN-elicited decrease in coronary perfusion pressure was monitored in isolated, constant-flow perfused rabbit hearts. NO was not detected in control tissues. In GTN-treated rabbits, NO formation was higher in organs than in vascular tissues and higher in venous than in arterial vessels. In isolated hearts, ventricular NO levels and decreases in coronary perfusion pressure achieved by GTN were closely correlated. Purified cytochrome P-450 catalyzed NO formation from GTN in a P-450-NADPH reductase- and NADPH-dependent fashion. CONCLUSIONS: Since GTN-derived NO formation in myocardial tissue correlates to the GTN-elicited vasodilator response, we conclude that GTN-derived NO detected in vivo correlates with the systemic effects of GTN. Therefore, the higher rate of NO formation detected in veins compared with arteries explains the preferential venodilator activity of GTN. High NO formation in cytochrome P-450-rich organs in vivo and efficient NO formation from GTN by cytochrome P-450 in vitro highlights the importance of this pathway for NO formation from GTN in the intact organism.

Animals↗
PubMed1995

Effect of ITF 296 on total effective vascular compliance in the anesthetized dog under autonomic blockade.

The effects of ITF 296 on venous tone, estimated as changes in total effective vascular compliance (TEVC), were investigated in the anesthetized, spontaneously breathing dog under autonomic blockade. TEVC was calculated from the correlation between the observed changes in central venous pressure (CVP) and the experimentally induced changes in blood volume during an 11-min cycle of whole blood infusion (2 ml/kg/min), withdrawal, and reinfusion. Stepwise increasing doses (short steady-state infusions) of ITF 296 (3, 10, and 30 micrograms/kg/min) resulted in a dose-dependent increase in TEVC (+27, +54, and +67%), whereas mean blood pressure (MBP) was reduced (-12, -23, and -33%, respectively). SIN-1 (10 micrograms/kg/min), administered at the end of the experiment as the reference NO-releasing compound, induced a rather complete venodilation (TEVC +182%) and a marked reduction in MBP (-47%). When nitroglycerin (NTG) was used as the reference compound at 1.5 micrograms/kg/min, a 67% increase in TEVC was observed. These results suggest that ITF 296 exerts a venodilator action but that this effect, and therefore the reduction in venous return, is less than that in response to nitrovasodilators such as SIN-1 or NTG. Furthermore, the vasodilator spectrum of ITF 296 compared to other nitrovasodilators is shifted somewhat more toward the arteriolar bed, with less pronounced venodilator effects, as indicated by the fall in MBP.

Anesthesia↗
PubMed1995

Effects of ITF 296 on epicardial coronary artery diameter during acute and long-term treatment in the conscious dog.

The action of the newly developed organic nitrate ITF 296 on large coronary arteries was investigated during acute and long-term treatment in the conscious dog, chronically instrumented for recording of large coronary artery diameter (CD), mean arterial blood pressure (MBP), and heart rate (HR). Short-term steady-state infusion of ITF 296 at doses of 0.1-30 micrograms/kg/min induced a dose-dependent increase in CD. Maximal coronary artery dilation (+11%) was reached at 10 micrograms/kg/min and no further increase was observed at 30 micrograms/kg/min. At the same doses no significant variations of MBP or HR were observed. Long-term administration (5-day infusion) of ITF 296 at 20 micrograms/kg/min resulted in a 10% increase in CD, and the effect was well maintained over the entire infusion period. MBP did not change significantly, whereas HR rose by 21% by the second day of infusion and remained elevated until the end of the treatment. These results confirm that ITF 296 is an effective dilator of large coronary arteries at doses devoid of peripheral hemodynamic effects. Moreover, the vasodilating effect on coronary arteries is well maintained during long-term administration, suggesting a reduced tolerance development.

Animals↗
PubMed1994

Subcellular localization and characterization of nitric oxide synthase(s) in endothelial cells: physiological implications.

Endothelial cells (EC) contain a constitutive Ca2+/calmodulin-dependent nitric oxide (NO) synthase (cNOS) which plays an important role in the local control of vascular tone. We compared the subcellular distribution of this enzyme in cultured and freshly isolated pig EC by determination of specific cNOS activity and immunoblot analysis. Similar studies were also performed with cultured and freshly isolated bovine and cultured human EC. Enzyme activity was predominantly (> 70%) associated with the particulate fraction of all EC types tested and was highest in freshly isolated porcine EC. Both specific cNOS activity and immunoreactivity were substantially higher (> 3-fold) in the microsomal as compared with the soluble fraction of all EC types tested. In freshly isolated pig EC, these two fractions also differed in terms of their Ca(2+)-dependency, pH optimum and inhibitor specificity. EC may thus contain either two different cNOS isoenzymes or a single enzyme, the conformation of which differs between the soluble and membrane-bound state. Moreover, detailed subcellular fractionation of freshly isolated pig EC revealed that the distribution of cNOS activity closely resembled that of the plasma membrane marker 5'-nucleotidase, suggesting that most, if not all, of the cNOS activity in these cells is associated with the plasma membrane. This localization might render the enzyme more susceptible to activation by physical stimuli, such as a shear stress-induced change in the fluidity of the plasma membrane. Moreover, the continuous exposure to shear stress in vivo may also upregulate cNOS expression in EC, since specific enzyme activity, immunoreactivity and basal NO release were significantly higher in freshly isolated EC as compared with cultured EC.

Amino Acid Oxidoreductases↗