Search PubMed⌕ Search

Biomedical subjects

J C Stoclet

Publications and source records attributed to J C Stoclet.

At least 127 records · Page 7Linked to original sources
PubMed1987

Quantitative method for the study of the morphological changes induced by vasoactive agents in single aortic myocytes grown in primary cultures.

Morphological changes of aortic myocytes grown in primary cultures on collagen gels, were induced by two adrenergic agonists, in the absence or presence of specific antagonists. Their effects were quantified by means of the measurement of the surface areas of the images of the cells. The alpha-adrenoreceptor agonist phenylephrine induced a concentration-dependent decrease of the surface areas of the images of the cells, which was competitively antagonized by phentolamine, and was reduced in the presence of CdCl2 (an inhibitor of calcium entry). The beta-adrenoreceptor agonist isoproterenol induced an increase in the surface areas of the images of prostaglandin F2-alpha pretreated cells and this effect was antagonized by propranolol. It is concluded that the morphological changes of myocytes in primary culture, induced by phenylephrine or isoproterenol can be quantified by the measurement of the surface areas of their images. These changes were in good agreement with the response of isolated thoracic aorta rings or strips to the same agonists with regard to their sensitivity, specificity, and kinetic features. Thus the measurement of the changes of the surface area of the images of myocytes induced by vasoactive agents constitutes a noninvasive, nondestructive method for the quantification of their responses to these agents at the single cell level.

Animals↗
PubMed1986

Phorbol ester inhibition of vasopressin-induced calcium efflux from cultured rat aortic myocytes.

The effect of the protein kinase C activator TPA was investigated on AVP-induced 45Ca release from rat aortic myocytes. In the nanomolar range TPA, but not 4 beta-phorbol, reduced the brief 45Ca efflux produced by AVP in the presence or in the absence of extracellular calcium. The maximal effect of TPA was to abolish the response to a half maximally active concentration of AVP, and to reduce by 50% the maximal response to the hormone. These results suggest that protein kinase C activation can exert a negative control on the early AVP-induced calcium mobilization in vascular smooth muscle.

Animals↗
PubMed1986

Selective inhibition of cyclic nucleotide phosphodiesterases of human, bovine and rat aorta.

Cyclic nucleotide phosphodiesterase (PDE) activity from the 105,000 g supernatant of human, bovine and rat aorta smooth muscle cells was resolved by DEAE-trisacryl chromatography into three major forms showing similar properties in each species. In addition to the two PDE forms previously characterized in vascular tissues (a cAMP-PDE and a calmodulin-dependent PDE), a cGMP-PDE, insensitive to calmodulin, was isolated and characterized in the aorta of the three species. Each isolated PDE form was differently inhibited by various chemical compounds, and these compounds produced effects on cyclic nucleotide levels in isolated rat aorta which could be expected from their inhibitory effect on isolated PDE forms. At concentrations non-selectively inhibiting the three isolated PDE forms (including the calmodulin-dependent one), IBMX (3-isobutyl-1-methylxanthine) and trequinsin markedly and dose-dependently increased both cAMP and cGMP aorta levels (up to 7-fold, in presence of 500 microM IBMX). By contrast selective inhibitors of cGMP-PDE or cAMP-PDE could only induce a moderate elevation (by 1.5-3-fold) in cGMP or cAMP levels, respectively. In the case of M&B 22,948, a highly specific and potent inhibitor of cGMP-PDE, a concentration-dependent increase in tissue cGMP levels was produced by concentrations (in the microM range) active in inhibiting the isolated enzyme. In the case of selective cAMP-PDE inhibitors (rolipram and Ro 20-1724), however, a significant increase in aorta cAMP content was induced only in the presence of drug concentrations which were much higher (200 and 500 microM, respectively) than those inhibiting the isolated enzyme (IC50:5 and 18 microM, respectively). Inhibitors of both cGMP-PDE and cAMP-PDE (dipyridamole, cilostamide and its derivative AAL 05) produced the same moderate effects as did the combination of a selective cGMP-PDE inhibitor and a selective cAMP-PDE inhibitor on the levels of both cGMP and cAMP. These results show that the three forms of PDE isolated from aortic smooth muscle retain properties that they exhibit in the tissue and which are similar in the three species examined, including man. They suggest that each form participates in a specific manner to the regulation of cAMP and cGMP concentrations in aorta smooth muscle cells.

1-Methyl-3-isobutylxanthine↗
PubMed1986

Alpha-adrenoceptor antagonistic and calcium antagonistic effects of nicergoline in the rat isolated aorta.

The activity of the alpha-adrenoceptor antagonist nicergoline, a molecule composed of two constituent parts, ergoline and bromonicotinic acid, was investigated in the rat isolated aorta. Nicergoline (10 nM-0.1 microM) displaced concentration-effect curves elicited by noradrenaline and phenylephrine to the right and inhibited maximal responses elicited by both alpha-adrenoceptor agonists without significantly affecting prostaglandin F2 alpha-induced contractions. Higher concentrations of nicergoline (1 microM-50 microM) displaced to the right the concentration-effect curves elicited by calcium in a depolarizing medium. This calcium antagonist activity was not shared by either of the constituent parts. Nicergoline 100 microM abolished the 45Ca influx induced into rat aorta by 100 mM K+-containing physiological solution. The selectivity of nicergoline for alpha 1-adrenoceptors seen in binding experiments also depends on the presence of the bromonicotinic moiety of the molecule. It is concluded that nicergoline, but not its substituent parts, displays both alpha 1-adrenoceptor and calcium antagonism. The latter property may account for some of the observed effects of this compound.

Animals↗
PubMed1986

Effects of bepridil and of its quaternary derivative on rat tail artery.

The effects of bepridil and its quaternary ammonium derivative (BN+) were compared, showing that: (i) both drugs inhibited K+-induced contractions with similar time courses and potencies, (ii) bepridil blocked the tonic but not the phasic component of contractions elicited by noradrenaline, whereas BN+ had no effect on noradrenaline-elicited contractions. These results, and the relative insensitivity of skinned taenia caeci to bepridil, suggest that this drug and BN+ do not act directly on contractile proteins but affect K+- and noradrenaline-induced calcium channel activities differentially.

Animals↗
PubMed1986

Mechanical responses of rat vascular smooth muscle cells to rat interferon.

Mechanical responses elicited by rat interferon (IFN) were investigated in rat vascular smooth muscle cells (RVSMC) in primary culture. IFN induced a dose-dependent reduction in the apparent cell surface area, which was quantitatively comparable to, or even greater than, the response elicited by the alpha-adrenergic agent, phenylephrine. Neither mock-IFN nor rat IFN-neutralized with specific antibodies elicited such a response. The sensitivity of RVSMC to the mechanical effect of IFN was species specific. In addition, the effect of IFN was inhibited by the Ca2+ entry blocker, verapamil.

Animals↗
PubMed1985

Cyclic AMP-dependent phosphorylation of a 16 kDa protein in a plasma membrane-enriched fraction of rat aortic myocytes.

Phosphorylation induced by cAMP-dependent protein kinase was examined in a plasma membrane-enriched fraction from control and beta-adrenergic-stimulated rat aortic myocytes. Phosphorylation of a 16 kDa protein which copurified with the plasma membrane marker (Na+ + K+)-ATPase was most prominent. It was decreased by pretreatment of the myocytes with isoproterenol and the effect of isoproterenol was inhibited by propranolol. Both phosphorylation induced by cAMP-dependent protein kinase and its inhibition by isoproterenol pretreatment declined in preparations exposed to endogenous phosphatase. These results provide strong evidence that beta-adrenergic stimulation of aortic myocytes induces in situ phosphorylation of a 16 kDa plasma membrane protein.

Animals↗
PubMed1985

Diphenylalkylamine calcium antagonists interact with alpha-adrenoceptor binding sites in aortic membranes.

Some interactions of calcium antagonists with [3H]prazosin and [3H]yohimbine binding sites were investigated in bovine aorta membranes. Diphenylalkylamines (flunarizine, cinnarizine and bepridil) acted as competitors of the two ligands with Ki values in the microM range. With the exception of verapamil, reference compounds (nifedipine, Bay-K 8644, diltiazem) and the peripheral benzodiazepine receptor antagonist PK 11195 did not displace the ligands. The apparent affinity of the diphenylalkylamines for alpha-adrenoceptor was consistent with the concentrations producing vasodilatation.

Animals↗
PubMed1985

Effect of vasoactive intestinal polypeptide (VIP) on cyclic AMP level and relaxation in rat isolated aorta.

The effects of vasoactive intestinal polypeptide (VIP) on cyclic nucleotide (cAMP and cGMP) levels and smooth muscle relaxation were investigated in rat isolated aorta and compared to those of the beta-adrenergic agonist isoprenaline. VIP increased the cAMP level of rat aorta in a concentration-dependent manner with an EC50 of 0.1 microM. VIP 1 microM maximally increased the cAMP level 7 fold, whereas the beta-adrenergic agonist isoprenaline 10 microM elevated the cAMP level only 2.5 fold. VIP 1 microM relaxed the precontracted rat aorta by only about 16% whereas isoprenaline 10 microM induced a relaxation of about 86%. VIP did not alter the cGMP level and its effect on cAMP content was not changed in the presence of indomethacin (5 microM). No quantitative correlation could thus be established between increases in total tissue levels of cAMP and the degree of relaxation in rat isolated aorta.

Animals↗
PubMed1985

Characterization of two distinct alpha-adrenoceptor binding sites in smooth muscle cell membranes from rat and bovine aorta.

In order to characterize postjunctional alpha adrenoceptor binding sites of aortic smooth muscle, the specific binding of (3H)prazosin and (3H)yohimbine to membranes prepared from the medial layers of rat and bovine thoracic aorta was investigated. Binding of (125I)-BE 2254 (2-[B-(4-hydroxyphenyl)-ethylaminomethyl] tetralone) and (3H)RX 781094 (idazoxan) was also examined in bovine membranes. Each of the ligands displayed saturable, specific binding to a single population of sites; the KD values of the respective ligands were similar in the two animal species. The number of (3H)prazosin and (125I)BE 2254 binding sites (160-190 fmol X mg protein-1 in the two species) was higher than the number of (3H)yohimbine and (3H)RX 781094 binding sites (110-120 fmol X mg protein-1 in the bovine and 50 fmol X mg protein-1 in the rat). Alpha-adrenoceptor ligands inhibited binding of the ligands with the following orders of potency:prazosin greater than BE 2254 greater than yohimbine greater than RX 781094 greater than clonidine in the case of (3H)-prazosin, and yohimbine greater than RX 781094 greater than clonidine greater than prazosin in the case of (3H)yohimbine. Methoxamine, in concentrations up to 10 microM, was without effect on the binding of either ligand. The absence or presence of Na+, K+ or Ca2+ added at physiological concentrations did not change the order of potency of displacing ligands whereas Ca2+ reduced by 50% the numbers of (3H)prazosin and (3H)-yohimbine sites and Na+ increased by 3-fold the affinity of (3H)yohimbine.(ABSTRACT TRUNCATED AT 250 WORDS)

Adrenergic alpha-Agonists↗
PubMed1985

Insensitivity of calcium-dependent endothelial stimulation in rat isolated aorta to the calcium entry blocker, flunarizine.

In rat aortic segments complete with endothelium, acetylcholine (1 microM) relaxed noradrenaline, phenylephrine and prostaglandin F2 alpha (PGF2 alpha)-induced contractions of various magnitudes. Maximal 1 microM phenylephrine-induced contractions were relaxed to a greater extent than were maximal contractions induced by the other two agonists. Contractions elicited by various concentrations of phenylephrine and PGF2 alpha in the presence of a maximal effective concentration of the calcium entry blocker flunarizine (3 microM) were relaxed by acetylcholine to about the same residual tension as were contractions elicited in the absence of flunarizine. Acetylcholine (1 microM) and phenylephrine (1 microM) increased tissue levels of guanosine cyclic 3'5'-monophosphate (cyclic GMP) by about 37 fold and 2 fold respectively. Preincubation of tissues in the absence of calcium abolished these agonist-induced increases in cyclic GMP levels, but preincubation with flunarizine had no significant effect on the increase in cyclic GMP level induced by the agonists. Pretreatment with flunarizine had no significant effect on the basal tissue level of cyclic GMP, but pretreatment in calcium-free solution reduced the basal tissue level of the cyclic nucleotide by about half. It is concluded that in rat aorta, endothelium-dependent acetylcholine-induced relaxation and endothelium-dependent acetylcholine and phenylephrine-induced increases in tissue levels of cyclic GMP, are dependent on extracellular calcium, but are not antagonized by flunarizine. This may indicate that if calcium channels of endothelial cells are activated by these agonists, their characteristics are not identical with those of the calcium channels of the smooth muscle cells.

Acetylcholine↗
PubMed1985

Modulation by endothelium of contractile responses in rat aorta in absence and presence of flunarizine.

The possible modulation by endothelium of phenylephrine- and prostaglandin F2 alpha-induced mobilization of calcium for contraction in the rat aorta has been investigated. Contractions elicited by these and other agonists are inhibited in the presence of endothelium. For any single concentration of phenylephrine in the presence of endothelium, the initial phasic components of contractions were significantly greater, the maximal contractions were achieved sooner and were less well maintained as compared to contractions elicited in the absence of endothelium. The kinetic characteristics of contractions stimulated by single concentrations of PGF2 alpha were similar in the presence and absence of endothelium and did not exhibit initial phasic components of contraction. Sub-maximal contractions-elicited by both PGF2 alpha and phenylephrine in the absence of endothelium were inhibited to a greater extent by flunarizine 3 microM than equieffective contractions elicited in the presence of endothelium. Maximal contractions elicited by phenylephrine (1 microM) were inhibited to a similar extent by flunarizine in the presence and absence of endothelium, but maximal contractions elicited by PGF2 alpha (30 microM) were inhibited by flunarizine to a greater extent in the presence than in the absence of endothelium. It is concluded that an endothelium-derived factor, perhaps distinct from endothelium-derived relaxing factor, can modulate the ability of both phenylephrine and PGF2 alpha to mobilize calcium for contraction. This modulatory effect is associated with an enhanced mobilization of intracellular calcium. Thus, submaximal concentrations of both agonists were less dependent on extracellular calcium than on intracellular calcium to elicit contractions in the presence of endothelium, as compared to contractions elicited in the absence of endothelium.

Animals↗
PubMed1984

Highly enriched, minimally disrupted plasma membrane vesicles from aortic myocytes grown in primary culture.

A plasma membrane-enriched fraction (fraction 1B) has been obtained from rat aortic myocytes grown in primary culture. Plasma membrane markers, 5'-nucleotidase and ouabain-sensitive (Na+ + K+)-ATPase, are enriched 4.1- and 8.7-fold, respectively, in this fraction. Although endoplasmic reticulum marker NADPH-cytochrome c reductase is the most enriched in mitochondrial and heavy sucrose density gradient fractions, substantial enrichment of this marker is also observed in membrane fraction 1. This membrane preparation therefore contains a certain quantity of endoplasmic reticulum. Cytochrome c oxidase is de-enriched by a factor of 0.04 in fraction 1, indicating that it is essentially clear of mitochondrial contamination. Homogenization of aortic media-intima layers using a whole-tissue technique induces greater disruption of mitochondria and subsequent contamination of membrane fractions than does the procedure for cell disruption. Analysis of electrophoretic gels, vesicle density distribution and electron micrographs of enriched membrane fractions provide evidence that plasma membrane enriched from cultured myocytes is less traumatized than comparable fractions obtained from intact tissue. The potential value of such a highly enriched, minimally disrupted plasma membrane preparation is discussed.

5'-Nucleotidase↗
PubMed1984

Age-related changes of in vivo beta-adrenergic responsiveness in normotensive and spontaneously hypertensive rats.

The variations of plasma cyclic AMP concentration caused by propranolol and isoproterenol were studied in order to investigate endogenous stimulation and responsiveness of the beta-adrenoceptor-cyclic AMP system in vivo, in 7 and 18 week old unanaesthetized male SHR and WKY. In both age groups, the basal cyclic AMP level was higher in SHR than in controls but was significantly reduced to a comparable value in the two strains after intraperitoneal injection of 2.5 mg X kg-1 propranolol, a dose which markedly depressed or even abolished the effect of isoproterenol. Cumulative dose-response curves of plasma cyclic AMP concentration obtained in another group of rats after successive subcutaneous injections of isoproterenol showed that the ED50 value of this drug was higher in SHR than in WKY and increased in a parallel manner in the two strains between 7 and 18 weeks of age (respectively from 0.21 +/- 0.01 to 0.34 +/- 0.02 mumol X kg-1 in WKY and from 0.32 +/- 0.02 to 0.52 +/- 0.03 mumol X kg-1 in SHR). At the same time blood pressure increased much less markedly in WKY (from 115 +/- 4 to 119 +/- 2 mm Hg) than in SHR (from 134 +/- 3 to 179 +/- 3 mm Hg). Altogether these results show that endogenous beta-adrenergic stimulation of the cyclic AMP system was higher in SHR of both ages in spite of a diminished responsiveness of this system to exogenous isoproterenol.

Aging↗
PubMed1984

Different effects of phenylephrine and clonidine on 86Rb efflux and on contraction in the rat caudal artery.

A large transitory efflux of 86Rb was induced by concentrations of phenylephrine (10(-6) and 10(-4)M) or clonidine (10(-4)M) which were able to produce phasic contraction. By contrast, a lower concentration of clonidine (10(-6) M), which could only induce a slower (tonic) contraction, only produced a weak but sustained 86Rb efflux. These results show qualitative and quantitative differences in the effects of the two alpha-agonists. They further support the view that phasic and tonic alpha-adrenergic responses may be related to different mechanisms.

Animals↗
PubMed1984

Dissociation between endothelium-mediated increases in tissue cGMP levels and modulation of aortic contractile responses.

Both methoxamine and clonidine elicited similar maximal contractions of rat isolated aorta in the absence of endothelium. These contractions were not associated with changes in tissue levels of cGMP or cAMP. In the presence of endothelium maximal methoxamine-induced contractions were not less than those elicited in the absence of endothelium but maximal clonidine-induced contractions were reduced to about 10% of those in the absence of endothelium. However, in the presence of endothelium both methoxamine and clonidine induced similar increases in tissue cGMP levels of about 1.5 to 2 fold; cAMP levels were unchanged. There is therefore a dissociation between endothelium-mediated inhibition of maximal contractile responses and increases in tissue levels of cGMP.

Adrenergic alpha-Agonists↗