0000000000126174
AUTHOR
M.a. Noguera
Amplifying effect of serotonin on contractile responses in rat aorta and depletion of intracellular Ca-stores
1. Serotonin, 1 microM, induces a contractile response in isolated rat aorta in the presence or absence of extracellular Ca. 2. In Ca-free media, the fast phasic contraction is lower in magnitude and further addition of serotonin evokes no response. 3. Recovery of the contractile response in Ca-free medium is obtained by a 40 min incubation in Ca-containing solution. 4. In Ca, Mg-free medium, the response to serotonin is significantly higher than that obtained in the presence of Mg. 5. An amplifying effect of serotonin on the contractile responses induced by serotonin itself or by noradrenaline was observed in Ca-containing but not in Ca-free solution.
Acute and Chronic Captopril, but Not Prazosin or Nifedipine, Normalize Alterations in Adrenergic Intracellular Ca2+ Handling Observed in the Mesenteric Arterial Tree of Spontaneously Hypertensive Rats
The effect of hypertension and acute (36-h) or chronic (from age 6 to 16 weeks) antihypertensive treatment with prazosin (2 mg kg(-1) per day), nifedipine (50 mg kg(-1) per day), or captopril (50 mg kg(-1) per day) on Ca2+ mobilization due to alpha1-adrenoceptor activation was analyzed in functional studies using arterial rings [four conductance/distributing vessels: aorta, main mesenteric, iliac, and tail arteries and two resistance vessels; first and second small mesenteric artery branches obtained from spontaneously hypertensive rats (SHR, 6 and 16 weeks old) and age-matched Wistar Kyoto rats (WKY)]. Maximal response to noradrenaline in the presence of extracellular Ca2+ is not affected …
Effect of Divalent Cations on the Contractile Response of Rat Aorta to Depolarization before and after Nifedipine Treatment
The influence of the divalent cations, Ca2+, Mg2+ and Ba2+, on the contractile response of the rat aorta to KCl and on the recovery of this response after nifedipine treatment was analyzed. KCl (80 mmol/l) promoted a two-phase (phasic and tonic) contractile response in Krebs solution but, as expected, no contractile response in Ca(2+)-free medium. In Mg(2+)-free medium, the phasic response to KCl was unaffected but the tonic one decreased slowly, suggesting that a long incubation time in the absence of Mg2+ (65 min) promotes a loss of or a change in the intracellular distribution of this ion that modifies Ca2+ entry through L channels or Ca2+ handling. Ba2+ (1.8 mmol/l) contracted the rat a…
Capacitative Ca2+ entry associated with α1-adrenoceptors in rat aorta
In rat aorta, depletion of internal Ca2+ stores by addition of noradrenaline (1 microM) induces a biphasic response (an initial phasic response and a tonic one) mediated by two different intracellular Ca2+ pools. This response cannot be repeated, suggesting a depletion of internal Ca2+ stores sensitive to noradrenaline. In absence of the agonist, this depletion is the signal for the entry of extracellular Ca2+, not only to refill the stores but also, under our experimental conditions, to activate the contractile proteins thus inducing an increase in the resting tone (IRT) that constitutes functional evidence of this Ca2+ entry. The ionic channels involved in the mechanism of the IRT have be…
Modulatory role of a constitutively active population of α1D-adrenoceptors in conductance arteries
A constitutively active population of α1D-adrenoceptors in iliac and proximal, distal, and small mesenteric rat arteries was studied. The increase in resting tone (IRT) that evidences it was observed only in iliac and proximal mesenteric and was inhibited by prazosin (pIC50 = 9.57), 5-methylurapidil (pIC50 = 7.61), and BMY 7378 (pIC50 = 8.77). Chloroethylchlonidine (100 μmol/l) did not affect IRT, but when added before the other antagonists it blocked their effect. The potency shown by BMY 7378 confirms the α1D-subtype as responsible for IRT. BMY 7378 displayed greater inhibition of adrenergic responses in iliac (pIC50 = 7.57 ± 0.11) and proximal mesenteric arteries (pIC50 = 8.05 ± 0.2) th…
Relationships between structure and vascular activity in a series of benzylisoquinolines
1 1In the present work, the properties of 3-methyl isoquinoline, 3,4-dihydropapaverine, tetrahydropapaverine and tetrahydropapaveroline were compared with those of papaverine and laudanosine. The work includes functional studies on rat isolated aorta contracted with noradrenaline, caffeine or KC1, and a determination of the affinity of the compounds for α1-adrenoceptors and calcium channel binding sites, with [#H]-prazosin, [#H]-nitrendipine and [#H]-(+)-cis-diltiazem binding to rat cerebral cortical membranes. The effects of papaverine derivatives on the different molecular forms of cyclic nucleotide phosphodiesterases (PDE) isolated from bovine aorta were also determined. 2 2The three pap…
Intervention of two voltage-dependent calcium-entry pathways in the contractile response to acetylcholine and KCl in rat uterus.
The contractile response of rat uterine smooth muscle was investigated. Verapamil and diltiazem concentration-dependently relax the sustained contractions induced by KCl (56 mmol/l) or acetylcholine (10(-4) mol/l). This inhibitory effect was not not freely reversed by washing the tissue and subsequently no contractile response was obtained in depolarized tissue, but a lower biphasic response (phasic and tonic) to acetylcholine was observed. Addition of cumulative concentrations of CaCl2 (1.2-19.2 mmol/l) induced a partial recovery of the contractile response to acetylcholine or KCl, but addition of MgCl2 (1.2-19.2 mmol/l did not. When the channel was reactivated by a third addition of KCl o…
Action of an extract from the seeds of Fraxinus excelsior L. on metabolic disorders in hypertensive and obese animal models.
Nuzhenide and GI3, the principal secoiridoids of an extract obtained from the seeds of Fraxinus excelsior L. (FXE), are believed to be the active compounds responsible for the previously reported hypoglycemic effects of this extract. In this study, the effects of FXE were studied in two animal models which are representative of metabolic disorders: spontaneously hypertensive rats (SHR) and obese Zucker rats. SHR were acutely treated (oral gavage) with different doses of FXE. In addition, SHR and Zucker rats were chronically fed (20 or 5 weeks, respectively) with standard chow supplemented with FXE. Acute treatment with FXE (200 mg per kg body weight) decreased systolic blood pressure as in …
Different and common intracellular calcium-stores mobilized by noradrenaline and caffeine in vascular smooth muscle.
Noradrenaline (NA) 1 microM and caffeine (CAF) 10 mM induce a contractile response in isolated rat aorta maintained at 37 degrees C either in the presence or absence of extracellular calcium. In Ca-free media the contractile response was reduced and contractile activity of CAF only occurred at 25 degrees C. NA induced a biphasic response in Ca-free medium, with a fast phasic contraction followed by a smaller more sustained contraction. The response induced by CAF consisted of a fast transient contraction which returned to a level below the resting tone. After washing, further addition of NA or CAF evoked no increase in smooth muscle tension. The influence of Mg-depletion in the extracellula…
Differences in the signaling pathways of α(1A)- and α(1B)-adrenoceptors are related to different endosomal targeting.
AIMS: To compare the constitutive and agonist-dependent endosomal trafficking of α(1A)- and α(1B)-adrenoceptors (ARs) and to establish if the internalization pattern determines the signaling pathways of each subtype. METHODS: Using CypHer5 technology and VSV-G epitope tagged α(1A)- and α(1B)-ARs stably and transiently expressed in HEK 293 cells, we analyzed by confocal microscopy the constitutive and agonist-induced internalization of each subtype, and the temporal relationship between agonist induced internalization and the increase in intracellular calcium (determined by FLUO-3 flouorescence), or the phosphorylation of ERK1/2 and p38 MAP kinases (determined by Western blot). RESULTS AND C…
β2- and β1-Adrenoceptor Expression Exhibits a Common Regulatory Pattern With GRK2 and GRK5 in Human and Animal Models of Cardiovascular Diseases.
To explore if genic expression of β(1)- or β(2)-adrenoceptors (ARs) exhibits a common regulatory pattern with G protein-coupled receptor kinase (GRK) 2, GRK3, or GRK5 expression, we determined messenger RNA levels for these genes in different tissues from human and animal models of cardiovascular disease. We measured genic expression by qRT polymerase chain reaction in the left and right ventricles or peripheral blood mononuclear cells from healthy (n = 21), hypertensive (n = 20), heart failure (n = 24), and heart transplanted patients (n = 17) or in left ventricle, peripheral blood mononuclear cells, and kidney from spontaneously hypertensive rats or L-N-methyl-arginine-induced hypertensiv…
Effects of different agents on the contractile response elicited by extracellular calcium after depletion of internal calcium stores in rat isolated aorta.
Abstract Noradrenaline, 1 μm, induced a sustained contractile response in rat isolated aorta in the presence and in the absence of extracellular Ca2+. After depleting the noradrenaline-sensitive intracellular Ca2+ stores, an increase in the basal tone of the aorta was observed during the incubation period in the presence of Ca2+ and in the absence of the agonist. We have tested the possible pathways through which Ca2+ enters the cell to refill the previously depleted Ca2+ pools, a process that is accompanied by an increase in tension. The magnitude of this increase does not depend on the presence of Mg2+ in the extracellular medium nor on the temperature, suggesting that it is mediated by a…
Effect of divalent cations on KCl- and noradrenaline-induced contractile responses in rat aorta after nifedipine treatment.
Abstract Nifedipine (1 μM) relaxed the sustained contractile responses induced by 1 μM noradrenaline or 60 mM KCl in rat aortic strips. After washing, a second addition of the spasmogens gave smaller tonic contractions than the first one. Even more, a third addition of KCl also gave a smaller contraction than the first one, but a complete recovery of the contractile response to noradrenaline was obtained by a third addition of this agonist. Application of cumulative amounts of Ca2+ or Ba2+ (2.4–24 mM) on the residual contraction in response to these agents after nifedipine treatment, but in the absence of the blocker, restored the magnitude of the contractile responses. Addition of cumulati…
Participation of Intracellular Calcium Stores in Serotonin-Induced Contractions in Rat Aorta
Serotonin 1 mumol/l induces a contractile response in the isolated rat aorta in both the presence or absence of extracellular Ca. The present study analyzes the influence of temperature and caffeine on subsequent serotonin-induced contractions. In Ca-free medium, the contraction elicited by serotonin was higher at 25 degrees C than at 37 degrees C. In addition, the existence of two independent intracellular Ca pools releasable by serotonin, one of them also sensitive to caffeine, is postulated. The results also showed that addition of serotonin decreases the contractile response to this agonist in Ca-free medium.
Cytosolic Ca2+ and Phosphoinositide Hydrolysis Linked to Constitutively Active α1d-Adrenoceptors in Vascular Smooth Muscle
In the present study, we analyzed changes in intracellular Ca2+ levels and inositol phosphate accumulation related to a population of alpha 1d-adrenoceptors in rat aorta resembling constitutively active receptors. Following intracellular Ca2+ store depletion by noradrenaline in Ca2+-free medium and removal of the agonist, restoration of extracellular Ca2+ induced four signals: a biphasic (transient and sustained) increase in [Ca2+]i, inositol phosphate accumulation, and a contractile response in the aorta. The transient increase in Ca2+, the inositol phosphate accumulation, and the contractile response were not observed in aortae incubated with prazosin or BMY 7378 [8-[2-[4-(2-methoxyphenyl…
The three α1-adrenoceptor subtypes show different spatio-temporal mechanisms of internalization and ERK1/2 phosphorylation
AbstractWe analyzed the kinetic and spatial patterns characterizing activation of the MAP kinases ERK 1 and 2 (ERK1/2) by the three α1-adrenoceptor (α1-AR) subtypes in HEK293 cells and the contribution of two different pathways to ERK1/2 phosphorylation: protein kinase C (PKC)-dependent ERK1/2 activation and internalization-dependent ERK1/2 activation. The different pathways of phenylephrine induced ERK phosphorylation were determined by western blot, using the PKC inhibitor Ro 31-8425, the receptor internalization inhibitor concanavalin A and the siRNA targeting β-arrestin 2. Receptor internalization properties were studied using CypHer5 technology and VSV-G epitope-tagged receptors. Activ…
a1D-Adrenoceptors are responsible for the high sensitivity and the slow time-course of noradrenaline-mediated contraction in conductance arteries.
The objective of this study was to determine whether the different time-course characteristics of α1-adrenoceptor-mediated contraction in arteries can be related to the subtypes involved. Contractile responses to noradrenaline (NA) were compared with inositol phosphate accumulation and extracellular signal-regulated kinase (ERK)1/2 phosphorylation after α1-agonist stimuli in the same vessels in the presence or absence of α1-antagonists in rat or in α1-subtype knockout (KO) mice. Aorta, where α1D-AR is the main functional subtype, had higher sensitivity to NA (in respect of inositol phosphate [IP], pERK1/2, and contractile response) than tail artery, where the α1A-adrenoceptor subtype is pre…
Role of cyclic nucleotide phosphodiesterase isoenzymes in contractile responses of denuded rat aorta related to various Ca 2+ sources
We have examined the cyclic nucleotide phosphodiesterase isoforms (PDE) involved in the contractile response of rat aorta to different agonists and different experimental procedures for use in functional studies. The inhibitory effect of AAL 05 on the different PDEs isolated from bovine aortic smooth muscle was examined. Compound AAL 05 appeared to be a selective PDE3 inhibitor. We analyzed the ability of the non-selective inhibitor IBMX (3-isobutyl-1-methylxanthine) and the isoenzyme selective inhibitors nimodipine (type 1), AAL 05 (6-(N-methyl-N-cyclohexyl butyl carboxamide) quinolin-2-one) and SKF 94120 (5-(4-acetamidophenyl) pyrazin-2(1H)-one; type3), rolipram (type4) and zaprinast (typ…
Effect of different treatments in calcium-free medium on basal tone and contractile responses of guinea pig tracheae.
Acetylcholine (ACh; 0.1 mmol/l) and KCl (80 mmol/l) induce a biphasic contractile response in isolated guinea pig tracheae maintained at 37 degrees C either in the presence or absence of extracellular Ca2+. Exposure of the tissue to Ca(2+)-free solution evokes a significant decrease in basal tone and the sources of Ca2+ appear to be decreased by prolonged agonist stimulation, and even more by successive agonist stimulation. After an incubation period of 20 min in Ca(2+)-containing solution, the response is restored. Mg(2+)-depletion in Ca(2+)-free medium increased the contractile response to ACh, but not to KCl, and delayed the tonic component of the next contraction elicited in Ca(2+)-cont…
Relaxant activity of three aporphine alkaloids from Annona cherimolia on isolated aorta of rat.
Abstract In the present study we tested the relaxant effect of three aporphine alkaloids—roemerine, anonaine and dehydroroemerine—isolated from the roots of Annona cherimolia, on isolated strips of rat thoracic aorta. All compounds completely relaxed KCl- and noradrenaline-induced contractions with different potencies depending on their structural characteristics. The experiments, carried out in Ca2+-free medium using two different agonists (noradrenaline and caffeine) which mobilize calcium intracellularly by different mechanisms of action, showed that the alkaloids made no contribution to intracellular calcium processes. The present study provides evidence that the relaxant effects produc…
Vasodilator Effects of Liriodenine and Norushinsunine, Two Aporphine Alkaloids Isolated from <i>Annona cherimolia,</i>in Rat Aorta
The effect of two aporphines, liriodenine and norushinsunine, isolated from Annona cherimolia, were studied in the rat aorta in order to examine their mechanism of action. Both alkaloids (10–7–10–4 mol/l) showed relaxant effects on the contractions elicited by 10–6 mol/l noradrenaline (NA) or 80 mmol/l KC1, but, while liriodenine showed a nonspecific relaxant action on both spasmogens, norushinsunine was more potent on KC1-induced contraction. In Ca2+-free medium, both alkaloids (0.1 mmol/l) inhibited the responses elicited by NA, but not those elicited by caffeine. This inhibitory action occurred when the alkaloids were present during the release of the Ca2+ internal stores or during the r…
Activation of α1A-adrenoceptors desensitizes the rat aorta response to phenylephrine through a neuronal NOS pathway, a mechanism lost with ageing
Background and purpose A NO-mediated desensitization of vasoconstrictor responses evoked by stimulation of α1 -adrenoceptors has been reported in different vessels. We investigated the involvement of each α1 -adrenoceptor subtype and constitutive NOS isoforms and the influence of ageing and hypertension on this process. Experimental approach Wistar and spontaneously hypertensive rats (SHR), 16, 32, 52 and 72 weeks-old, were used to evaluate the desensitization process. Expression of α1 -adrenoceptor subtypes, endothelial NOS (eNOS) and neuronal NOS (nNOS) were determined in rat aorta and left ventricle (LV). Expression levels were also evaluated in LV of a group of heart failure patients wi…
A possible structural determinant of selectivity of boldine and derivatives for the alpha 1A-adrenoceptor subtype.
1. The selectivity of action of boldine and the related aporphine alkaloids, predicentrine (9-O-methylboldine) and glaucine (2,9-O-dimethylboldine) and alpha 1-adrenoceptor subtypes was studied by examining [3H]-prazosin competition binding in rat cerebral cortex. WB 4101 and benoxathian were used as selective alpha 1A-adrenoceptor antagonists. 2. In the competition experiments [3H]-prazosin (0.2 nM) binding was inhibited by WB 4101 and benoxathian. The inhibition curves displayed shallow slopes which could be subdivided into high and low affinity components (pKi = 9.92 and 8.29 for WB 4101, 9.35 and 7.94 for benoxathian). The two antagonists recognized approximately 37% of the sites with h…
Actions of alpha-adrenoceptor blocking agents on two types of intracellular calcium stores mobilized by noradrenaline in rat aorta.
In isolated rat aortic strips noradrenaline induces a biphasic contractile response in Ca-free medium, associated with two different intracellular calcium pools, one of which is common to caffeine. We analyzed the mechanisms involved in the depletion and repletion of both intracellular Ca pools sensitive to noradrenaline in different experimental procedures in presence of prazosin, phentolamine and yohimbine. At 37 degrees C the alpha-adrenergic blocking agents inhibited contractile responses to noradrenaline in Ca-free medium, with prazosin being highly selective. alpha 2-adrenoceptors probably do not participate in the release of Ca from internal stores, as no contractile response was obs…
Different β-adrenoceptor subtypes coupling to cAMP or NO/cGMP pathways: implications in the relaxant response of rat conductance and resistance vessels
Background and Purpose To analyse the relative contribution of β1-, β2- and β3-adrenoceptors (Adrb) to vasodilatation in conductance and resistance vessels, assessing the role of cAMP and/or NO/cGMP signalling pathways. Experimental Approach Rat mesenteric resistance artery (MRA) and aorta were used to analyse the Adrb expression by real-time-PCR and immunohistochemistry, and for the pharmacological characterization of Adrb-mediated activity by wire myography and tissue nucleotide accumulation. Key Results The mRNAs and protein for all Adrb were identified in endothelium and/or smooth muscle cells (SMCs) in both vessels. In MRA, Adrb1 signalled through cAMP, Adrb3 through both cAMP and cGMP…