Browsing by Author "Donoso, MV"

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    A2B adenosine receptor mediates human chorionic vasoconstriction and signals through arachidonic acid cascade
    (2005) Donoso, MV; López, RL; Miranda, R; Briones, R; Huidobro-Toro, JP
    Because adenosine is a vascular tone modulator, we examined the effect of adenosine and congeners in the vascular reactivity of isolated human placental vessels and in perfused cotyledons. We characterized its vasomotor action and tentatively identified the receptor subtypes and their intracellular signaling mechanisms. We recorded isometric tension from the circular layer of chorionic vessel rings maintained under 1.5 g of basal tension or precontracted with KCl. The relative order of potency of adenosine and structural analogs is consistent with the expression of A2B receptors, 5'-(N-ethylcarboxamido) adenosine (NECA) being the most potent. The maximal contraction ranged from 45% to 60% of the KCl standard response, except for an A(2A) receptor agonist that did not exceed 15%. Consistently, NECA was 100-fold more potent than adenosine to raise the perfusion pressure of ex vivo perfused cotyledons. In contrast, a selective A(3) receptor agonist relaxed precontracted rings of chorionic vessels. Whereas a selective A3 receptor antagonist was ineffective to antagonize adenosine-induced contraction, A(2) or A(1) receptor antagonists reduced adenosine-induced vasoconstriction concentration-dependently. Denudation of the endothelial layer reduced adenosine- and NECA-induced contractions by 50-70%. Furthermore, indomethacin reduced adenosine- or NECA-induced contractions concentration-dependently in intact and endothelium-denuded rings. A thromboxane receptor antagonist blocked adenosine- and NECA-induced contractions in intact and endothelium-denuded rings, suggesting the involvement of an arachidonic acid metabolite as the mediator of the vasoconstriction. We propose that adenosine A(2B) receptors mediate the adenosine- induced contraction vasomotor effect in human chorionic vessels and that this involves synthesis of a thromboxane receptor activator or a related prostanoid.
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    BIBP 3226, suramin and prazosin identify neuropeptide Y, adenosine 5'-triphosphate and noradrenaline as sympathetic cotransmitters in the rat arterial mesenteric bed
    (1997) Donoso, MV; Steiner, M; HuidobroToro, JP
    The physiological role of neuropeptide Y (NPY) and extracellular adenosine 5'-triphosphate (ATP) in sympathetic neurotransmission is becoming increasingly clear, To assess whether NPY and ATP act as cotransmitters together with noradrenaline (NA) in the sympathetic nerves of the superior mesenteric artery, the changes in perfusion pressure of the arterial mesenteric bed caused by nerve stimulation were recorded, Depolarization of the perivascular superior mesenteric arterial nerves caused frequency-and time-dependent increases in the perfusion pressure that were abolished by guanethidine, which implied the sympathetic origin of these responses. Independent perfusion with either 500 nM BIBP 3226, an NPY Y-1 antagonist; 3 mu M suramin, a competitive purinoceptor antagonist; or 0.1 nM prazosin, a competitive alpha-1 adrenoceptor antagonist, evoked approximately a 30% reduction in the rise in perfusion pressure caused by the 20-to 30-Hz electrical depolarization of the perimesenteric arterial nerves, Prazosin (0.1 nM) blocked the increases in perfusion pressure caused by electrical stimulation of the perimesenteric nerves but did not significantly reduce the vasomotor effect of exogenous NA, Likewise, 5-methyl urapidil and chloroethylclonidine, alpha-1 adrenoceptor antagonists with selectivity for the alpha-1A and alpha-1B receptor subtypes, respectively, concentration-dependently decreased the increase in perfusion pressure elicited by electrical stimulation of the perimesenteric nerves at concentrations lower than that required to block the vasoconstriction elicited by exogenous NA. The combined perfusion of 3 mu M suramin plus 0.1 nM prazosin did not result in a complete inhibition of the physiological response. Only upon the simultaneous application of BIBP plus suramin plus prazosin was the rise in perfusion pressure abolished. These results support the working hypothesis that the sympathetic nerves of the rat mesenteric bed release NPY, ATP and NA that act as postjunctional cotransmitters in this neuroeffector junction.
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    Neuropeptide Y is a vasoconstrictor and adrenergic modulator in the hamster microcirculation by acting on neuropeptide Y-1 and Y-2 receptors
    (1995) Boric, MP; Martinez, A; Donoso, MV; HuidobroToro, JP
    The microvascular effects of neuropeptide Y, and two analogs with preferential affinity for different neuropeptide Y receptor subtypes, were assessed by intravital microscopy on the hamster cheek pouch. The interaction of neuropeptide Y and its analogs with noradrenaline was also studied. Superfusion with 0.1-300 nM neuropeptide Y caused a concentration-dependent reduction in microvascular conductance that was paralleled by reductions in arteriolar and venular diameters. These effects of neuropeptide Y were equipotent with noradrenaline, but slower to develop and longer-lasting than that of noradrenaline. Neuropeptide Y did not affect permeability to macromolecules, as measured by extravasation of fluorescent dextran. The neuropeptide Y Y-1 receptor agonist, [Leu(31),Pro(34)]neuropeptide Y, mimicked neuropeptide Y with similar potency but shorter duration, while neuropeptide Y-(13-36), a neuropeptide Y Y-2 receptor agonist, was at least 10-fold less potent than neuropeptide Y to induce a delayed and prolonged reduction in microvascular conductance. The joint superfusion of 1 nM neuropeptide Y plus 0.1 mu M noradrenaline did not cause synergism, nor even summation of effects, but reduced the contractile effect of noradrenaline. No synergism was observed after a 10 min priming with 1 nM neuropeptide Y, followed by its joint application with 0.1 mu M noradrenaline, but a significant vasodilation and hyperemia ensued upon stopping noradrenaline application. Priming with 1 nM [Leu(31),Pro(34)]neuropeptide Y prolonged noradrenaline vasoconstriction without evidence of hyperemia. In contrast, priming with 1 nM neuropeptide Y-(13-36) significantly antagonized noradrenaline vasoconstriction. These findings indicate that both neuropeptide Y receptor subtypes are present in arterioles and venules of the hamster, and suggest that their activation with neuropeptide Y induces a rapid (Y-1 receptor subtype activation) and a delayed (Y-2 receptor subtype activation) vasocontractile response. The interaction with noradrenaline is complex, without evidence for synergism, but neuropeptide Y Y-2 receptor activation seems to antagonize noradrenaline and/or to facilitate auto-regulatory vasodilation after the catecholamine-induced vasoconstriction.
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    Neuropeptide Y is released from human mammary and radial vascular biopsies and is a functional modulator of sympathetic cotransmission
    (2004) Donoso, MV; Miranda, R; Irarrázaval, MJ; Huidobro-Toro, JP
    The role of neuropeptide Y (NPY) as a modulator of the vasomotor responses mediated by sympathetic cotransmitters was examined by electrically evoking its release from the perivascular nerve terminals of second- to third-order human blood vessel biopsies and by studying the peptide-induced potentiation of the vasomotor responses evoked by exogenous adenosine 5' triphosphate (ATP) and noradrenaline (NA). Electrical depolarization of nerve terminals in mammary vessels and radial artery biopsies elicited a rise in superfusate immunoreactive NPY (ir-NPY), which was chromatographically identical to a standard of human NPY (hNPY); a second peak was identified as oxidized hNPY. The amount released corresponds to 4-6% of the total NPY content in these vessels. Tissue extracts also revealed two peaks; hNPY accounted for 68-85% of the ir-NPY, while oxidized hNPY corresponded to 7-15%. The release process depended on extracellular calcium and on the frequency and duration of the electrical stimuli; guanethidine blocked the release, confirming the peptide's sympathetic origin. Assessment of the functional activity of the oxidized product demonstrated that while it did not change basal tension, the NA-evoked contractions were potentiated to the same extent as with native hNPY. Moreover, NPY potentiated both the vasomotor action of ATP or NA alone and the vasoconstriction elicited by the simultaneous application of both cotransmitters. RT-PCR detected the mRNA coding for the NPY Y-1 receptor. In summary, the release of hNPY or its oxidized species, elicited by nerve terminal depolarization, coupled to the potentiation of the sympathetic cotransmitter vasomotor responses, highlights the modulator role of NPY in both arteries and veins, strongly suggesting its involvement in human vascular sympathetic reflexes. Copyright (C) 2004 S. Karger AG, Basel.
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    Neuropeptide Y Y-1 receptors are involved in the vasoconstriction caused by human sympathetic nerve stimulation
    (1997) Racchi, H; Schliem, AJ; Donoso, MV; Rahmer, A; Zuniga, A; Guzman, S; Rudolf, K; HuidobroToro, JP
    Neuropeptide Y, a novel neurotransmitter, interacts with selective membrane receptors to cause vasoconstriction. Frequency- and concentration-dependent isometric contractions were observed in human inferior mesenteric artery and vein mounted rings that were stimulated with either electrical pulses (70 V, 0.5 ms, 2.5-20 Hz) or noradrenaline. The antagonism elicited by 100 nM tetrodotoxin and 1 mu M guanethidine confirmed the neuronal and sympathetic origins of the vasomotor response. Incubation with BIBP 3226 ((R)-N-2-(diphrnacetyl)-N-(4-hydroxyphenyl)-methyl-D-arginineamide), a selective neuropeptide Y Y, receptor antagonist, significantly reduced the vasoconstriction. The incomplete antagonist activity of BIBP 3226 tends to support the hypothesis of sympathetic co-transmission involving neuropeptide Y, adenosine 5'-triphosphate and noradrenaline. These findings were confirmed in parallel studies using rat superior mesenteric artery and vein ring preparations. (C) 1997 Elsevier Science B.V.
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    Pharmacological characterization of the ET(A) receptor in the vascular smooth muscle comparing its analogous distribution in the rat mesenteric artery and in the arterial mesenteric bed
    (1996) Donoso, MV; Faundez, H; Rosa, G; Fournier, A; Edvinsson, L; HuidobroToro, JP
    The potency of ET-1, ET-2, and ET-3 to contract the isolated perfused rat arterial mesenteric bed was 2.73 +/- 0.57, 1.63 +/- 0.32, and 144 +/- 30 nM, respectively. The vasomotor effect of the ETs was slow in onset, persistent but reversible. Sarafotoxin S6b mimicked the ETs with a potency twofold lower than ET-1; sarafotoxin S6c and the C-terminal hexapeptide of ET-1 was inactive. ET(B) agonists such as IRL-1620 and AGETB-89 were inactive as vasoconstrictors within the range of concentrations examined. Minor chemical modifications of ET-1 amino acids residues in position 7 or 21 decreased significantly the peptide potency; ET-I analogues with one or none of the disulfide bonds resulted inactive. The vasomotor effect of ETs was blocked in a competitive, reversible, and selective manner by FR 139317 and BQ-123, the latter being about threefold less potent than the former antagonist. The potency of FR 139317 was 20-fold higher to antagonize ET-3 than ET-1, and threefold higher to block ET-2 than ET-1. In strict analogy to FR 139317, BQ-123 was 12-fold more potent to antagonize ET-3 than ET-1, and fourfold more potent to antagonize ET-2 than ET-1. Upon removal of the endothelial cell layer, the vasomotor potency of ET-1 or the antagonist potency of FR 139317 remained unaltered, suggesting that the vasomotor receptors are localized in the arterial smooth muscles. The ET-l-induced vasomotor responses desensitized, an effect not crossed to noradrenaline (NA); perfusion with 10 mu M indomethacin did not alter the vasomotor potency of ET-1, excluding the participation of eicosanoids in the arteriolar effects of ET-1. In isolated rings of the rat mesenteric artery, set to record isometric contractions of the circular muscular layer, the potency of the ETs and their structural analogues was as follows: ET-2 = ET-1 = sarafotoxin S6b > ET-3 > sarafotoxin S6c. The C-terminal hexapeptide of ET-1 and [Ala(1,3,11,15)]ET-1 were inactive. The ET-1-induced vasoconstriction was antagonized in a concentration-dependent fashion by FR 139317. These results allow to conclude that the ET(A) receptors present in the arterial mesenteric circulation are localized in the vascular smooth muscle of the large-sized arteries as well as the smaller arterioles and precapillary vessels of the rat arterial mesenteric bed. Copyright (C) 1996 Elsevier Science Inc.
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    Stimulation of the sympathetic perimesenteric arterial nerves releases neuropeptide Y potentiating the vasomotor activity of noradrenaline: Involvement of neuropeptide Y-Y-1 receptors
    (1997) Donoso, MV; Brown, N; Carrasco, C; Cortes, V; Fournier, A; HuidobroToro, JP
    Neuropeptide Y (NPY) appears to be involved in the sympathetic regulation of vascular tone. To assess the putative role of NPY in mesenteric circulation, the release and biological effect of NPY were examined after electrical stimulation of perimesenteric arterial nerves, Nerve stimulation with trains of 2-30 Hz increased the perfusion pressure of the arterially perfused rat mesenteric bed in a frequency-and time-dependent fashion, Trains of 15-30 Hz significantly displaced to the left, approximately threefold, the noradrenaline (NA)-induced presser concentration-response curve, in addition to increasing significantly its efficacy. Perfusion with 10 nM exogenous NPY mimicked the electrical stimulation effect, causing a threefold leftward shift of the NA concentration-response curve and increasing the maximal NA response. These effects were antagonized by 100 nM BIBP 3226, indicating the activity of NPY-Y-1 receptors. Electrical stimulation of the perimesenteric nerves released immunoreactive NPY (ir-NPY) in a frequency-dependent fashion; the ir-NPY coelutes with synthetic NPY as confirmed by HPLC. Both the electrically induced presser response and the calcium-dependent release of NPY were obliterated in preparations perfused with 1 mu M guanethidine or in rats pretreated intravenously for 48 h with 6-hydroxydopamine, thus revealing the sympathetic origin of these phenomena. Only a small proportion of the total NPY content in the perimesenteric arterial nerves is released after electrical stimulation. Chromatographic studies of the physiological sources of the ir-NPV support that NPY fragments are generated via peptidase degradation. The present findings demonstrate that NPY is re[eased from the perimesenteric arterial sympathetic nerves and acts, via the activation of NPY-Y-1 receptors, as the mediator responsible for the potentiation of NA's effect on perfusion pressure in the isolated rat mesenteric bed.
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    Synergism between neuropeptide Y and norepinephrine highlights sympathetic cotransmission
    (1999) Cortés, V; Donoso, MV; Brown, N; Fanjul, R; López, C; Fournier, A; Huidobro-Toro, JP
    Although abundant literature supports the notion that neuropeptide Y (NPY) synergizes in vivo and in vitro, the vasomotor activity elicited by norepinephrine (NE), the converse interaction (i.e,, the adrenergic modulation of the NPY vasomotor response) has been less characterized. To assess whether NE synergizes the vasomotor effect of NPY, the rat arterial mesenteric bed was chosen as a model experimental system. Mesenteries were precontracted with NE and few minutes later were pelf used with exogenous NPY. Under these conditions, NPY contracted the arterial mesenteric bed with an EC50 value of 0.72 +/- 0.06 nM. NPY was unable to contract this vascular territory without an agonist-induced precontraction, Other agonists, such as endothelin-1, a synthetic analog of prostaglandin F-2 alpha, or 5-hydroxytryptamine, also were effective primers because in their presence, NPY was a potent vasoconstrictor. In contrast, mesenteries precontracted with KCI failed to evidence the NPY-induced rise in perfusion pressure. Two structural analogs of NPY, PW and [Leu(31),Pro(34)]NPY, mimicked the activity of NPY. The NPY fragment 13-36 did not elicit such a response. All NPY analogs exhibited less efficacy and potency relative to NPY. The NPY- and related structural analog-induced vasoconstriction was competitively and reversibly antagonized by BIBP 3226; the pA(2) of the NPY interaction was 7.0. The application of 0.1 to 1 mu M BIBP 3226 or 0.1 to 10 nM prazosin at the peak of the NPY vasomotor response elicited a gradual blockade of the vasoconstriction. Although BIBP 3226 blocked the increase in perfusion pressure elicited by NPY, leaving unaffected the NE-induced tone, 10 nM prazosin blocked the full response, including the NE-induced component. Tissue preincubation with 200 nM nifedipine abolished the NPY-induced vasoconstriction; likewise, the acute application of 10 to 100 nM nifedipine blocked gradually the maximal NPY-induced contraction. Removal of the mesenteric endothelial layer increased the potency of NPY by 2-fold; it also slightly potentiated the antagonist activity of BIBP 3226, The synergism between NPY and NE backs the principle of sympathetic cotransmission.
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    α2-adrenoceptors control the release of noradrenaline but not neuropeptide Y from perivascular nerve terminals
    (2002) Donoso, MV; Carvajal, A; Paredes, A; Tomic, A; Koenig, CS; Huidobro-Toro, JP
    Neuropeptide Y (NPY) and noradrenaline (NA) are co-transmitters at many sympathetic synapses, but it is not yet clear if their release is independently regulated. To address this question, we quantified the electrically evoked release of these co-transmitters from perivascular nerve terminals to the mesenteric circulation in control and drug-treated rats. 6-Hydroxydopamine reduced the tissue content and the electrically evoked release of ir-NPY and NA as well as the rise in perfusion pressure. A 0.001 mg/kg reserpine reduced the content of ir-NPY and NA, but did not modify their release nor altered the rise in perfusion pressure elicited by the electrical stimuli. However, 0.1 mg/kg reserpine reduced both the content and release of NA but decreased only the content but not the release of ir-NPY; the rise in perfusion pressure was halved. Clonidine did not affect the release of ir-NPY while it lowered the outflow of NA, not altering the rise in perfusion pressure elicited by the electrical stimuli. Yohimbine, did not modify the release of ir-NPY but increased the NA outflow, it antagonized the clonidine effect. Therefore, presynaptic alpha(2)-adrenoceptors modulate the release of NA but not NPY, implying separate regulatory mechanisms. (C) 2002 Elsevier Science Inc. All rights reserved.