Contraction Of Small Arteries Research Articles

TP and/or EP3 receptors mediate the vasoconstrictor and pressor responses of prostaglandin F2α in mice and/or humans.

The vasoconstrictor and/or pressor effects of prostaglandin (PG)F2α participate in the development of vascular pathologies and limit the clinical use of the agent. This study aimed to determine the receptor types responsible for the vasoconstrictor activity of PGF2α and whether they mediate the pressor response evoked by the prostanoid under in vivo conditions. Experiments were performed on genetically altered mice and/or on vessels from these mice or humans. Here we show that deletion of the thromboxane-prostanoid receptor (TP-/-) abolished or drastically diminished the contraction to PGF2α in isolated mouse vessels (some of which were resistance arteries) and reduced the elevation in blood pressure evoked by the prostanoid under in vivo conditions. In accordance, TP antagonism abolished the contraction in small arteries of human omentum. Further deletion of E prostanoid receptor type 3 (EP3-/-) removed the PGF2α-evoked contraction that remained in some TP-/- arteries and added to the effect of TP-/- on the elevation in blood pressure evoked by the prostanoid under in vivo conditions. In contrast, the uterine contraction to PGF2α mediated via the F prostanoid receptor (FP) was unaltered in TP-/-/EP3-/- mice. These results demonstrate that the non-FP receptors TP and/or EP3 mediate the vasoconstrictor and pressor effects of PGF2α, which are still of concern under clinical conditions.-Liu, B., Li, J., Yan, H., Tian, D., Li, H., Zhang, Y., Guo, T., Wu, X., Luo, W., Zhou, Y. TP and/or EP3 receptors mediate the vasoconstrictor and pressor responses of prostaglandin F2α in mice and/or humans.

The α2 isoform Na,K-ATPase modulates contraction of rat mesenteric small artery via cSrc-dependent Ca2+ sensitization.

The Na,K-ATPase is involved in a large number of regulatory activities including cSrc-dependent signalling. Upon inhibition of the Na,K-ATPase with ouabain, cSrc activation is shown to occur in many cell types. This study tests the hypothesis that acute potentiation of agonist-induced contraction by ouabain is mediated through Na,K-ATPase-cSrc signalling-dependent sensitization of vascular smooth muscle cells to Ca2+ . Agonist-induced rat mesenteric small artery contraction was examined in vitro under isometric conditions and in vivo in anaesthetized rats. Arterial wall tension and [Ca2+ ]i in vascular smooth muscle cells were measured simultaneously. Changes in cSrc and myosin phosphatase targeting protein 1 (MYPT1) phosphorylation were analysed by Western blot. Protein expression was examined with immunohistochemistry. The α1 and α2 isoforms of the Na,K-ATPase were transiently downregulated by siRNA transfection in vivo. Ten micromolar ouabain, but not digoxin, potentiated contraction to noradrenaline. This effect was not endothelium-dependent. Ouabain sensitized smooth muscle cells to Ca2+ , and this was associated with increased phosphorylation of cSrc and MYPT1. Inhibition of tyrosine kinase by genistein, PP2 or pNaKtide abolished the potentiating effect of ouabain on arterial contraction and Ca2+ sensitization. Downregulation of the Na,K-ATPase α2 isoform made arterial contraction insensitive to ouabain and tyrosine kinase inhibition. Data suggest that micromolar ouabain potentiates agonist-induced contraction of rat mesenteric small artery via Na,K-ATPase-dependent cSrc activation, which increases Ca2+ sensitization of vascular smooth muscle cells by MYPT1 phosphorylation. This mechanism may be critical for acute control of vascular tone.

Pannexin-1 channels do not regulate α1-adrenoceptor-mediated vasoconstriction in resistance arteries

Recent reports have provided evidence for a new concept that in small resistance arteries α1D-adrenoceptor-mediated contraction is intimately linked to pannexin-1 (Px1) hemichannels that open to allow the release of ATP, from the smooth muscle effector cell, that acts back on P2Y purinoceptors to cause contraction. This concept mainly relied on using mefloquine 10–20μM as a putative selective Px1 channel-blocking agent to completely inhibit the contraction to phenylephrine, but not K+ 40mM. Lower concentrations of mefloquine had no effect. The purpose of the present study was to explore the specificity of mefloquine for Px1 channels and the role of these channels in small artery contraction. In mouse and rat isolated small resistance arteries, either pressurised or set up for wire myography, the effects of mefloquine on contractions to K+, phenylephrine and a range of vasoconstrictor agents were assessed and compared with the Px1 channel inhibitor carbenoxolone. Mefloquine had a wide range of inhibitory actions at 10–20μM, some 200-fold above the concentrations previously shown to inhibit expressed Px1 channel activity. Mefloquine 3–10μM inhibited phenylephrine, U46619, vasopressin, endothelin-1, sympathetic nerve stimulation and K+ 40mM-mediated contractions in rat and mouse small mesenteric, and mouse thoracodorsal, arteries. Carbenoxolone 1–100μM did not inhibit the contractile responses to these agents in small resistance arteries. The present study demonstrates that in small resistance arteries there is no evidence that Px1 channels releasing ATP have any role in the constrictor actions of α1-adrenoceptor activation.

Pulmonary vasoconstrictor influence of endothelin in exercising swine depends critically on phosphodiesterase 5 activity

Both phosphodiesterase 5 (PDE5) inhibition and endothelin (ET) receptor blockade have been shown to induce pulmonary vasodilation. However, little is known about the effect of combined blockade of these two vasoconstrictor pathways. Since nitric oxide (NO) exerts its pulmonary vasodilator influence via production of cyclic guanosine monophosphate (cGMP) as well as through inhibition of ET, we hypothesized that interaction between the respective signaling pathways precludes an additive vasodilator effect. We tested this hypothesis in chronically instrumented swine exercising on a treadmill by comparing the vasodilator effect of the PDE5 inhibitor EMD360527, the ETA/ETB antagonist tezosentan, and combined EMD360527 and tezosentan. In the systemic circulation, vasodilation by tezosentan and EMD360527 was additive, both at rest and during exercise, resulting in a 17 ± 2% drop in blood pressure. In the pulmonary circulation, both EMD360527 and tezosentan produced vasodilation. However, tezosentan produced no additional pulmonary vasodilation in the presence of EMD360527, either at rest or during exercise. Moreover, in isolated preconstricted porcine pulmonary small arteries (∼300 μm) EMD360527 (1 nM-10 μM) induced dose-dependent vasodilation, whereas tezosentan (1 nM-10 μM) failed to elicit vasodilation irrespective of the presence of EMD360527. However, both PDE5 inhibition and 8Br-cGMP, but not 8Br-cAMP, blunted pulmonary small artery contraction to ET and its precursor Big ET in vitro. In conclusion, in healthy swine, either at rest or during exercise, PDE5 inhibition and the associated increase in cGMP produce pulmonary vasodilation that is mediated in part through inhibition of the ET pathway, thereby precluding an additional vasodilator effect of ETA/ETB receptor blockade in the presence of PDE5 inhibition.

Arteriolar vascular smooth muscle cells: Mechanotransducers in a complex environment

Contraction of small artery (diameters typically less than 250μm) vascular smooth muscle cells (VSMCs) plays a critical role in local control of blood flow and arterial pressure through its affect on vascular caliber. Specifically, contraction of small arteries in response to increased intraluminal pressure is referred to as the myogenic response and represents an important role for mechanotransduction. Critical questions remain as to how changes in pressure are sensed by VSMCs and transduced across the cell membrane to tune the contractile state of the cell. Recent studies suggest a pivotal role for interactions between VSMCs and extracellular matrix (ECM) proteins. Thus, pressure-induced deformation of ECM proteins and their cell surface receptors (for example, integrins) may initiate contraction and cytoskeletal remodeling through modulation of ion channels, membrane depolarization, increased intracellular Ca2+ and actomyosin crossbridge cycling. Importantly, it is argued that the contractile properties of small artery VSMCs reflect an intimate and integrated interaction with their extracellular environment and the three-dimensional structure of the vessel wall.

Uridine adenosine tetraphosphate (Up4A) as a novel coronary vasodilator in health and disease: Role of purinergic P1 and P2 receptors

Up4A is a potent endothelium‐derived contracting factor that contains purine and pyrimidine moieties and activates purinergic P2X and P2Y receptors. Up4A has recently been found to also have vasodilator effects. Here, we evaluated Up4A in isolated coronary small arteries from normal (N) swine and swine after myocardial infarction (MI), and studied the purinergic receptor subtypes involved. Surprisingly, Up4A (10−9–10−5 M) failed to induce contraction in small arteries from N and MI, but produced dose‐dependent relaxation in vessels from N and MI preconstricted with U46619. Sensitivity to Up4A was lower in MI than N. In N, Up4A induced vasodilation was reduced by nitric oxide (NO) inhibition with L‐NAME to a similar extent as by nonselective P1 antagonist 8PT and A2A antagonist SCH58261, while nonselective P2 antagonist PPADS, P2X1 antagonist MRS2159, and P2Y1 antagonist MRS2179 had no effect. In MI, dilation by Up4A was blunted by P2‐ (PPADS, MRS2159, MRS2179) and P1‐inhibition (8PT, SCH58261) and L‐NAME. Q‐PCR confirmed the presence of coronary P2X1, P2Y1 and A2A receptors. Thus, Up4A exerts a dilator rather than a constrictor effect in coronary microvessels which in N occurs via P1 (particularly A2A) mediated activation of NO, and both P1 and P2 in MI. These findings indicate that the mechanism of vasodilation by Up4A is different in the diseased heart.Supported by China Scholarship Council (2009624027)

Sympathetic neurogenic Ca2+ signalling in rat arteries: ATP, noradrenaline and neuropeptide Y

The sympathetic nervous system (SNS) plays an essential role in the control of total peripheral vascular resistance by controlling the contraction of small arteries. The SNS also exerts long-term trophic influences in health and disease; SNS hyperactivity accompanies most forms of human essential hypertension, obesity and heart failure. At their junctions with smooth muscle cells, the peri-arterial sympathetic nerves release ATP, noradrenaline (NA) and neuropeptide Y (NPY) onto smooth muscle cells. Confocal Ca(2+) imaging studies reveal that ATP and NA each produce unique types of postjunctional Ca(2+) signals and consequent smooth muscle cell contractions. Neurally released ATP activates postjunctional P2X(1) receptors to produce local, non-propagating Ca(2+) transients, termed 'junctional Ca(2+) transients', or 'jCaTs'. Neurally released NA binds to alpha(1)-adrenoceptors and can activate Ca(2+) waves or more uniform global changes in [Ca(2+)]. Neurally released NPY does not appear to produce Ca(2+) transients directly, but significantly modulates NA-induced Ca(2+) signalling. The neural release of ATP and NA, as judged by postjunctional Ca(2+) signals, electrical recording of excitatory junction potentials and carbon fibre amperometry to measure NA, varies markedly with the pattern of nerve activity. This probably reflects both pre- and postjunctional mechanisms, which are not yet fully understood. These phenomena, together with different temporal patterns of sympathetic nerve activity in different regional circulations, are probably an important mechanistic basis of the important selective regulation of regional vascular resistance and blood flow by the sympathetic nervous system.

Sympathetically evoked Ca2+signaling in arterial smooth muscle

The sympathetic nervous system plays an essential role in the control of total peripheral vascular resistance and blood flow, by controlling the contraction of small arteries. Perivascular sympathetic nerves release ATP, norepinephrine (NE) and neuropeptide Y. This review summarizes our knowledge of the intracellular Ca2+ signals that are activated by ATP and NE, acting respectively on P2X1 and alpha1-adrenoceptors in arterial smooth muscle. Each neurotransmitter produces a unique type of post-synaptic Ca2+ signal and associated contraction. The neural release of ATP and NE is thought to vary markedly with the pattern of nerve activity, probably reflecting both pre- and post-synaptic mechanisms. Finally, we show that Ca2+ signaling during neurogenic contractions activated by trains of sympathetic nerve fiber action potentials are in fact significantly different from that elicited by simple bath application of exogenous neurotransmitters to isolated arteries (a common experimental technique), and end by identifying important questions remaining in our understanding of sympathetic neurotransmission and the physiological regulation of contraction of small arteries.

Targeting Small Arteries of Hypertensive Status with Novel ATP-Sensitive Potassium Channel Openers&#

The excessive contraction of small arteries under high blood pressure is the main contributor to the pathological change of hypertension. Current anti-hypertensive drugs, which lack a selective effect on small arteries in a hypertensive state, may cause many adverse effects. We have developed a novel opener of ATP-sensitive potassium channels, iptakalim, the vasorelaxing action of which is determined by the hypertensive status of small arteries. In conscious stroke-prone, spontaneously hypertensive rats and "two-kidneys, one-clip" renal hypertensive dogs, this compound produced long-lasting hypotensive effects, with no tolerance. Furthermore, it exerted a protective effect against hypertensive damage to target organs. Given its potency and fewer side effects, iptakalim hydrochloride is a promising drug for the treatment of hypertension.

Different roles of ryanodine receptors and inositol (1,4,5)-trisphosphate receptors in adrenergically stimulated contractions of small arteries.

The functions of ryanodine receptors (RyRs) and inositol (1,4,5)-trisphosphate receptors [Ins(1,4,5)P(3)Rs] in adrenergically activated contractions of pressurized rat mesenteric small arteries were investigated. Caffeine (20 mM) but not phenylephrine (PE; 10 microM) facilitated the depletion of smooth muscle sarcoplasmic reticulum (SR) Ca(2+) stores by ryanodine (40 microM). In ryanodine-treated SR-depleted arteries, 1) Ca(2+) sparks were absent, 2) low concentrations of PE failed to elicit either vasoconstriction or normal asynchronous propagating Ca(2+) waves, and 3) high [PE] induced abnormally slow oscillatory contractions (vasomotion) and synchronous Ca(2+) oscillations. In ryanodine-treated SR-depleted arteries denuded of endothelium, high [PE] induced steady contraction and steady elevation of intracellular [Ca(2+)]. In contrast, 2-aminoethyl diphenylborate (2-APB), a putative blocker of Ins(1,4,5)P(3)Rs, produced opposite effects to ryanodine: 1) Ca(2+) sparks were present; 2) Ca(2+) waves were absent; 3) caffeine-releasable Ca(2+) stores were intact; and 4) PE, even at high concentrations on endothelial-denuded arteries, failed to elicit contraction, asynchronous Ca(2+) waves, or synchronous Ca(2+) oscillations or maintained elevated [Ca(2+)]. We conclude that 1) Ins(1,4,5)P(3)Rs are essential for adrenergically induced asynchronous Ca(2+) waves and the associated steady vasoconstriction, 2) RyRs are not appreciably opened during adrenergic activation (because PE did not facilitate the development of the effects of ryanodine), and 3) Ins(1,4,5)P(3)Rs are not essential for Ca(2+) sparks. This provides an explanation of the fact that adrenergic stimulation decreases the frequency of Ca(2+) sparks (previously reported) while simultaneously increasing the frequency of asynchronous propagating Ca(2+) waves; different SR Ca(2+)-release channels are involved.

Purinergic and adrenergic Ca2+ transients during neurogenic contractions of rat mesenteric small arteries.

Contraction of small arteries is regulated by the sympathetic nervous system, but the Ca2+ transients during neurally stimulated contraction of intact small arteries have not yet been recorded. We loaded rat mesenteric small arteries with the fluorescent Ca2+ indicator fluo-4 and mounted them in a myograph that permitted simultaneous (i) high-speed confocal imaging of fluorescence from individual smooth muscle cells, (ii) electrical stimulation of perivascular nerves, and (iii) recording of isometric tension. Sympathetic neuromuscular transmission was achieved by electrical field stimulation (EFS) (frequency, 10 Hz; pulse voltage, 40 V; pulse duration, 0.2 ms) in the presence of capsaicin and scopolamine (to inhibit 'sensory' and cholinergic nerves, respectively). During the first 20 s of EFS, force rose to a small peak and then declined. During this time, junctional Ca2+ transients (jCaTs) were present at relatively high frequency. We have previously attributed jCaTs to influx of Ca2+ through post-junctional P2X receptors activated by ATP. Propagating asynchronous Ca2+ waves, previously associated with bath-applied alpha1-adrenoceptor agonists, were not initially present. During the next 2.5 min of EFS, force rose slowly, and asynchronous propagating Ca2+ waves appeared. The selective alpha1-adrenoceptor antagonist prazosin abolished both the slowly developing contraction and the Ca2+ waves, but reduced the initial transient contraction by only ~25 %. During 3 min of EFS in prazosin, the frequency of jCaTs declined markedly; at sites at which at least one jCaT occurred, the average probability of a jCaT was 0.008 +/- 0.002 pulse-1 in the first 20 s and 0.0007 +/- 0.0002 pulse-1 in the last 20 s. We suggest that (i) ATP released from sympathetic varicosities activates the initial, transient, contraction and the activator Ca2+ is derived largely from jCaTs, and (ii) sympathetically released noradrenaline (NA) activates the later, major contraction through mechanisms involving alpha1-adrenoceptors and which are associated with propagating Ca2+ waves.

Serotonin-induced contraction in mesenteric resistance arteries: signaling and changes in deoxycorticosterone acetate-salt hypertension.

Large arteries from hypertensive subjects are hyperresponsiveness to 5-hydroxytryptamine (5-HT). We tested the hypothesis that small arteries (225 micro ID) have a profile similar to conduit arteries, including signal transduction mechanisms and the 5-HT receptor subtype(s) mediating arterial contraction in normal and high blood pressure. Aorta and mesenteric arteries from Sprague-Dawley (232+/-6 micro ID), sham (229+/-7 micro ID; systolic blood pressure, 120+/-2 mm Hg), or deoxycorticosterone acetate (DOCA)-salt rats (255+/-11 micro ID, 192+/-8 mm Hg) were mounted in a wire-based myograph. In resistance arteries from Sprague-Dawley rats, the 5-HT2A receptor mediated contraction; agonists of the 5-HT1B, 5-HT1D, 5-HT1F, and 5-HT2B receptor were inactive. The tyrosine kinase inhibitor genistein (5 micromol/L, 4.8-fold rightward shift), PD 098,059 (10 micromol/L, 3.2-fold shift), phospholipase C inhibitor NCDC (100 micromol/L), and nifedipine (50 nmol/L) reduced maximum 5-HT-induced contraction in small arteries (4.5% and 53% control, respectively). As in aorta, 5-HT had a decrease in threshold (100-fold lower), increase in potency (11.6-fold leftward shift), and increase in efficacy (140% sham response) in small arteries from DOCA-salt rats compared with sham. Unlike in aorta, 5-HT-induced contraction in DOCA-salt small arteries was shifted competitively by the 5-HT2A receptor antagonist ketanserin (-log K(B) [mol/L] for both sham and DOCA-salt, 9.25+/-0.1), and contraction to the 5-HT2B agonist BW723C86 was not observed. Thus, the 5-HT2A receptor remains the contractile receptor in hypertension in small arteries. Although similarities were observed for large and small arteries, differences under the condition of DOCA-salt hypertension exist that may determine serotonergic compounds effective in lowering blood pressure.

Noradrenaline-Induced Paxillin Phosphorylation, ERK Activation and MEK-Regulated Contraction in Intact Rat Mesenteric Arteries

In rat mesenteric arteries, noradrenaline (NA) induces a time-dependent increase in tyrosine phosphorylation of a number of proteins, one of which was identified as paxillin. NA-induced protein tyrosine phosphorylation was ablated by tyrosine kinase inhibition, virtually unaffected by protein kinase C (PKC) inhibition or PKC downregulation and was mimicked by KCl. NA also caused a time-dependent activation of the extracellular signal-regulated kinases (ERK)1 and ERK2. These responses were blocked by the ERK-activating kinase (MEK) inhibitor PD98059 and by tyrosine kinase inhibition but only modestly attenuated by PKC downregulation or inhibition. Pretreatment of cannulated mesenteric arteries (50 mm Hg internal pressure) with PD98059 significantly reduced the contractile responsiveness of the vessels to NA (1.56 ± 0.14 µM, EC₅₀ control; 3.32 ± 0.49 µM, EC₅₀ + PD98059, p < 0.01). Thus, NA induces time-dependent increases in protein-tyrosine phosphorylation and ERK activation in rat mesenteric arteries that could suggest a role for Ca²⁺-dependent non-receptor tyrosine kinases and ERKs in the response of small arteries to NA. In addition, the modulation of NA-induced mesenteric artery contraction by inhibition of the MEK/ERK pathway further implicates ERK in the regulation of, though perhaps not the mediation of NA-induced small artery contraction.

Endothelin-1-induced contraction of mesenteric small arteries is mediated by ryanodine receptor Ca2+ channels and cyclic ADP-ribose.

Contraction of vascular smooth muscle by endothelin-1 is dependent on extracellular and intracellular Ca2+. However, the role of ryanodine-sensitive Ca2+ stores in endothelin-1-induced contraction is unknown. Vascular contraction was measured in mesenteric small arteries (200-300 microm intraluminal diameter) isolated from Sprague-Dawley rats and maintained at a constant intraluminal pressure of 40 mm Hg. The presence of functional ryanodine receptor Ca2+ release channels (RyRC) was demonstrated by the finding that ryanodine (10 microM), which locks the RyRC in a subconductance state, produced significant contraction of small arteries in the presence of 15 mM KCl. This effect was inhibited by dantrolene (10 microM), a RyRC inhibitor. Dantrolene significantly reduced the ET(A) receptor-mediated contraction to endothelin-1 (10(-11)-10(-9) M). The ability of dantrolene to reverse contraction induced by endothelin-1 was also determined. Dantrolene (1-10 microM) produced concentration-dependent relaxation of vessels precontracted to 38+/-3% of resting diameter with endothelin-1 but had no effect in vessels precontracted to a similar degree with phenylephrine or KCl. Because activation of RyRC may be dependent on production of cyclic ADP-ribose, the effect of nicotinamide (2 mM), an inhibitor of ADP-ribosyl cyclase, on contraction to endothelin-1 was determined. Nicotinamide had an inhibitory effect similar to that produced by dantrolene. A combination of nicotinamide and dantrolene had no greater effect than either agent alone, suggesting a common pathway for cyclic ADP-ribose and RyRC. In summary, endothelin-1 induces contraction of small mesenteric arteries through ET(A) receptor-mediated production of cyclic ADP-ribose and activation of RyRC.

Altered angiotensin II-induced small artery contraction during the development of hypertension in spontaneously hypertensive rats

This study assessed whether the angiotensin-II (Ang II)-induced contractile responsiveness of resistance arteries is altered during the development of hypertension in spontaneously hypertensive rats (SHR). Structural parameters and Ang II-stimulated contraction were determined in small mesenteric arteries from 6-week-old (phase of developing hypertension) and 21-week-old SHR (phase of established hypertension), compared with age-matched Wistar-Kyoto rats (WKY). To ascertain whether effects were specific for Ang II, contractile responses to another vasoactive agonist, vasopressin (AVP), were also determined. Systolic blood pressure was measured in conscious rats by the tail-cuff method. Segments of third-order mesenteric arteries (≈200 μm in diameter and 2 mm in length) were mounted in a pressurized system with the intraluminal pressure maintained at 45 mm Hg. Blood pressure was significantly increased in SHR ( P < .001) and was higher in adult than in young SHR ( P < .001). Ang II dose-dependently increased contraction, with responses significantly greater ( P < .05) in SHR than in age-matched WKY. SHR, in the early phase of hypertension, exhibited significantly augmented contractile responses (E max = 70 ± 5%), compared with SHR with established hypertension (E max = 33 ± 5%). These effects were not generalized, as responses to AVP were not significantly different between young and adult SHR. Functional Ang II-elicited alterations were associated with structural modifications: 6-week-old SHR had smaller media to lumen ratio compared with 21-week-old SHR (8.1% ± 0.17% v 10.6% ± 0.20%, P < .01). In young SHR vessels the media cross-sectional area was unchanged relative to age-matched WKY rats, suggesting eutrophic remodeling (remodeling index 101.4% v 93.3% young v adult), whereas the cross-sectional area of adult vessels was increased in comparison to WKY rats, suggesting mild hypertrophic remodeling (growth index −1.0% v 15.2%, young v adult). In conclusion, the present study demonstrates that in SHR with early hypertension and slight medial thickening, Ang II-mediated vascular contractile responsiveness is significantly augmented compared with SHR with established hypertension and more severe vascular structural changes. These findings indicate attenuation, as hypertension progresses, of the initially enhanced vascular reactivity to Ang II that is present during the development of hypertension in SHR

Role of AT2 receptors in angiotensin II-stimulated contraction of small mesenteric arteries in young SHR.

This study assesses the receptor subtype (AT1 and AT2) through which angiotensin II (Ang II) mediates contraction in small arteries of young and adult spontaneously hypertensive rats (SHR). Segments of third-order mesenteric arteries ( approximately 200 microm in lumen diameter) were mounted in a pressurized system. Systolic blood pressure and media:lumen ratio of small arteries were significantly greater (P<0.001) in young SHR and adult SHR than in age-matched Wistar-Kyoto rats (WKY). Ang II-induced contractile effects were significantly increased (P<0.05) in young SHR compared with age-matched WKY. AT1 blockade with losartan, and combined AT1 and AT2 blockade with losartan and PD123319, abolished Ang II-stimulated contraction in young and adult rats. AT2 blockade (PD123319) significantly reduced (P<0.01) Ang II-elicited contraction in young SHR but had no effect in WKY or adult SHR, indicating that AT2 receptors may contribute to Ang II-induced contraction in young SHR. To determine the Ang receptor status in rat mesenteric vessels, AT1 and AT2 receptor mRNA expression was determined by reverse transcription-polymerase chain reaction. AT1 and AT2 receptor protein expression were detected by Western blot analysis. AT1 receptor mRNA was equally expressed in age-matched rats, but expression was significantly lower in young rats compared with adult rats. AT2 receptor mRNA was weakly expressed in WKY and adult SHR. In vessels from young SHR, AT2 receptor mRNA expression was significantly increased compared with the other groups. AT1 receptor protein was equally expressed in adult rats of both strains but was undetectable in young rats. AT2 receptor protein was only detectable in young rats, with the magnitude of expression greater in SHR than WKY. In conclusion, Ang II-stimulated contractile responses are augmented in vessels from young SHR. These effects are reduced by selective AT2 blockade and abolished by AT1 blockade, indicating that both Ang receptor subtypes are involved in contraction in young SHR. In WKY and adult SHR, losartan, but not PD123319, inhibited Ang II-induced contraction, indicating the exclusive involvement of AT1 receptors. Thus, in SHR, in the phase of developing hypertension, enhanced Ang II-stimulated vascular contraction may be associated with changes in Ang II receptor status, as evidenced pharmacologically and by increased vascular AT2 receptor mRNA and protein expression.

Rhythmic contractions of isolated small arteries from rat: role of calcium.

In order to investigate the mechanisms behind rhythmic contractions in small arteries of the mesenteric arcade from Wistar rats, the calcium dependency of the oscillations in response to noradrenaline activation was tested on isolated vessels. Application of 1 microM ryanodine or 30 microM TMB-8 (procedures known to inhibit Ca2+ release from intracellular stores) totally abolished the rhythmic activity, even though the antagonists had opposite effects on the amplitude of the contractile response to noradrenaline. Verapamil (1 microM) or felodipine (1 nM) (agents known to inhibit influx of extracellular Ca2+) also abolished the oscillations and reduced the maximal noradrenaline response by about 40%. Reducing the extracellular Ca2+ concentration to 0.1 mM reduced the amplitude of the noradrenaline response to a similar extent as 1 nM felodipine, but did not eliminate the oscillations. This may indicate that the effect of calcium entry blockers was to eliminate the voltage-dependency of Ca2+ inflow rather than just reducing the Ca2+ level. Manoeuvres that would increase the cytosolic Ca2+ concentration (exposure to caffeine or to the calcium agonist BAY-K 8644) increased the frequency of the oscillations. These observations indicate an important role, not only for voltage-operating channels, but also for intracellular calcium stores in the generation of rhythmic contractions in these small arteries. Oscillations appear to be generated by an interplay between membrane activation and intracellular calcium stores.

Regulation of diacylglycerol metabolism by vasoconstrictor hormones in intact small arteries.

The initiation of receptor-mediated small artery contraction is dependent on inositol 1,4,5-trisphosphate-stimulated release of stored calcium. The role of the other product of inositol lipid hydrolysis, 1,2-diacylglycerol, in maintaining contraction remains controversial. Therefore, we have determined the contractile response of rat subcutaneous small arteries (< 300 microns i.d.), when mounted as ring preparations in a myograph, to noradrenaline, angiotensin II, KCl-induced membrane depolarization, and a cell-permeable diglyceride, dioctanoylglycerol. In parallel experiments, the conversion of this diglyceride to dioctanoylphosphatidate was studied in 32P-labeled vessels. Dioctanoylglycerol produced a slow-onset sustained contraction that was dependent on extracellular calcium. This was accompanied by the generation of the lipid dioctanoylphosphatidate. Noradrenaline and KCl induced rapid-onset sustained contractions and increased the production of dioctanoylphosphatidate (75% and 91%, respectively). In addition, dioctanoylglycerol levels were reduced (41%) after noradrenaline stimulation, suggesting activation of diacylglycerol kinase. In contrast, the contractile response to angiotensin II was transient, and this agonist did not significantly affect the conversion of dioctanoylglycerol to phosphatidate. Noradrenaline markedly increased (fourfold) the formation of endogenous phosphatidate, whereas endogenous 1,2-diacylglycerol was increased (47%) with angiotensin II. These results demonstrate that phosphatidate formation is regulated by vasoconstrictor hormones during receptor-mediated contraction, independent of diglyceride mass. Modulation of the levels of lipid second messengers downstream from phospholipid hydrolysis may represent a mechanism by which agonists that act through the same signaling system produce different contractile responses.

Differential reactivity to 5-hydroxytryptamine in canine coronary arteries.

Responses to 5-hydroxytryptamine (5HT) were compared in large [2.1-3.0 mm outside diameter (OD)], medium (1.5-1.8 mm OD) and small (0.5-1.0 mm OD) isolated canine coronary arteries. 5HT produced contraction of large and medium arteries, with the maximal response averaging 35.1 +/- 4.0 and 21.0 +/- 3.3%, respectively, of the contraction to 100 mM KCl. Endothelial removal increased the response to 5HT, with the maximal response averaging 43.6 +/- 12.6 and 32.4 +/- 7.5%, respectively, of the 100 mM KCl contraction. Small arteries did not contract significantly to 5HT in the presence or absence of endothelium. However, 5HT (10(-6) M) contracted small arteries that were contracted with 30 mM KCl, averaging 130 +/- 3% of the original contraction to KCl. This further contraction to 5HT was slightly potentiated by removal of the endothelium. We conclude that, unlike larger epicardial arteries, coronary arteries less than 1 mm OD are unresponsive to 5HT under resting conditions. Failure of small arteries to contract to 5HT cannot be explained by an inhibitory influence of the endothelium. However, 5HT enhances the contraction of small arteries to K+, and this response tends to be augmented by endothelial removal.

The structural mechanism of arterial contraction demonstrated through the adrenalin effect upon the coronary arteries.

Despite numerous clinical conceptions concerning the pathogenetic role of vascular spasm, we have, until to-day. no knowledge of the slruclural phenomena occurring in arterial contraction and spasm. Contraction of small arteries in the heart muscle of dogs submitted to the influence of adrenalin are made evident by means of special fixation method. The histological picture of the arterial contraction shows a characteristic distribution of muscular cells and elastic elements in the arterial wall. This structure is identical with that observed in the closure apparatus of certain small arteries regulating arteriovenous anastomoses. From a general point of view. the observations may open a fertile perspective not only for investigations in the* physiological field, but also for analysis and interpretation of certain pathological findings in cardiovascular and renal disorders. © 1952 S. Karger AG, Basel.