Angiotensin II-induced inhibition of calcium currents in hamster submandibular ganglion neurons.

Angiotensin II (Ang II) is one of the most important vasoconstrictive hormones but is also known to act as a neuromodulator and a neurotransmitter in the central and peripheral nervous systems. In a previous study, we have shown that Ang II, via AT1 receptors, induced depolarization by inhibition of M-type K(+) channels and SK channels in submandibular ganglion (SMG) neurons. In this study, we investigated the effects of Ang II on calcium channel current (I(Ca)) in acutely dissociated SMG neurons by the patch-clamp technique using the whole-cell configuration. Ang II inhibited total I(Ca) by 32.1+/-2.7%. The half-maximum inhibitory concentration (IC(50)) of Ang II for inhibiting I(Ca) was 0.8 microM. In the presence of 1 microM losartan, which is a selective antagonist of AT1 receptors, the effect of Ang II was attenuated (7.6+/-1.5%). Application of a strong depolarizing voltage prepulse did not affect the Ang II-induced inhibition of I(Ca) (32.8+/-2.8%). Intracellular dialysis of GDP-beta-S attenuated the inhibition of I(Ca) (6.8+/-2.1%). The mean percentage inhibitions of L-, N- and P/Q-type VDCCs by Ang II were 29.1+/-1.7, 16.3+/-6.0 and 1.2+/-0.8%, respectively, of the total I(Ca).     

PI3-kinase inhibitors abolish the enhanced chronotropic effects of angiotensin II in spontaneously hypertensive rat brain neurons

Angiotensin II (Ang II), acting at Ang II type 1 receptors (AT1Rs), increases the firing rate of neurons from Wistar-Kyoto (WKY) rat brain via protein kinase C (PKC)- and calcium-calmodulin kinase II (CaMKII)-dependent mechanisms. The objectives of this study were twofold; first, to compare the Ang-II-stimulated increase in firing of neurons from WKY and spontaneous hypertensive rats (SHR) and second, to elucidate the signaling mechanisms involved. Action potentials were measured in neurons cultured from SHR and WKY rat brains using the whole cell configuration of the patch-clamp technique in the current-clamp mode. Ang II (100 nM) caused three- and sixfold increases in neuronal firing rate in WKY rat and SHR neurons, respectively; effects that were abolished by the AT1R antagonist Losartan (1 microM). Co-administration of calphostin C (10 microM, a PKC inhibitor) and KN-93 (10 microM, a CaMKII inhibitor) completely blocked this Ang II action in WKY rat neurons, while they caused only a approximately 50% attenuation in SHR neurons. The residual increase in firing rate produced by Ang II in SHR neurons was blocked by inhibitors of phosphatidylinositol 3 kinase (PI3-kinase), either LY 294002 (10 microM) or wortmannin (100 nM). These observations suggest that a PI3-kinase signaling pathway may be responsible for the enhanced chronotropic effect produced by Ang II in SHR neurons.     

Angiotensin II angiogenic effect in vivo involves vascular endothelial growth factor- and inflammation-related pathways

Although accumulating lines of evidence indicate the proangiogenic role of angiotensin II (Ang II), little is known about the molecular mechanisms associated with such an effect. This study aimed to identify molecular events involved in Ang II-induced angiogenesis in the Matrigel model in mice. C57Bl/6 female mice received a subcutaneous injection of either Matrigel or Matrigel with Ang II (10(-7) M) alone, with Ang II and an AT1 receptor antagonist (candesartan, 10(-6) M), or with Ang II and AT2 receptor antagonist (PD123319, 10(-6) M). After 14 days, angiogenesis was assessed in the Matrigel-plug by histological evaluation and cellular counting. Ang II increased by 1.9-fold the number of cells within the Matrigel (p < 0.01 versus control). Immunohistological analysis revealed the presence of macrophages, endothelial and smooth muscle cells, and the development of vascular-like structure. Such an angiogenic effect was associated with an increase in vascular endothelial growth factor (VEGF) (1.5-fold, p < 0.01), endothelial nitric oxide (eNOS) (1.7-fold, p < 0.01), and cyclooxygenase-2 (1.4-fold, p < 0.05) protein levels measured by Western blotting. Conversely, Ang II treatment did not affect MMP-9 and MMP-2 activity, assessed by zymography. Blockade of AT1 receptor completely prevented the Ang II-induced angiogenesis and protein regulations, whereas that of AT2 was ineffective. Administration of VEGF neutralizing antibody (2.5 microg ip twice a week) and cyclooxygenase-2 selective inhibitor (nimesulide, 30 mg/L) also hampered Ang II proangiogenic effect. In addition, Ang II-induced cell ingrowth was impaired by treatment with nitric oxide synthase inhibitor (L-NAME, 10 mg/kg/day) and in eNOS-deficient mice. Therefore, in an in vivo model, Ang II induced angiogenesis through AT1 receptor, which involved activation of VEGF/eNOS-related pathway and of the inflammatory process.     

Regulation of alveolar fluid clearance and ENaC expression in lung by exogenous angiotensin II

Angiotensin II (Ang II) has been demonstrated as a pro-inflammatory effect in acute lung injury, but studies of the effect of Ang II on the formation of pulmonary edema and alveolar filling remains unclear. Therefore, in this study the regulation of alveolar fluid clearance (AFC) and the expression of epithelial sodium channel (ENaC) by exogenous Ang II was verified. SD rats were anesthetized and were given Ang II with increasing doses (1, 10 and 100 μg/kg per min) via osmotic minipumps, whereas control rats received only saline vehicle. AT1 receptor antagonist ZD7155 (10 mg/kg) and inhibitor of cAMP degeneration rolipram (1 mg/kg) were injected intraperitoneally 30 min before administration of Ang II. The lungs were isolated for measurement of alveolar fluid clearance. The mRNA and protein expression of ENaC were detected by RT-PCR and Western blot. Exposure to higher doses of Ang II reduced AFC in a dose-dependent manner and resulted in a non-coordinate regulation of α-ENaC vs. the regulation of β- and γ-ENaC, however Ang II type 1 (AT1) receptor antagonist ZD7155 prevented the Ang II-induced inhibition of fluid clearance and dysregulation of ENaC expression. In addition, exposure to inhibitor of cAMP degradation rolipram blunted the Ang II-induced inhibition of fluid clearance. These results indicate that through activation of AT(1) receptor, exogenous Ang II promotes pulmonary edema and alveolar filling by inhibition of alveolar fluid clearance via downregulation of cAMP level and dysregulation of ENaC expression.     

Chronotropic effect of angiotensin II via type 2 receptors in rat brain neurons

Previously, we determined that angiotensin II (Ang II) elicits an Ang II type 2 (AT(2)) receptor-mediated increase of neuronal delayed rectifier K(+) (I(KV)) current in neuronal cultures from newborn rat hypothalamus and brain stem. This requires generation of lipoxygenase (LO) metabolites of arachidonic acid (AA) and activation of serine/threonine phosphatase type 2A (PP-2A). Enhancement of I(KV) results in a decrease in net inward current during the action potential (AP) upstroke as well as shortening of the refractory period, which may lead to alterations in neuronal firing rate. Thus, in the present study, we used whole-cell current clamp recording methods to investigate the AT(2) receptor-mediated effects of Ang II on the firing rate of cultured neurons from the hypothalamus and brain stem. At room temperature, these neurons exhibited spontaneous APs with an amplitude of 77.72 +/- 2.7 mV (n = 20) and they fired at a frequency of 0.8 +/- 0.1 Hz (n = 11). Most cells had a prolonged early after-depolarization that followed an initial fully developed AP. Superfusion of Ang II (100 nM) plus losartan (LOS, 1 microM) to block Ang II type 1 receptors elicited a significant chronotropic effect that was reversed by the AT(2) receptor inhibitor PD 123,319 (1 microM). LOS alone had no effect on any of the parameters measured. The chronotropic effect of Ang II was reversed by the general LO inhibitor 5,8,11,14-eicosatetraynoic acid (10 microM) or by the selective PP-2A inhibitor okadaic acid (1 nM) and was mimicked by the 12-LO metabolite of AA 12-(S)-hydroxy-(5Z, 8Z, 10E, 14Z)-eicosatetraynoic acid. These data indicate that Ang II elicits an AT(2) receptor-mediated increase in neuronal firing rate, an effect that involves generation of LO metabolites of AA and activation of PP-2A.     

Involvement of Src tyrosine kinase and mitogen-activated protein kinase in the facilitation of calcium channels in rat nucleus of the tractus solitarius by angiotensin II

It is recognized that brain contains all the components of the renin–angiotensin systems (RAS). The nucleus of the tractus solitarius (NTS) is known to play a major role in the regulation of cardiovascular, respiratory, gustatory, hepatic and swallowing functions. Voltage-dependent Ca²⁺ channels (VDCCs) serve as crucial mediators of membrane excitability and Ca²⁺-dependent functions such as neurotransmitter release, enzyme activity and gene expression. The purpose of this study was to investigate the effects of angiotensin II (Ang II) on VDCC currents ( I _Ca) in the NTS using patch-clamp recording methods. An application of Ang II caused facilitation of L-type I _Ca in a concentration-dependent manner with an EC₅₀ of 167 n m and a Hill coefficient of 1.73. AT₁ receptor antagonist losartan antagonized the Ang II-induced facilitation of I _Ca. Intracellular dialysis of the Gα_i-protein antibody attenuated the Ang II-induced facilitation of I _Ca. Both Src tyrosine kinase inhibitor and mitogen-activated protein kinase (MAPK) inhibitor attenuated the Ang II-induced facilitation of I _Ca. p38 MAPK inhibitor also attenuated the Ang II-induced facilitation of I _Ca. These results indicate that Ang II facilitates L-type VDCCs via Gα_i-proteins involving Src tyrosine kinase and p38 MAPK kinase mediated by AT₁ receptors in NTS.     

Up-regulation of AT(1) and AT(2) receptors in postinfarcted hypertrophied myocytes and stretch-mediated apoptotic cell death

To determine whether up-regulation of AT(1) and AT(2) receptors occurred in hypertrophied myocytes after infarction and whether AT(2) played a role in stretch-mediated apoptosis, left ventricular myocytes were dissociated from the surviving portion of the wall 8 days after coronary occlusion and cardiac failure in rats. Control cells were obtained from sham-operated animals. Myocytes were stretched in an equibiaxial stretch apparatus and angiotensin II (Ang II) formation and cell death were measured 3 and 12 hours later. AT(1) and AT(2) proteins were evaluated in freshly isolated myocytes and after stretch. The effects of AT(1) and AT(2) antagonists on stretch-induced Ang II synthesis and apoptosis were also established. Myocardial infarction increased AT(1) and AT(2) in myocytes and stretch further up-regulated these receptors. Ang II levels were higher in postinfarcted myocytes and this peptide increased with the duration of stretch in both groups of cells. Similarly, apoptosis increased with time in control and postinfarcted myocytes. Absolute values of Ang II and apoptosis were greater in myocytes from infarcted hearts at 3 and 12 hours after stretch. Addition of AT(1) blocker to cultures inhibited stretch-activated apoptosis in both myocyte populations as well as the generation of Ang II in postinfarcted myocytes. In contrast, AT(2) antagonists had no impact on these cellular events. In conclusion, Ang II stimulated cell death through AT(1) receptor activation, whereas ligand binding to AT(2) receptor did not alter Ang II concentration and apoptosis in normal and postinfarcted hypertrophied myocytes.     

坎地沙坦阻断血管紧张素Ⅱ介导的原代急性髓样白血病细胞增殖的作用及机制

目的: 观察血管紧张素Ⅱ 1型受体(AT1R)拮抗剂坎地沙坦抑制血管紧张素Ⅱ(Ang Ⅱ)介导的原代急性髓样白血病(AML)细胞增殖的作用及机制。方法: MTT法观察Ang Ⅱ对原代AML细胞、正常骨髓单个核细胞增殖的影响以及坎地沙坦和AT2R拮抗剂 PD123319对AngII促原代AML细胞增殖的拮抗作用; Western blotting法观察坎地沙坦和PI3K抑制剂对原代AML细胞Akt磷酸化的影响。结果: AngII能剂量和时间依赖性促进原代AML细胞增殖(P<0.05),而对正常骨髓无此作用。坎地沙坦随浓度和时间依赖性阻断Ang II作用下白血病细胞增殖(P<0.05)。PI3K抑制剂可抑制Ang II促进原代AML细胞的增殖(P<0.05),坎地沙坦能明显下调Ang II增加原代AML细胞Akt的磷酸化水平(P<0.05)。结论: 坎地沙坦通过抑制PI3K/Akt信号转导途径抑制Ang II/AT1R介导的白血病细胞增殖。     

Activation of PKC increases Na+-K+ pump current in ventricular myocytes from guinea pig heart

 We have previously shown activation of α₁-adrenergic receptors increases Na⁺-K⁺ pump current (I _p) in guinea pig ventricular myocytes, and the increase is eliminated by blockers of phosphokinase C (PKC). In this study we examined the effect of activators of PKC on I _p. Phorbol 12-myristate 13-acetate (PMA), a PKC activator, increased I _P at each test potential without shifting its voltage dependence. The concentration required for a half-maximal response (K _0.5) was 6 μM at 15 nM cytosolic [Ca²⁺] ([Ca²⁺]_i) and13 nM at 314 nM [Ca²⁺]_i. The maximal increase at either [Ca²⁺]_i was about 30%. Another activator of PKC, 1,2-dioctanoyl-sn-glycerol (diC₈), increased I _p similarly. The effect of PMA on I _P was eliminated by the PKC inhibitor staurosporine, but not by the peptide PKI, an inhibitor of protein kinase A (PKA). PMA and α₁-adrenergic agonist effects both were sensitive to [Ca²⁺]_i, blocked by PKC inhibitors, unaffected by PKA inhibition, and increased I _p uniformly at all voltages. However, they differed in that α₁-activation caused a maximum increase of 15% vs 30% via PMA, and α₁-effects were less sensitive to [Ca²⁺]_i than PMA effects. These results demonstrate that activation of PKC causes an increase in I _p in guinea pig ventricular myocytes. Moreover, they suggest that the coupling of α₁-adrenergic activation to I _p is entirely through PKC, however α₁-activation may be coupled to a specific population of PKC whereas PMA is a more global agonist. Received: 21 August 1998 / Received after revision: 7 December 1998 / Accepted: 11 December 1998     

Sustained beta-adrenergic stimulation increased L-type Ca2+ channel expression in cultured quiescent ventricular myocytes

The abundance of voltage-gated L-type Ca2+ channels is altered by beta-adrenergic receptor (beta-AR) stimulation and by an elevation of the intracellular Ca2+ concentration in cardiac myocytes. In whole animal, chronic beta-AR stimulation or pacing heart results in various changes in the abundance of the channel, but it reduces the beta-AR responsiveness of the L-type channel. Because beta-AR stimulation facilitates the L-type calcium channels, it is difficult in the whole animal to study the effects of beta-AR and Ca2+ influx on the upregulation of the L-type channel independently of each other, which makes the culture of nonbeating adult myocytes an attractive model. We found that culturing quiescent adult rabbit ventricular myocytes with isoproterenol (ISO, 2 microM) for 72 h or more caused a significant increase in the expression of mRNA coding for the L-type channel alpha(1C) subunit by approximately twofold as compared to time-matched controls, and it was followed by a 1.8-fold increase in the Ca2+ current density at 96 h. Somewhat surprisingly, an acute application of 1 microM ISO increased the current amplitude even in ISO-treated cells. The increase in the current density, induced by sustained beta-AR stimulation, was blocked by a beta-AR antagonist, propranolol (10 microM), but not by a Ca2+ antagonist, nitrendipine (10 microM). In addition, the effects were reproduced by forskolin (10 microM), but not by a Ca2+ agonist, Bay-K 8644 (2 microM). Taken together, these results suggest that sustained beta-AR stimulation upregulates L-type channel expression, but does not alter the beta-AR responsiveness of the channel in quiescent myocytes.     

Mechanisms of angiotensin II signaling on cytoskeleton of podocytes

Podocytes are significant in establishing the glomerular filtration barrier. Sustained rennin–angiotensin system (RAS) activation is crucial in the pathogenesis of podocyte injury and causes proteinuria. This study demonstrates that angiotensin II (Ang II) caused a reactive oxygen species (ROS)-dependent rearrangement of cortical F-actin and a migratory phenotype switch in cultured mouse podocytes with stable Ang II type 1 receptor (AT1R) expression. Activated small GTPase Rac-1 and phosphorylated ezrin/radixin/moesin (ERM) proteins provoked Ang II-induced F-actin cytoskeletal remodeling. This work also shows increased expression of Rac-1 and phosphorylated ERM proteins in cultured podocytes, and in glomeruli of podocyte-specific AT1R transgenic rats (Neph-hAT1 TGRs). The free radical scavenger DMTU eliminated Ang II-induced cell migration, ERM protein phosphorylation and cortical F-actin remodeling, indicating that ROS mediates the influence of Rac-1 on podocyte AT1R signaling. Heparin, a potent G-coupled protein kinase 2 inhibitor, was found to abolish ERM protein phosphorylation and cortical F-actin ring formation in Ang II-treated podocytes, indicating that phosphorylated ERM proteins are the cytoskeletal effector in AT1R signaling. Moreover, Ang II stimulation triggered down-regulation of α actinin-4 and reduced focal adhesion expression in podocytes. Signaling inhibitor assay of Ang II-treated podocytes reveals that Rac-1, RhoA, and F-actin reorganization were involved in expressional regulation of α actinin-4 in AT1R signaling. With persistent RAS activation, the Ang II-induced phenotype shifts from being dynamically stable to adaptively migratory, which may eventually exhaust podocytes with a high actin cytoskeletal turnover, causing podocyte depletion and focal segmental glomerulosclerosis. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Effects of mibefradil, a selective T-type Ca2+ channel antagonist, on sino-atrial node and ventricular myocardia

The effects of mibefradil, a non-dihydropyridine Ca2+ channel antagonist, on the action potential configuration of isolated rabbit sino-atrial node preparations, membrane currents of guinea-pig ventricular myocytes and the contractile force of isolated ventricular papillary muscles were examined. In sino-atrial node preparations, 10 microM mibefradil decreased the slope of the pacemaker depolarization (phase 4 depolarization) and maximum rate of rise, and shifted the threshold potential to the positive direction with no effect on action potential duration. In ventricular myocytes, 1 microM mibefradil inhibited the T-type Ca2+ current by about 40% while it had no effect on the L-type Ca2+ current. At 10 microM, mibefradil inhibited the L-type and T-type Ca2+ currents by about 40% and 90%, respectively. Mibefradil had no effect on contractile force at concentrations up to 1 microM. Thus, mibefradil was shown to produce potent prolongation of the pacemaker depolarization, mainly through inhibition of the T-type Ca2+ current. It is suggested that the T-type Ca2+ current may not be involved in ventricular contraction.     

Protein kinase C suppresses spontaneous, transient, outwards K+ currents through modulation of the Na/Ca exchanger in guinea-pig gastric myocytes

The effect of protein kinase C (PKC) on the Ca2+-activated K+ current (IK,Ca) in guinea-pig gastric myocytes was studied using the whole-cell voltage-clamp technique. At a holding potential of 0 mV, IK,Ca, recorded as spontaneous, transient, outwards currents (STOCs), was markedly inhibited, both in mean amplitude (54 +/- 5%) and frequency (60 +/- 8%) by 1 microM phorbol 12, 13 dibutyrate (PDBu, n = 6). These effects were antagonized by pretreatment with 10 nM bisindolylmaleimide I (n = 5), a selective inhibitor of PKC. The possibility that the inhibition of STOCs was due to direct channel inhibition by PKC was addressed using inside-out or open-cell-attached patch-clamp techniques, the latter established using beta-escin. PDBu did not alter the conductance or open probability of the KCa channel in any mode, suggesting that PKC does not inhibit the KCa channel directly. To study the involvement of the Na/Ca exchanger in the inhibition of STOCs by PDBu, its operation was prevented by replacing Na+ in the internal solution by tris(hydroxymethyl)aminomethane (TRIS) and external Na+ by equimolar K+ and Ca2+-activated inwards K+ currents recorded at a holding potential of 0 mV. Neither the mean amplitude (96 +/- 8%) nor the frequency of these currents was inhibited significantly by 1 microM PDBu (n = 5). Like PDBu, 5 microM 2-(2-[4-(4-nitrobenzyloxy)phenyl]ethyl) isothiourea methanesulphonate (KB-R7943), a selective inhibitor of the reverse mode Na/Ca exchanger, also inhibited the mean amplitude (45 +/- 6%) and frequency (26 +/- 2%) of STOCs at the holding potential of 0 mV (n=6). The results suggest that the suppression of STOCs by PKC is mediated by inhibition of the Na/Ca exchanger.     

Endothelium-derived contracting factors mediate the Ang II-induced endothelial dysfunction in the rat aorta: preventive effect of red wine polyphenols

Angiotensin II (Ang II)-induced hypertension is associated with vascular oxidative stress and an endothelial dysfunction. This study examined the role of reactive oxygen species (ROS) and endothelium-derived contracting factors in Ang II-induced endothelial dysfunction and whether these effects are prevented by red wine polyphenols (RWPs), a rich source of natural antioxidants. Rats were infused with Ang II for 14 days. RWPs were administered in the drinking water 1 week before and during the Ang II infusion. Arterial pressure was measured in conscious rats. Vascular reactivity was assessed in organ chambers and cyclooxygenase-1 (COX-1) and COX-2 expression by Western blot and immunofluorescence analyses. Ang II-induced hypertension was associated with blunted endothelium-dependent relaxations and induction of endothelium-dependent contractions in the presence of nitro-L-arginine in response to acetylcholine (Ach). These effects were not affected by the combination of membrane permeant analogs of superoxide dismutase and catalase but were abolished by the thromboxane A₂ (TP) receptor antagonist GR32191B and the COX-2 inhibitor NS-398. The COX-1 inhibitor SC-560 also prevented contractile responses to Ach. Ang II increased the expression of COX-1 and COX-2 in the aortic wall. RWPs prevented Ang II-induced hypertension, endothelial dysfunction, and upregulation of COX-1 and COX-2. Thus, Ang II-induced endothelial dysfunction cannot be explained by an acute formation of ROS reducing the bioavailability of nitric oxide but rather by COX-dependent formation of contracting factors acting on TP receptors. RWPs are able to prevent the Ang II-induced endothelial dysfunction mostly due to their antioxidant properties.     

Angiotensin II stimulates proliferation of adventitial fibroblasts cultured from rat aortic explants

It has been proposed that the local renin-angiotensin system is activated in the adventitia after vascular injury. However, the physiological role of Angiotensin II (Ang II) in the adventitia has not been studied at a cellular level. This study was designed to assess the role of Ang II in the growth response of cultured adventitial fibroblasts (AFs). Adventitial explants of the rat thoracic aorta showed outgrowth of AFs within 5-7 days. Ang II caused hyperplastic response of AF cultures. The Ang II-induced mitogenic response of AFs was mediated primarily by the AT1 receptor. Ang II caused a rapid induction of immediate early genes (c-fos, c-myc and jun B). Induction of c-fos expression was fully blocked by an AT1 receptor antagonist but not by an AT2 receptor antagonist. Epidermal growth factor (EGF), platelet-derived growth factor-BB (PDGF-BB) and basic fibroblast growth factor (bFGF) induced DNA synthesis in AFs. Co-stimulation of AFs with the growth factors and Ang II potentiated the incorporation of 3H-thymidine into DNA. Results from this study indicate that Ang II causes mitogenesis of AFs via AT1 receptor stimulation and potentiates the responses to other mitogens. These data suggest that the Ang II may play an important role in regulating AF function during vascular remodeling following arterial injury.     

Role of Ca2+-Activated Cl− Current in Ventricular Action Potentials of Sheep During Adrenoceptor Stimulation

Adrenoceptor stimulation enhances repolarising and depolarising membrane currents to different extents in cardiac myocytes. We investigated the opposing effects of the repolarising Ca(2+)-activated Cl(-) current (I(Cl(Ca))) and depolarising L-type Ca(2+) current (I(Ca,L)) on the action potential configuration of sheep ventricular myocytes stimulated with noradrenaline. Whole-cell current-clamp recordings revealed that noradrenaline accelerated and prolonged phase-1 repolarisation. We define the minimal potential at the end of phase-1 repolarisation as "notch level". Noradrenaline (1 microM) caused the notch level to fall from 14 +/- 2.6 to 7.8 +/- 2.8 mV (n = 24), but left action potential duration, resting membrane potential or action potential amplitude unaffected. Whole-cell voltage-clamp recordings showed that 1 microM noradrenaline increased both I(Ca,L) and I(Cl(Ca)), but it had no significant effect on the principal K(+) currents. Blockage of I(Cl(Ca)) by 0.5 mM 4,4'-diisothiocyanatostilbene-2,2'-disulphonic acid (DIDS) in both the absence and the presence of noradrenaline abolished phase-1 repolarisation. In the presence of noradrenaline, DIDS caused elevation of the plateau phase amplitude and an increase in the action potential duration. In conclusion, elevation of the plateau phase amplitude and action potential prolongation associated with an increased I(Ca,L) upon adrenoceptor stimulation is prevented by an increased I(Cl(Ca)) in sheep ventricular myocytes. Experimental Physiology (2001) 86.2, 151-159.     

Dual repressive effect of angiotensin II-type 1 receptor blocker telmisartan on angiotensin II-induced and estradiol-induced uterine leiomyoma cell proliferation

BACKGROUND: Although uterine leiomyomas or fibroids are the most common gynecological benign tumor and greatly affect reproductive health and well-being, the pathophysiology and epidemiology of uterine leiomyomas are poorly understood. Elevated blood pressure has an independent, positive association with risk for clinically detected uterine leiomyoma. Angiotensin II (Ang II) is a key biological peptide in the renin-angiotensin system that regulates blood pressure. METHODS: In this study, we investigated the potential role of Ang II (1-1000 nM) in the proliferation of rat ELT-3 leiomyoma cells in vitro. RT-PCR and western blot analysis with cell proliferation and DNA transfection assays were performed to determine the mechanism of action of Ang II. RESULTS: Ang II induced ELT-3 leiomyoma cell proliferation (P < 0.01) and the expression of Ang II type 1 receptor (AT(1)R) and AT(2)R mRNA and protein was confirmed. Regarding the intracellular signaling pathway, the Ang II-induced cell proliferation was AT(1)R-, epidermal growth factor receptor-, extracellular-regulated kinase- and protein kinase C-dependent but was not dependent on the AT(2)R or phosphatidylinositol-3 kinase or JAK kinase. The AT(1)R blocker telmisartan, effectively repressed Ang II-induced and estradiol-induced cell proliferation (P < 0.01). AT(1)R, but not AT(2)R, plays a role in Ang II-induced ELT-3 cell proliferation. CONCLUSIONS: These experimental findings in vitro highlight the potential role of Ang II in the proliferation of leiomyoma cells.