Block of Inward Rectifying K+ Channels (KIR) Inhibits Bradykinin-Induced Vasodilatation in Human Forearm Resistance Vasculature
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《动脉硬化血栓血管生物学》
From the Department of Clinical Pharmacology, GKT Division of Cardiovascular Medicine, St Thomas’ Hospital, London, United Kingdom.
Correspondence to J.M. Ritter, Department of Clinical Pharmacology, St Thomas’ Hospital, Lambeth Palace Road, London, SE1 7EH, UK. E-mail james.ritter@kcl.ac.uk
Abstract
Objective— To investigate the possible involvement of inward rectifying K+ channels (KIR) in the response of human resistance vessels to bradykinin in vivo.
Methods and Results— Drugs were administered via the brachial artery in healthy male volunteers and forearm blood flow was measured by venous occlusion plethysmography. Inhibition of KIR by barium chloride (4 μmol min–1) alone or with additional inhibition of Na+/K+ ATPase (ouabain 2.7 μmol min–1) reduced responses to bradykinin (30 pmol min–1), by 26±8.3% and 36±7.2%, respectively (each P<0 0.05). Barium with ouabain plus inhibitors of prostaglandin (PG) and nitric oxide synthesis inhibited but did not abolish responses to bradykinin (51±2.8% inhibition; P<0.01); norepinephrine (240 pmol min–1) caused similar reduction of baseline blood flow, as did this combination of inhibitors, but did not significantly inhibit the response to bradykinin. Barium plus ouabain did not significantly reduce responses to acetylcholine or albuterol.
Conclusion— A component of the vasodilator response to bradykinin in human forearm vasculature is mediated by KIR.
The possible involvement of inward-rectifying K+ channels (KIR) in the action of bradykinin was investigated by administering drugs via the brachial artery in healthy men. Barium selectively inhibited the forearm blood flow response to bradykinin, indicating that a component of this response is mediated by KIR.
Key Words: bradykinin ? barium ? forearm vasculature ? inward-rectifying potassium channels ? hyperpolarizing factor
Introduction
Potassium ion (K+) channel activity is an important determinant of the membrane potential of vascular smooth muscle and, thus, of vascular tone. Inward-rectifying K+ channels (KIR) in vascular smooth muscle differ from other vascular smooth muscle K+ channels (Ca2+-activated K+ channels, KCa, voltage-dependent K+ channels, KV, and ATP-sensitive K+ channels, KATP) in their unique current voltage properties and the consequent hyperpolarizing effect of increased external K+.1–3 KIR channels play a role in K+-mediated dilation of rat coronary and cerebral arteries4 and contribute to basal vasodilator tone in human forearm resistance vasculature.5 KIR is more sensitive to Ba2+ than other K+ channels, half-block being achieved at 2 μmol L–1 at –60 mV in rat cerebral artery vascular smooth muscle,6 50-times more potent than on KATP channels, >500-times as potent as on KV channels, and >5000-times as potent as on KCa channels.7 Brachial artery infusion of barium chloride in a dose that increases the local mean plasma concentration of Ba2+ to 50 μmol L–1 inhibits the vasodilator response to infused potassium chloride by 60±9%.5 Electrogenic Na+/K+ exchange also contributes to hyperpolarizing responses to K+, and ouabain, an inhibitor of Na+/K+ ATPase, inhibits forearm vasodilator responses to KCl by 33% in healthy men.8 The combination of Ba2+ plus ouabain in the same doses inhibits responses to K+ almost completely.5 Coinfusion of barium chloride with ouabain thus provides a pharmacological tool to investigate whether increased extracellular K+ concentration contributes to vasodilator responses.
Activation of vascular smooth muscle KIR and Na+/K+ ATPase by K+ released from endothelial cells causes endothelium-dependent hyperpolarization in rat hepatic arteries in vitro,9 but the contribution, if any, of K+ to endothelium-dependent vasodilator agonists in vivo is less clearly established. Bradykinin is an endothelium-dependent vasodilator. In addition to releasing prostacyclin10 and nitric oxide (NO)11 from endothelial cells, it causes endothelium-dependent hyperpolarization of human coronary artery despite inhibition of prostaglandin (PG) and NO synthesis.12 When infused via the brachial artery, it is a potent vasodilator in human forearm vasculature13 by an action on B2 receptors.14 Bradykinin-induced vasodilation in this vascular bed is not inhibited by aspirin15 and is incompletely blocked by L-NG monomethyl-arginine (L-NMMA).16–19 Ouabain does not inhibit forearm responses to bradykinin in normotensive subjects but does significantly reduce such responses in patients with essential hypertension.19 In the present investigation, we determined effects of Ba2+ with or without ouabain on bradykinin-induced vasodilation in healthy normotensive men without known risk factors for atheromatous vascular disease. Experiments were performed with or without inhibitors of NO and PG synthesis. Norepinephrine was used to control for nonspecific physiological antagonism caused by vasoconstriction caused by the inhibitors. Sensitivity of bradykinin to Ba2+ plus ouabain was compared with 2 other endothelium-dependent agonists (acetylcholine and albuterol) that activate NO synthesis by distinct mechanisms in this vascular bed.20–22
Methods
The St Thomas’ Hospital Research Ethics Committee approved the studies and all subjects gave written informed consent. Healthy men aged 33±11 years (mean±SD), nonsmokers using no medications, were invited to take part. Subjects were normotensive (blood pressure <130/80 mm Hg) and normocholesterolemic (total cholesterol <5.2 mmol L–1). Forearm studies were performed as described previously.5 Blood flow was measured in both arms using venous occlusion plethysmography, and drugs dissolved in physiological saline were infused into the left brachial artery via a 27-gauge steel cannula. Each protocol was performed in a separate group of subjects to limit exposure to barium. Barium infusions (4 μmol min–1) were given for 6 minutes. In each protocol, saline was first administered for 18 minutes to establish baseline flow. Vasodilators were infused for 3 minutes. Bradykinin (30 pmol min–1) was administered, followed by saline for 18 minutes, to re-establish baseline flow. Inhibitors were then administered for 3 minutes alone and for a final 3 minutes with bradykinin. Inhibitors were: Ba2+ alone in the first protocol; Ba2+ with ouabain (2.7 μmol min–1) in the second; Ba2+, indomethacin (0.34 μmol min–1),23 ouabain, and L-NMMA (64 μmol min–1)24 in the third; and norepinephrine (240 pmol min–1) in the fourth. In control experiments to exclude desensitization, bradykinin was infused twice as before but with saline rather than inhibitor. In 2 separate studies, acetylcholine (33 nmol min–1) or albuterol (1 nmol min–1) were given instead of bradykinin, and Ba2+ with ouabain tested as a potential inhibitor. Results are expressed as means±SEM. Percent inhibition was calculated for each individual subject as: [(FBFcontrol –FBFinhib)/ (FBFcontrol)] x 100%, where FBFcontrol and FBFinhib were the forearm blood flow in the absence and presence of inhibitor. Differences were analyzed using Student paired t test (2-sided). P<0.05 was taken as significant.
Results
There were no adverse events or electrocardiographic changes. Arterial blood pressure and blood flow in the noninfused arm did not change significantly. Mean blood flows in the infused arm are summarized in Table 1. Bradykinin increased blood flow similarly in each protocol, and each inhibitor reduced blood flow significantly. When co-infused with norepinephrine, the vasodilator effect of bradykinin was not significantly inhibited. Ba2+ alone inhibited the vasodilator response to bradykinin by 26±8.3% (P<0.05), barium plus ouabain inhibited the response by 36±7.2% (P<0.05), and barium plus ouabain, indomethacin, and L-NMMA inhibited the response by 51±2.8% (P<0.01). When bradykinin was infused twice using the same protocol but in the absence of inhibitors, there was no evidence of desensitization: the response to the first versus second infusion was 8.6±1.7 versus 8.6±1.8 mL min–1 100 mL forearm–1 (n=6, P=NS). Ba2+ plus ouabain did not significantly reduce vasodilator responses to acetylcholine or to albuterol (Table 2, which also shows the effect of these inhibitors on bradykinin for comparison).
TABLE 1. Effects of Inhibitors of KIR, Na+/K+ ATPase, NO Synthase, and Cyclooxygenase on Responses to Bradykinin
TABLE 2. Effects of Inhibition of KIR and Na+/K+ ATPase on Responses to Bradykinin, Acetylcholine, and Albuterol
Discussion
The main finding is that Ba2+ (plasma concentration 50 μmol L–1)5 selectively inhibits forearm blood flow responses to bradykinin. Norepinephrine does not significantly inhibit responses to bradykinin, and Ba2+ plus ouabain does not significantly inhibit acetylcholine or albuterol. This implicates KIR in the vasodilator response to bradykinin in human forearm resistance vasculature. Ba2+ with ouabain in the doses used almost completely abolishes vasodilator responses to K+5, but in the present experiments coinfusion of ouabain with Ba2+ does not completely inhibit the response to bradykinin, and responses to bradykinin are not abolished even when Ba2+ and ouabain are given with indomethacin and L-NMMA in doses that block forearm PG synthesis and nitric oxide-mediated responses to acetylcholine.23,24 The simplest explanation is that bradykinin dilates this vascular bed partly but not entirely through activation of KIR. The residual vasodilator response to bradykinin in the presence of inhibitors points to the possible involvement of a direct vasodilator action of bradykinin on vascular smooth muscle or of mediator pathways distinct from NO, prostaglandins, or K+ ions.
Activation of KIR by bradykinin could be via increased K+ concentration in the interstitial extracellular space in resistance arteries. The present experiments do not define the cellular origin of such increased interstitial K+ concentration. This could be the endothelium, as in rat hepatic artery in vitro,9,25 consistent with an EDHF/K+-mediated mechanism of bradykinin in the forearm in vivo. This agrees with the observation that a dose of tetraethylammonium expected to give a plasma concentration of 2x10–4 mol L–1 inhibits but does not abolish responses to bradykinin in this vascular bed.18 This concentration of tetraethylammonium could inhibit K+ efflux from endothelium via an action on KCa channels, where it produces half the maximum block at 2x10–4 mol L–1.7
Limitations
A constraint was our concern to limit the exposure of volunteers to Ba2+. The total infused dose of 24 μmol is less than one-tenth the chronic oral reference dose calculated by the Environmental Protection Agency. Limiting the dose of Ba2+ in this way meant that we were not able to explore its effects on a range of doses of bradykinin. A limitation of norepinephrine as a control is that it can inhibit KIR in vascular smooth muscle; other vasoconstrictors may influence KIR by an effect on membrane potential. Another limitation is that the present experiments do not identify the cellular distribution of the KIR channel involved in the vasodilator action of bradykinin in the forearm. This may be important because this channel is expressed not only in vascular smooth muscle but also in endothelial cells.26
In conclusion, a dose of Ba2+ that selectively inhibits KIR inhibits forearm vasodilator responses to bradykinin in healthy men, evidence that a component of this response is mediated by activation of KIR. This is consistent with bradykinin acting through K+ and/or another KIR-dependent EDHF in human forearm resistance vessels in vivo.
Acknowledgments
This work was supported by the British Heart Foundation.
Received June 8, 2004; accepted November 3, 2004.
References
Bradley KK, Jaggar JH, Bonev AD, Heppner TJ, Flynn ERM, Nelson MT, Horowitz B. Kir 2.1 encodes the inward rectifier potassium channel in rat arterial smooth muscle cells. J Physiol. 1999; 515: 639–651.
Zaritsky JJ, Eckman DM, Wellman GC, Nelson MT, Schwartz TL. Targeted disruption of Kir 2.1 and Kir 2.2 genes reveals the essential role of the inwardly rectifying K+ current in K+-mediated vasodilation. Circ Res. 2000; 87: 160–166.
Lopatin AN, Makhina EN, Nichols CG. Potassium channel block by cytoplasmic polyamines as the mechanism of intrinsic rectification. Nature. 1994; 372: 366–369.
Knot, HJ, Zimmerman PA, Nelson MT. Extracellular K+-induced hyperpolarizations and dilatations of rat coronary and cerebral arteries involve inward rectifier K+ channels. J Physiol (Lond). 1996; 492: 419–430.
Dawes M, Sieniawska C, Delves T et al. Barium reduces resting blood flow and inhibits potassium-induced vasodilation in the human forearm, Circulation. 2002; 105: 1323–1328.
Quayle JM, McCarron JG, Asbury JR, Nelson MT. Inward rectifier K+ currents in smooth muscle cells from rat resistance-sized cerebral arteries. Am J Physiol. 1993; 265: C1363–C1370.
Nelson MT, Quayle JM. Physiological roles and properties of potassium channels in arterial smooth muscle. Am J Physiol. 1995; 268: C799–C822.
Robinson BF, Phillips RJW, Wilson PN, Chiodini PL. Effect of local infusion of ouabain on human forearm vascular resistance and on response to potassium, verapamil and sodium nitroprusside. J Hypertens. 1983; 1: 165–169.
Edwards G, Dora KA, Gardener MJ, Garland CJ, Weston AH. K+ is an endothelium-derived hyperpolarizing factor in rat arteries, Nature. 1998; 396: 269–272.
Hong SL. Effect of bradykinin and thrombin on prostacyclin synthesis in endothelial cells from calf and pig aorta and human umbilical cord vein. Thromb Res. 1980; 18: 787–795.
Palmer RMJ, Ashton DS, Moncada S. Vascular endothelial cells synthesize nitric oxide from L-arginine. Nature. 1988; 333: 664–666.
Nakashima M, Mombouli J-V, Taylor AA, Vanhoutte PM. Endothelium-dependent hyperpolarization caused by bradykinin in human coronary arteries, J Clin Invest. 1993; 92: 2867–2871.
Fox RH, Goldsmith R, Kidd DJ. Lewis platelet glycoprotein (GP). Bradykinin as a vasodilator in man. J Physiol. 1961; 157: 589–602.
Cockcroft JR, Chowienczyk PJ, Brett SE, Bender N, Ritter JM. Inhibition of bradykinin-induced vasodilation in human forearm vasculature by icatibant, a potent B2-receptor antagonist. Br J Clin Pharmacol. 1994; 38: 317–322.
Benjamin N, Cockcroft JR, Collier JG, Dollery CT, Ritter JM, Webb DJ. Local inhibition of converting enzyme and vascular responses to angiotensin and bradykinin in the human forearm. J Physiol. 1989; 412: 543–555.
Cockcroft JR, Chowienczyk PJ, Brett SE, Ritter JM. Effect of NG monomethyl L arginine on kinin-induced vasodilation in the human forearm. Br J Clin Pharmacol. 1994; 38: 307–310.
O’Kane KP, Webb DJ, Collier JG, Vallance PJ. Local L-NG monomethyl-arginine attenuates the vasodilator action of bradykinin in the human forearm, Br J Clin Pharmacol. 1994; 38: 311–315.
Honing MLH, Smits P, Morrison PJ, Rabelink TJ. Bradykinin-induced vasodilation of human forearm resistance vessels is primarily mediated by endothelium-dependent hyperpolarisation. Hypertension. 2000; 35: 1314–1318.
Taddei S, Ghiadoni L, Virdis A, Buralli S, Salvetti A. Vasodilation to bradykinin is mediated by an ouabain-sensitive pathway as a compensatory mechanism for impaired nitric oxide availability in essential hypertensive patients. Circulation. 1999; 100: 1400–1405.
Vallance P, Collier J, Moncada S. Effects of endothelium-derived nitric oxide on peripheral arteriolar tone in man. Lancet. 1989; 2: 997–1000.
Dawes M, Chowienczyk PJ, Ritter JM. Effects of inhibition of the L-arginine/ nitric oxide pathway on vasodilation caused by ?-adrenergic agonists in human forearm. Circulation. 1997; 95: 2293–2297.
Ferro A, Queen LR, Priest RM, Xu B, Ritter JM, Poston P, Ward JPT. Activation of nitric oxide synthase by ?2-adrenoceptors in human umbilical vein endothelium in vitro. Br J Pharmacol. 1999; 126: 1872–1880.
Wilson JR, Kapoor SC. Contribution of prostaglandins to exercise-induced vasodilation in humans. Am J Physiol. 1993; 265: H171–H175.
Dawes M, Chowienczyk PJ, Ritter JM. Quantitative aspects of the inhibition by NG monomethyl L-arginine of responses to endothelium-dependent vasodilators in human forearm vasculature. Br J Pharmacol. 2001; 134: 939–944.
Busse R, Edwards G, Félétou M, Fleming I, Vanhoutte PM, Weston AH. EDHF: Bringing the concepts together. Trends in Pharmacological Sciences. 2002; 23: 374–380.
Nilius B, Droogmans G. Ion channels and their functional role in vascular endothelium. Physiol Rev. 2001; 81: 1415–1459.(R. Dwivedi; S. Saha; P.J.)
Correspondence to J.M. Ritter, Department of Clinical Pharmacology, St Thomas’ Hospital, Lambeth Palace Road, London, SE1 7EH, UK. E-mail james.ritter@kcl.ac.uk
Abstract
Objective— To investigate the possible involvement of inward rectifying K+ channels (KIR) in the response of human resistance vessels to bradykinin in vivo.
Methods and Results— Drugs were administered via the brachial artery in healthy male volunteers and forearm blood flow was measured by venous occlusion plethysmography. Inhibition of KIR by barium chloride (4 μmol min–1) alone or with additional inhibition of Na+/K+ ATPase (ouabain 2.7 μmol min–1) reduced responses to bradykinin (30 pmol min–1), by 26±8.3% and 36±7.2%, respectively (each P<0 0.05). Barium with ouabain plus inhibitors of prostaglandin (PG) and nitric oxide synthesis inhibited but did not abolish responses to bradykinin (51±2.8% inhibition; P<0.01); norepinephrine (240 pmol min–1) caused similar reduction of baseline blood flow, as did this combination of inhibitors, but did not significantly inhibit the response to bradykinin. Barium plus ouabain did not significantly reduce responses to acetylcholine or albuterol.
Conclusion— A component of the vasodilator response to bradykinin in human forearm vasculature is mediated by KIR.
The possible involvement of inward-rectifying K+ channels (KIR) in the action of bradykinin was investigated by administering drugs via the brachial artery in healthy men. Barium selectively inhibited the forearm blood flow response to bradykinin, indicating that a component of this response is mediated by KIR.
Key Words: bradykinin ? barium ? forearm vasculature ? inward-rectifying potassium channels ? hyperpolarizing factor
Introduction
Potassium ion (K+) channel activity is an important determinant of the membrane potential of vascular smooth muscle and, thus, of vascular tone. Inward-rectifying K+ channels (KIR) in vascular smooth muscle differ from other vascular smooth muscle K+ channels (Ca2+-activated K+ channels, KCa, voltage-dependent K+ channels, KV, and ATP-sensitive K+ channels, KATP) in their unique current voltage properties and the consequent hyperpolarizing effect of increased external K+.1–3 KIR channels play a role in K+-mediated dilation of rat coronary and cerebral arteries4 and contribute to basal vasodilator tone in human forearm resistance vasculature.5 KIR is more sensitive to Ba2+ than other K+ channels, half-block being achieved at 2 μmol L–1 at –60 mV in rat cerebral artery vascular smooth muscle,6 50-times more potent than on KATP channels, >500-times as potent as on KV channels, and >5000-times as potent as on KCa channels.7 Brachial artery infusion of barium chloride in a dose that increases the local mean plasma concentration of Ba2+ to 50 μmol L–1 inhibits the vasodilator response to infused potassium chloride by 60±9%.5 Electrogenic Na+/K+ exchange also contributes to hyperpolarizing responses to K+, and ouabain, an inhibitor of Na+/K+ ATPase, inhibits forearm vasodilator responses to KCl by 33% in healthy men.8 The combination of Ba2+ plus ouabain in the same doses inhibits responses to K+ almost completely.5 Coinfusion of barium chloride with ouabain thus provides a pharmacological tool to investigate whether increased extracellular K+ concentration contributes to vasodilator responses.
Activation of vascular smooth muscle KIR and Na+/K+ ATPase by K+ released from endothelial cells causes endothelium-dependent hyperpolarization in rat hepatic arteries in vitro,9 but the contribution, if any, of K+ to endothelium-dependent vasodilator agonists in vivo is less clearly established. Bradykinin is an endothelium-dependent vasodilator. In addition to releasing prostacyclin10 and nitric oxide (NO)11 from endothelial cells, it causes endothelium-dependent hyperpolarization of human coronary artery despite inhibition of prostaglandin (PG) and NO synthesis.12 When infused via the brachial artery, it is a potent vasodilator in human forearm vasculature13 by an action on B2 receptors.14 Bradykinin-induced vasodilation in this vascular bed is not inhibited by aspirin15 and is incompletely blocked by L-NG monomethyl-arginine (L-NMMA).16–19 Ouabain does not inhibit forearm responses to bradykinin in normotensive subjects but does significantly reduce such responses in patients with essential hypertension.19 In the present investigation, we determined effects of Ba2+ with or without ouabain on bradykinin-induced vasodilation in healthy normotensive men without known risk factors for atheromatous vascular disease. Experiments were performed with or without inhibitors of NO and PG synthesis. Norepinephrine was used to control for nonspecific physiological antagonism caused by vasoconstriction caused by the inhibitors. Sensitivity of bradykinin to Ba2+ plus ouabain was compared with 2 other endothelium-dependent agonists (acetylcholine and albuterol) that activate NO synthesis by distinct mechanisms in this vascular bed.20–22
Methods
The St Thomas’ Hospital Research Ethics Committee approved the studies and all subjects gave written informed consent. Healthy men aged 33±11 years (mean±SD), nonsmokers using no medications, were invited to take part. Subjects were normotensive (blood pressure <130/80 mm Hg) and normocholesterolemic (total cholesterol <5.2 mmol L–1). Forearm studies were performed as described previously.5 Blood flow was measured in both arms using venous occlusion plethysmography, and drugs dissolved in physiological saline were infused into the left brachial artery via a 27-gauge steel cannula. Each protocol was performed in a separate group of subjects to limit exposure to barium. Barium infusions (4 μmol min–1) were given for 6 minutes. In each protocol, saline was first administered for 18 minutes to establish baseline flow. Vasodilators were infused for 3 minutes. Bradykinin (30 pmol min–1) was administered, followed by saline for 18 minutes, to re-establish baseline flow. Inhibitors were then administered for 3 minutes alone and for a final 3 minutes with bradykinin. Inhibitors were: Ba2+ alone in the first protocol; Ba2+ with ouabain (2.7 μmol min–1) in the second; Ba2+, indomethacin (0.34 μmol min–1),23 ouabain, and L-NMMA (64 μmol min–1)24 in the third; and norepinephrine (240 pmol min–1) in the fourth. In control experiments to exclude desensitization, bradykinin was infused twice as before but with saline rather than inhibitor. In 2 separate studies, acetylcholine (33 nmol min–1) or albuterol (1 nmol min–1) were given instead of bradykinin, and Ba2+ with ouabain tested as a potential inhibitor. Results are expressed as means±SEM. Percent inhibition was calculated for each individual subject as: [(FBFcontrol –FBFinhib)/ (FBFcontrol)] x 100%, where FBFcontrol and FBFinhib were the forearm blood flow in the absence and presence of inhibitor. Differences were analyzed using Student paired t test (2-sided). P<0.05 was taken as significant.
Results
There were no adverse events or electrocardiographic changes. Arterial blood pressure and blood flow in the noninfused arm did not change significantly. Mean blood flows in the infused arm are summarized in Table 1. Bradykinin increased blood flow similarly in each protocol, and each inhibitor reduced blood flow significantly. When co-infused with norepinephrine, the vasodilator effect of bradykinin was not significantly inhibited. Ba2+ alone inhibited the vasodilator response to bradykinin by 26±8.3% (P<0.05), barium plus ouabain inhibited the response by 36±7.2% (P<0.05), and barium plus ouabain, indomethacin, and L-NMMA inhibited the response by 51±2.8% (P<0.01). When bradykinin was infused twice using the same protocol but in the absence of inhibitors, there was no evidence of desensitization: the response to the first versus second infusion was 8.6±1.7 versus 8.6±1.8 mL min–1 100 mL forearm–1 (n=6, P=NS). Ba2+ plus ouabain did not significantly reduce vasodilator responses to acetylcholine or to albuterol (Table 2, which also shows the effect of these inhibitors on bradykinin for comparison).
TABLE 1. Effects of Inhibitors of KIR, Na+/K+ ATPase, NO Synthase, and Cyclooxygenase on Responses to Bradykinin
TABLE 2. Effects of Inhibition of KIR and Na+/K+ ATPase on Responses to Bradykinin, Acetylcholine, and Albuterol
Discussion
The main finding is that Ba2+ (plasma concentration 50 μmol L–1)5 selectively inhibits forearm blood flow responses to bradykinin. Norepinephrine does not significantly inhibit responses to bradykinin, and Ba2+ plus ouabain does not significantly inhibit acetylcholine or albuterol. This implicates KIR in the vasodilator response to bradykinin in human forearm resistance vasculature. Ba2+ with ouabain in the doses used almost completely abolishes vasodilator responses to K+5, but in the present experiments coinfusion of ouabain with Ba2+ does not completely inhibit the response to bradykinin, and responses to bradykinin are not abolished even when Ba2+ and ouabain are given with indomethacin and L-NMMA in doses that block forearm PG synthesis and nitric oxide-mediated responses to acetylcholine.23,24 The simplest explanation is that bradykinin dilates this vascular bed partly but not entirely through activation of KIR. The residual vasodilator response to bradykinin in the presence of inhibitors points to the possible involvement of a direct vasodilator action of bradykinin on vascular smooth muscle or of mediator pathways distinct from NO, prostaglandins, or K+ ions.
Activation of KIR by bradykinin could be via increased K+ concentration in the interstitial extracellular space in resistance arteries. The present experiments do not define the cellular origin of such increased interstitial K+ concentration. This could be the endothelium, as in rat hepatic artery in vitro,9,25 consistent with an EDHF/K+-mediated mechanism of bradykinin in the forearm in vivo. This agrees with the observation that a dose of tetraethylammonium expected to give a plasma concentration of 2x10–4 mol L–1 inhibits but does not abolish responses to bradykinin in this vascular bed.18 This concentration of tetraethylammonium could inhibit K+ efflux from endothelium via an action on KCa channels, where it produces half the maximum block at 2x10–4 mol L–1.7
Limitations
A constraint was our concern to limit the exposure of volunteers to Ba2+. The total infused dose of 24 μmol is less than one-tenth the chronic oral reference dose calculated by the Environmental Protection Agency. Limiting the dose of Ba2+ in this way meant that we were not able to explore its effects on a range of doses of bradykinin. A limitation of norepinephrine as a control is that it can inhibit KIR in vascular smooth muscle; other vasoconstrictors may influence KIR by an effect on membrane potential. Another limitation is that the present experiments do not identify the cellular distribution of the KIR channel involved in the vasodilator action of bradykinin in the forearm. This may be important because this channel is expressed not only in vascular smooth muscle but also in endothelial cells.26
In conclusion, a dose of Ba2+ that selectively inhibits KIR inhibits forearm vasodilator responses to bradykinin in healthy men, evidence that a component of this response is mediated by activation of KIR. This is consistent with bradykinin acting through K+ and/or another KIR-dependent EDHF in human forearm resistance vessels in vivo.
Acknowledgments
This work was supported by the British Heart Foundation.
Received June 8, 2004; accepted November 3, 2004.
References
Bradley KK, Jaggar JH, Bonev AD, Heppner TJ, Flynn ERM, Nelson MT, Horowitz B. Kir 2.1 encodes the inward rectifier potassium channel in rat arterial smooth muscle cells. J Physiol. 1999; 515: 639–651.
Zaritsky JJ, Eckman DM, Wellman GC, Nelson MT, Schwartz TL. Targeted disruption of Kir 2.1 and Kir 2.2 genes reveals the essential role of the inwardly rectifying K+ current in K+-mediated vasodilation. Circ Res. 2000; 87: 160–166.
Lopatin AN, Makhina EN, Nichols CG. Potassium channel block by cytoplasmic polyamines as the mechanism of intrinsic rectification. Nature. 1994; 372: 366–369.
Knot, HJ, Zimmerman PA, Nelson MT. Extracellular K+-induced hyperpolarizations and dilatations of rat coronary and cerebral arteries involve inward rectifier K+ channels. J Physiol (Lond). 1996; 492: 419–430.
Dawes M, Sieniawska C, Delves T et al. Barium reduces resting blood flow and inhibits potassium-induced vasodilation in the human forearm, Circulation. 2002; 105: 1323–1328.
Quayle JM, McCarron JG, Asbury JR, Nelson MT. Inward rectifier K+ currents in smooth muscle cells from rat resistance-sized cerebral arteries. Am J Physiol. 1993; 265: C1363–C1370.
Nelson MT, Quayle JM. Physiological roles and properties of potassium channels in arterial smooth muscle. Am J Physiol. 1995; 268: C799–C822.
Robinson BF, Phillips RJW, Wilson PN, Chiodini PL. Effect of local infusion of ouabain on human forearm vascular resistance and on response to potassium, verapamil and sodium nitroprusside. J Hypertens. 1983; 1: 165–169.
Edwards G, Dora KA, Gardener MJ, Garland CJ, Weston AH. K+ is an endothelium-derived hyperpolarizing factor in rat arteries, Nature. 1998; 396: 269–272.
Hong SL. Effect of bradykinin and thrombin on prostacyclin synthesis in endothelial cells from calf and pig aorta and human umbilical cord vein. Thromb Res. 1980; 18: 787–795.
Palmer RMJ, Ashton DS, Moncada S. Vascular endothelial cells synthesize nitric oxide from L-arginine. Nature. 1988; 333: 664–666.
Nakashima M, Mombouli J-V, Taylor AA, Vanhoutte PM. Endothelium-dependent hyperpolarization caused by bradykinin in human coronary arteries, J Clin Invest. 1993; 92: 2867–2871.
Fox RH, Goldsmith R, Kidd DJ. Lewis platelet glycoprotein (GP). Bradykinin as a vasodilator in man. J Physiol. 1961; 157: 589–602.
Cockcroft JR, Chowienczyk PJ, Brett SE, Bender N, Ritter JM. Inhibition of bradykinin-induced vasodilation in human forearm vasculature by icatibant, a potent B2-receptor antagonist. Br J Clin Pharmacol. 1994; 38: 317–322.
Benjamin N, Cockcroft JR, Collier JG, Dollery CT, Ritter JM, Webb DJ. Local inhibition of converting enzyme and vascular responses to angiotensin and bradykinin in the human forearm. J Physiol. 1989; 412: 543–555.
Cockcroft JR, Chowienczyk PJ, Brett SE, Ritter JM. Effect of NG monomethyl L arginine on kinin-induced vasodilation in the human forearm. Br J Clin Pharmacol. 1994; 38: 307–310.
O’Kane KP, Webb DJ, Collier JG, Vallance PJ. Local L-NG monomethyl-arginine attenuates the vasodilator action of bradykinin in the human forearm, Br J Clin Pharmacol. 1994; 38: 311–315.
Honing MLH, Smits P, Morrison PJ, Rabelink TJ. Bradykinin-induced vasodilation of human forearm resistance vessels is primarily mediated by endothelium-dependent hyperpolarisation. Hypertension. 2000; 35: 1314–1318.
Taddei S, Ghiadoni L, Virdis A, Buralli S, Salvetti A. Vasodilation to bradykinin is mediated by an ouabain-sensitive pathway as a compensatory mechanism for impaired nitric oxide availability in essential hypertensive patients. Circulation. 1999; 100: 1400–1405.
Vallance P, Collier J, Moncada S. Effects of endothelium-derived nitric oxide on peripheral arteriolar tone in man. Lancet. 1989; 2: 997–1000.
Dawes M, Chowienczyk PJ, Ritter JM. Effects of inhibition of the L-arginine/ nitric oxide pathway on vasodilation caused by ?-adrenergic agonists in human forearm. Circulation. 1997; 95: 2293–2297.
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