Hydrogen sulfide dilates cerebral arterioles by activating smooth muscle cell plasma membrane KATP channels

Hydrogen sulfide (H(2)S) is a gaseous signaling molecule that appears to contribute to the regulation of vascular tone and blood pressure. Multiple potential mechanisms of vascular regulation by H(2)S exist. Here, we tested the hypothesis that piglet cerebral arteriole smooth muscle cells generate ATP-sensitive K(+) (K(ATP)) currents and that H(2)S induces vasodilation by activating K(ATP) currents. Gas chromatography/mass spectrometry data demonstrated that after placing Na(2)S, an H(2)S donor, in solution, it rapidly (1 min) converts to H(2)S. Patch-clamp electrophysiology indicated that pinacidil (a K(ATP) channel activator), Na(2)S, and NaHS (another H(2)S donor) activated K(+) currents at physiological steady-state voltage (-50 mV) in isolated cerebral arteriole smooth muscle cells. Glibenclamide, a selective K(ATP) channel inhibitor, fully reversed pinacidil-induced K(+) currents and partially reversed (～58%) H(2)S-induced K(+) currents. Western blot analysis indicated that piglet arterioles expressed inwardly rectifying K(+) 6.1 (K(ir)6.1) channel and sulfonylurea receptor 2B (SUR2B) K(ATP) channel subunits. Pinacidil dilated pressurized (40 mmHg) piglet arterioles, and glibenclamide fully reversed this effect. Na(2)S also induced reversible and repeatable vasodilation with an EC(50) of ～30 μM, and this effect was partially reversed (～55%) by glibenclamide. Vasoregulation by H(2)S was also studied in pressurized resistance-size cerebral arteries of mice with a genetic deletion in the gene encoding SUR2 (SUR2 null). Pinacidil- and H(2)S-induced vasodilations were smaller in arterioles of SUR2 null mice than in wild-type controls. These data indicate that smooth muscle cell K(ATP) currents control newborn cerebral arteriole contractility and that H(2)S dilates cerebral arterioles by activating smooth muscle cell K(ATP) channels containing SUR2 subunits.     

Adenosine and hypoxic dilation of rat coronary small arteries: roles of the ATP-sensitive potassium channel, endothelium, and nitric oxide

The aims of the study were to examine the roles of the ATP-sensitive potassium (K(ATP)) channel, the endothelium, and nitric oxide (NO) in the responses of rat coronary small arteries to adenosine and hypoxia. Segments of rat coronary vessel were investigated in vitro using pressure myography; all vessels studied developed stable spontaneous myogenic tone during equilibration. Glibenclamide (a K(ATP) channel inhibitor) reversed pinacidil but not 2-deoxyglucose-induced dilation. Both adenosine and hypoxia dilated the vessels, and glibenclamide did not reverse these responses. Endothelial removal or N(G)-nitro-L-arginine methyl ester (L-NAME) inhibited the dilation to adenosine by approximately 50%; subsequent addition of glibenclamide was without effect. Hypoxic dilation was completely inhibited by endothelium removal or L-NAME. We conclude that adenosine- and hypoxia-induced dilation of rat coronary arteries does not appear to involve the K(ATP) channel. Adenosine-induced dilation is partially and hypoxic dilation is completely dependent on endothelium-derived NO.     

Arginine vasopressin inhibits Kir6.1/SUR2B channel and constricts the mesenteric artery via V1a receptor and protein kinase C

Kir6.1/SUR2B channel is the major isoform of K(ATP) channels in the vascular smooth muscle. Genetic disruption of either subunit leads to dysregulation of vascular tone and regional blood flows. To test the hypothesis that the Kir6.1/SUR2B channel is a target molecule of arginine vasopressin (AVP), we performed studies on the cloned Kir6.1/SUR2B channel and cell-endogenous K(ATP) channel in rat mesenteric arteries. The Kir6.1/SUR2B channel was expressed together with V1a receptor in the HEK-293 cell line. Whole cell currents of the transfected HEK cells were activated by K(ATP) channel opener pinacidil and inhibited by K(ATP) channel inhibitor glibenclamide. AVP produced a concentration-dependent inhibition of the pinacidil-activated currents with IC(50) 2.0 nM. The current inhibition was mediated by a suppression of the open-state probability without effect on single-channel conductance. An exposure to 100 nM PMA, a potent PKC activator, inhibited the pinacidil-activated currents, and abolished the channel inhibition by AVP. Such an effect was not seen with inactive phorbol ester. A pretreatment of the cells with selective PKC blocker significantly diminished the inhibitory effect of AVP. In acutely dissociated vascular smooth myocytes, AVP strongly inhibited the cell-endogenous K(ATP) channel. In isolated mesenteric artery rings, AVP produced concentration-dependent vasoconstrictions with EC(50) 6.5 nM. At the maximum effect, pinacidil completely relaxed vasoconstriction in the continuing exposure to AVP. The magnitude of the AVP-induced vasoconstriction was significantly reduced by calphostin-C. These results therefore indicate that the Kir6.1/SUR2B channel is a target molecule of AVP, and the channel inhibition involves G(q)-coupled V1a receptor and PKC.     

Pharmaco-topology of sulfonylurea receptors. Separate domains of the regulatory subunits of K(ATP) channel isoforms are required for selective interaction with K(+) channel openers

The differential responsiveness of (SUR1/K(IR)6.2)(4) pancreatic beta-cell versus (SUR2A/K(IR)6.2)(4) sarcolemmal or (SUR2B/K(IR)6. 0)(4) smooth muscle cell K(ATP) channels to K(+) channel openers (KCOs) is the basis for the selective prevention of hyperinsulinemia, myocardial infarction, and acute hypertension. KCO-stimulation of K(ATP) channels is a unique example of functional coupling between a transport ATPase and a K(+) inward rectifier. KCO binding to SUR is Mg-ATP-dependent and antagonizes the inhibition of (K(IR)6.0)(4) pore opening by nucleotides. Patch-clamping of matched chimeric human SUR1-SUR2A/K(IR)6.2 channels was used to identify the SUR regions that specify the selective response of sarcolemmal versus beta-cell channels to cromakalim or pinacidil versus diazoxide. The SUR2 segment containing the 12th through 17th predicted transmembrane domains, TMD12-17, confers sensitivity to the benzopyran, cromakalim, and the pyridine, pinacidil, whereas an SUR1 segment which includes TMD6-11 and the first nucleotide-binding fold, NBF1, controls responsiveness to the benzothiadiazine, diazoxide. These data are incorporated into a functional topology model for the regulatory SUR subunits of K(ATP) channels.     

Evidence for presence of ATP-sensitive K+ channels in rat colonic smooth muscle cells.

Coexpression of sulfonylurea receptor (SUR) and inward-rectifying K+ channel (Kir6.1 or 6.2) subunit yields ATP-sensitive K+ (K(ATP)) channels. Three subtypes of SUR have been cloned: pancreatic (SUR1), cardiac (SUR2A), and vascular smooth muscle (SUR2B). The distinct responses to K+ channel openers (KCOs) produced in different tissues may depend on the SUR isoform of K(ATP) channel. Therefore, we investigated the effects of pinacidil and diazoxide, two KCOs, on K(ATP) currents in intestinal smooth muscle cells of the rat colon (circular layer) using whole-cell voltage clamp. Pinacidil stimulated a time-independent K+ current evoked by various test potentials from a holding potential of -70 mV. The reversal potential of the stimulated current was about -75 mV, which is close to the equilibrium potential for K+ (E(K)). Both pinacidil and diazoxide dose-dependently stimulated K+ currents (evoked by ramp pulses), with EC50 values of 1.3 and 34.2 microM, respectively. The stimulated current was completely reversed by glybenclamide (3 microM). Since the EC50 values are close to those reported for vascular smooth muscle (VSM) cells, the SUR subtype may be similar to that in VSM cells, and could form the functional K(ATP) channel in rat colonic smooth muscle cells.     

Protein kinase C isoform-dependent modulation of ATP-sensitive K+ channels in mitochondrial inner membrane

The ATP-sensitive K(+) (K(ATP)) channels in both sarcolemmal (sarcK(ATP)) and mitochondrial inner membrane (mitoK(ATP)) are the critical mediators in cellular protection of ischemic preconditioning (IPC). Whereas cardiac sarcK(ATP) contains Kir6.2 and sulfonylurea receptor (SUR)2A, the molecular identity of mitoK(ATP) remains elusive. In the present study, we tested the hypothesis that protein kinase C (PKC) may promote import of Kir6.2-containing K(ATP) into mitochondria. Fluorescence imaging of isolated mitochondria from both rat adult cardiomyocytes and COS-7 cells expressing recombinant Kir6.2/SUR2A showed that Kir6.2-containing K(ATP) channels were localized in mitochondria and this mitochondrial localization was significantly increased by PKC activation with phorbol 12-myristate 13-acetate (PMA). Fluorescence resonance energy transfer microscopy further revealed that a significant number of Kir6.2-containing K(ATP) channels were localized in mitochondrial inner membrane after PKC activation. These results were supported by Western blotting showing that the Kir6.2 protein level in mitochondria from COS-7 cells transfected with Kir6.2/SUR2A was enhanced after PMA treatment and this increase was inhibited by the selective PKC inhibitor chelerythrine. Furthermore, functional analysis indicated that the number of functional K(ATP) channels in mitochondria was significantly increased by PMA, as shown by K(ATP)-dependent decrease in mitochondrial membrane potential in COS-7 cells transfected with Kir6.2/SUR2A but not empty vector. Importantly, PKC-mediated increase in mitochondrial Kir6.2-containing K(ATP) channels was blocked by a selective PKCepsilon inhibitor peptide in both COS-7 cells and cardiomyocytes. We conclude that the K(ATP) channel pore-forming subunit Kir6.2 is indeed localized in mitochondria and that the Kir6.2 content in mitochondria is increased by activation of PKCepsilon. PKC isoform-regulated mitochondrial import of K(ATP) channels may have significant implication in cardioprotection of IPC.     

Cerebral artery K(ATP)- and K(Ca)-channel activity and contractility: changes with development

The present study was designed to test the hypothesis that in cerebral arteries of the fetus, ATP-sensitive (K(ATP)) and Ca(2+)-activated K(+) channels (K(Ca)) play an important role in the regulation of intracellular Ca(2+) concentration ([Ca(2+)](i)) and that this differs significantly from that of the adult. In main branch middle cerebral arteries (MCA) from near-term fetal ( approximately 140 days) and nonpregnant adult sheep, simultaneously we measured norepinephrine (NE)-induced responses of vascular tension and [Ca(2+)](i) in the absence and presence of selective K(+)-channel openers/blockers. In fetal MCA, in a dose-dependent manner, both the K(ATP)-channel opener pinacidil and the K(Ca)-channel opener NS 1619 significantly inhibited NE-induced tension [negative logarithm of the half-maximal inhibitory concentration (pIC(50)) = 5.0 +/- 0.1 and 8.2 +/- 0.1, respectively], with a modest decrease of [Ca(2+)](i). In the adult MCA, in contrast, both pinacidil and NS 1619 produced a significant tension decrease (pIC(50) = 5.1 +/- 0.1 and 7.6 +/- 0.1, respectively) with no change in [Ca(2+)](i). In addition, the K(Ca)-channel blocker iberiotoxin (10(-7) to 10(-6) M) resulted in increased tension and [Ca(2+)](i) in both adult and fetal MCA, although the K(ATP)-channel blocker glibenclamide (10(-7) to 3 x 10(-5) M) failed to do so. Of interest, administration of 10(-7) M iberiotoxin totally eliminated vascular contraction and increase in [Ca(2+)](i) seen in response to 10(-5) M ryanodine. In precontracted fetal cerebral arteries, activation of the K(ATP) and K(Ca) channels significantly decreased both tension and [Ca(2+)](i), suggesting that both K(+) channels play an important role in regulating L-type channel Ca(2+) flux and therefore vascular tone in these vessels. In the adult, K(ATP) and the K(Ca) channels also appear to play an important role in this regard; however, in the adult vessel, activation of these channels with resultant vasorelaxation can occur with no significant change in [Ca(2+)](i). These channels show differing responses to inhibition, e.g., K(Ca)-channel inhibition, resulting in increased tension and [Ca(2+)](i), whereas K(ATP)-channel inhibition showed no such effect. In addition, the K(Ca) channel appears to be coupled to the sarcoplasmic reticulum ryanodine receptor. Thus differences in plasma membrane K(+)-channel activity may account, in part, for the differences in the regulation of contractility of fetal and adult cerebral arteries.     

Modulation of the trafficking efficiency and functional properties of ATP-sensitive potassium channels through a single amino acid in the sulfonylurea receptor

Mutations in the sulfonylurea receptor 1 (SUR1), a subunit of ATP-sensitive potassium (K(ATP)) channels, cause familial hyperinsulinism. One such mutation, deletion of phenylalanine 1388 (DeltaPhe-1388), leads to defects in both trafficking and MgADP response of K(ATP) channels. Here we investigated the biochemical features of Phe-1388 that control the proper trafficking and function of K(ATP) channels by substituting the residue with all other 19 amino acids. Whereas surface expression is largely dependent on hydrophobicity, channel response to MgADP is governed by multiple factors and involves the detailed architecture of the amino acid side chain. Thus, structural features in SUR1 required for proper channel function are distinct from those required for correct protein trafficking. Remarkably, replacing Phe-1388 by leucine profoundly alters the physiological and pharmacological properties of the channel. The F1388L-SUR1 channel has increased sensitivity to MgADP and metabolic inhibition, decreased sensitivity to glibenclamide, and responds to both diazoxide and pinacidil. Because this conservative amino acid substitution occurs in the SUR2A and SUR2B isoforms, the mutation provides a mechanism by which functional diversities in K(ATP) channels are generated.     

Fetal and adult cerebral artery K(ATP) and K(Ca) channel responses to long-term hypoxia.

High-altitude long-term hypoxia (LTH) alters cerebral vascular contractile and relaxation responses in both fetus and adult. We tested the hypotheses that LTH-mediated vascular responses were secondary to altered K+ channel function and that in the fetus these responses differ from those of the adult. In middle cerebral arteries (MCA) from both nonpregnant adult and fetal (approximately 140 days gestation) sheep, which were either acclimatized to high altitude (3,820 m) or sea-level controls, we measured norepinephrine (NE)-induced contractions and intracellular Ca2+ concentration ([Ca2+]i) simultaneously, in the presence or absence of different K+ channel openers or blockers. In adult MCA, LTH was associated with approximately 20% decrease in NE-induced tension and [Ca2+]i, with a significant increase in Ca2+ sensitivity. In contrast, in fetal MCA, LTH failed to affect significantly NE-induced contraction or [Ca2+]i but significantly decreased the ATP-sensitive K+ (K(ATP)) channel and Ca2+-activated K+ (K(Ca)) channel-mediated relaxation. The significant effect of K(ATP) and K(Ca) channel activators on the relaxation responses and the fact that K+ channels play a key role in myogenic tone support the hypotheses that K+ channels play an important role in hypoxia-mediated responses. These results also support the hypothesis of significant developmental differences with maturation from fetus to adult.     

Endosomal KATP channels as a reservoir after myocardial ischemia: a role for SUR2 subunits

ATP-sensitive K(+) (K(ATP)) channels, composed of inward rectifier K(+) (Kir)6.x and sulfonylurea receptor (SUR)x subunits, are expressed on cellular plasma membranes. We demonstrate an essential role for SUR2 subunits in trafficking K(ATP) channels to an intracellular vesicular compartment. Transfection of Kir6.x/SUR2 subunits into a variety of cell lines (including h9c2 cardiac cells and human coronary artery smooth muscle cells) resulted in trafficking to endosomal/lysosomal compartments, as assessed by immunofluorescence microscopy. By contrast, SUR1/Kir6.x channels efficiently localized to the plasmalemma. The channel turnover rate was similar with SUR1 or SUR2, suggesting that the expression of Kir6/SUR2 proteins in lysosomes is not associated with increased degradation. Surface labeling of hemagglutinin-tagged channels demonstrated that SUR2-containing channels dynamically cycle between endosomal and plasmalemmal compartments. In addition, Kir6.2 and SUR2 subunits were found in both endosomal and sarcolemmal membrane fractions isolated from rat hearts. The balance of these K(ATP) channel subunits shifted to the sarcolemmal membrane fraction after the induction of ischemia. The K(ATP) channel current density was also increased in rat ventricular myocytes isolated from hearts rendered ischemic before cell isolation without corresponding changes in subunit mRNA expression. We conclude that an intracellular pool of SUR2-containing K(ATP) channels exists that is derived by endocytosis from the plasma membrane. In cardiac myocytes, this pool can potentially play a cardioprotective role by serving as a reservoir for modulating surface K(ATP) channel density under stress conditions, such as myocardial ischemia.     

PKA phosphorylation of SUR2B subunit underscores vascular KATP channel activation by beta-adrenergic receptors

ATP-sensitive K(+) (K(ATP)) channels are activated by several vasodilating hormones and neurotransmitters through the PKA pathway. Here, we show that phosphorylation at Ser1387 of the SUR2B subunit is critical for the channel activation. Experiments were performed in human embryonic kidney (HEK) 293 cells expressing the cloned Kir6.1/SUR2B channel. In whole cell patch, the Kir6.1/SUR2B channel activity was stimulated by isoproterenol via activation of beta(2) receptors. This effect was blocked in the presence of inhibitors for adenylyl cyclase or PKA. Similar channel activation was seen by exposing inside-out patches to the catalytic subunit of PKA. Because none of the previously suggested PKA phosphorylation sites accounted for the channel activation, we performed systematic mutational analysis on Kir6.1 and SUR2B. Two serine residues (Ser1351, Ser1387) located in the NBD2 of SUR2B were critical for the channel activation. In vitro phosphorylation experiments showed that Ser1387 but not Ser1351 was phosphorylated by PKA. The PKA-dependent activation of cell-endogenous K(ATP) channels was observed in acutely dissociated mesenteric smooth myocytes and isolated mesenteric artery rings, where activation of these channels contributed significantly to the isoproterenol-induced vasodilation. Taken together, these results indicate that the Kir6.1/SUR2B channel is a target of beta(2) receptors and that the channel activation relies on PKA phosphorylation of SUR2B at Ser1387.     

Regulation of ATP-sensitive potassium channel function by protein kinase A-mediated phosphorylation in transfected HEK293 cells

ATP-sensitive potassium (K(ATP)) channels regulate insulin secretion, vascular tone, heart rate and neuronal excitability by responding to transmitters as well as the internal metabolic state. K(ATP) channels are composed of four pore-forming alpha-subunits (Kir6.2) and four regulatory beta-subunits, the sulfonylurea receptor (SUR1, SUR2A or SUR2B). Whereas protein kinase A (PKA) phosphorylation of serine 372 of Kir6.2 has been shown biochemically by others, we found that the phosphorylation of T224 rather than S372 of Kir6.2 underlies the catalytic subunits of PKA (c-PKA)- and the D1 dopamine receptor-mediated stimulation of K(ATP) channels expressed in HEK293 cells. Specific changes in the kinetic properties of channels treated with c-PKA, as revealed by single-channel analysis, were mimicked by aspartate substitution of T224. The T224D mutation also reduced the sensitivity to ATP inhibition. Alteration of channel gating and a decrease in the apparent affinity for ATP inhibition thus underlie the positive regulation of K(ATP) channels by PKA phosphorylation of T224 in Kir6.2, which may represent a general mechanism for K(ATP) channel regulation in different tissues.     

Delivery of genes encoding cardiac K(ATP) channel subunits in conjunction with pinacidil prevents membrane depolarization in cells exposed to chemical hypoxia-reoxygenation

Metabolic injury is a complex process affecting various: tissues with membrane depolarisation recognised as a common trigger event leading to cell death. To examine whether, under metabolic challenge, membrane potential homeostasis can be maintained by an activator of channel proteins, we here delivered Kir6.2 and SUR2A genes, which encode cardiac K(ATP) channel subunits, into a somatic cell line lacking native K(ATP) channels (COS-7 cells). Chemical hypoxia-reoxygenation was simulated in COS-7 cells by addition and removal of the mitochondrial poison 2,4 dinitrophenol (DNP). The membrane potential of COS-7 cells at rest was -31 +/- 3 mV. This value did not change following 3 min-long exposure to DNP (-32 +/- 4 mV). In contrast, washout of DNP induced significant membrane depolarisation (-17 +/- 2 mV). Delivery of Kir6.2/SUR2A genes did not change cellular response to hypoxia-reoxygenation. Similarly, pinacidil, potassium channel opener, did not have effect on hypoxia-reoxygenation-induced membrane depolarisation in cells lacking recombinant K(ATP) channel subunits. However, gene delivery combined with pinacidil prevented membrane depolarisation induced by hypoxia-reoxygenation. This effect of pinacidil, in cells expressing Kir6.2/SUR2A, was observed regardless of whether pinacidil was added only during hypoxia or reoxygenation. The present study demonstrates that combined use of K(ATP) channel subunits gene delivery and pharmacological targeting of recombinant proteins can be used to efficiently control membrane potential under hypoxia-reoxygenation.     

Different binding properties and affinities for ATP and ADP among sulfonylurea receptor subtypes, SUR1, SUR2A, and SUR2B

ATP-sensitive potassium (K(ATP)) channels, composed of sulfonylurea receptor (SURx) and Kir6.x, play important roles by linking cellular metabolic state to membrane potential in various tissues. Pancreatic, cardiac, and vascular smooth muscle K(ATP) channels, which consist of different subtypes of SURx, differ in their responses to cellular metabolic state. To explore the possibility that different interactions of SURx with nucleotides cause differential regulation of K(ATP) channels, we analyzed the properties of nucleotide-binding folds (NBFs) of SUR1, SUR2A, and SUR2B. SURx in crude membrane fractions was incubated with 8-azido-[alpha-(32)P]ATP or 8-azido-[gamma-(32)P]ATP under various conditions and was photoaffinity-labeled. Then, SURx was digested mildly with trypsin, and partial tryptic fragments were immunoprecipitated with antibodies against NBF1 and NBF2. Some nucleotide-binding properties were different among SUR subtypes as follows. 1) Mg(2+) dependence of nucleotide binding of NBF2 of SUR1 was high, whereas those of SUR2A and SUR2B were low. 2) The affinities of NBF1 of SUR1 for ATP and ADP, especially for ATP, were significantly higher than those of SUR2A and SUR2B. 3) The affinities of NBF2 of SUR2B for ATP and ADP were significantly higher than those of SUR2A. This is the first biochemical study to analyze and compare the nucleotide-binding properties of NBFs of three SUR subtypes, and our results suggest that their different properties may explain, in part, the differential regulation of K(ATP) channel subtypes. The high nucleotide-binding affinities of SUR1 may explain the high ability of SUR1 to stimulate pancreatic K(ATP) channels. It is also suggested that the C-terminal 42 amino acids affect the physiological roles of SUR2A and SUR2B by changing the nucleotide-binding properties of their NBFs.     

Regulation of the ATP-sensitive potassium channel subunit, Kir6.2, by a Ca2+-dependent protein kinase C

The activity of ATP-sensitive potassium (K(ATP)) channels is governed by the concentration of intracellular ATP and ADP and is thus responsive to the metabolic status of the cell. Phosphorylation of K(ATP) channels by protein kinase A (PKA) or protein kinase C (PKC) results in the modulation of channel activity and is particularly important in regulating smooth muscle tone. At the molecular level the smooth muscle channel is composed of a sulfonylurea subunit (SUR2B) and a pore-forming subunit Kir6.1 and/or Kir6.2. Previously, Kir6.1/SUR2B channels have been shown to be inhibited by PKC, and Kir6.2/SUR2B channels have been shown to be activated or have no response to PKC. In this study we have examined the modulation of channel complexes formed of the inward rectifier subunit, Kir6.2, and the sulfonylurea subunit, SUR2B. Using a combination of biochemical and electrophysiological techniques we show that this complex can be inhibited by protein kinase C in a Ca(2+)-dependent manner and that this inhibition is likely to be as a result of internalization. We identify a residue in the distal C terminus of Kir6.2 (Ser-372) whose phosphorylation leads to down-regulation of the channel complex. This inhibitory effect is distinct from activation which is seen with low levels of channel activity.     

Alteration of ATP-sensitive K+ channels in rabbit aortic smooth muscle during left ventricular hypertrophy

We investigated the impairment of ATP-sensitive K(+) (K(ATP)) channels in aortic smooth muscle cells (ASMCs) from isoproterenol-induced hypertrophied rabbits. The amplitude of K(ATP) channels induced by the K(ATP) channel opener pinacidil (10 μM) was greater in ASMCs from control than from hypertrophied animals. In phenylephrine-preconstricted aortic rings, pinacidil induced relaxation in a dose-dependent manner. The dose-dependent curve was shifted to the right in the hypertrophied (EC(50): 17.80 ± 3.28 μM) compared with the control model (EC(50): 6.69 ± 2.40 μM). Although the level of Kir6.2 subtype expression did not differ between ASMCs from the control and hypertrophied models, those of the Kir6.1 and SUR2B subtypes were decreased in the hypertrophied model. Application of the calcitonin-gene related peptide (100 nM) and adenylyl cyclase activator forskolin (10 μM), which activates protein kinase A (PKA) and consequently K(ATP) channels, induced a K(ATP) current in both control and hypertrophied animals; however, the K(ATP) current amplitude did not differ between the two groups. Furthermore, PKA expression was not altered between the control and hypertrophied animals. These results suggests that the decreased K(ATP) current amplitude and K(ATP) channel-induced vasorelaxation in the hypertrophied animals were attributable to the reduction in K(ATP) channel expression but not to changes in the intracellular signaling mechanism that activates the K(ATP) current.     

Pharmacological modulation of K(ATP) channels

Pharmacological modulation of ATP-sensitive K+ (K(ATP)) channels is used in the treatment of a number of clinical conditions, including type 2 diabetes and angina. The sulphonylureas and related drugs, which are used to treat type 2 diabetes, stimulate insulin secretion by closing K(ATP) channels in pancreatic beta-cells. Agents used to treat angina, by contrast, act by opening K(ATP) channels in vascular smooth and cardiac muscle. Both the therapeutic K(ATP) channel inhibitors and the K(ATP) channel openers target the sulphonylurea receptor (SUR) subunit of the K(ATP) channel, which exists in several isoforms expressed in different tissues (SUR1 in pancreatic beta-cells, SUR2A in cardiac muscle and SUR2B in vascular smooth muscle). The tissue-specific action of drugs that target the K(ATP) channel is attributed to the properties of these different SUR subtypes. In this review, we discuss the molecular basis of tissue-specific drug action, and its implications for clinical practice.