Ageing is associated with a decrease in the number of sarcolemmal ATP-sensitive K+ channels in a gender-dependent manner

The opening of sarcolemmal K(ATP) channels is considered to be an important endogenous cardioprotective mechanism. On the other hand, age-dependent changes in the myocardial susceptibility to ischemia and hypoxia have been observed in different species, including humans. Here, we have hypothesized that aging might be associated with the changes in sarcolemmal K(ATP) channels. Therefore, the main objective of the present study was to establish whether aging changes expression of cardiac sarcolemmal ATP-sensitive K+ (K(ATP)) channels. RT-PCR using primers specific for K(ATP) channel subunits, Kir6.2, Kir6.1 and SUR2A subunits was performed using total RNA from guinea-pig ventricular tissue. Whole cell electrophysiology was done on isolated guinea-pig ventricular cardiomyocytes. Western blotting using anti-Kir6.2 and anti-SUR2A antibodies was performed on cardiac membrane fraction. Tissue and cells were harvested from young and old, male and female guinea-pigs. RT-PCR analysis did not reveal significant age-related changes in levels of Kir6.1 or Kir6.2 mRNAs. However, levels of SUR2A were significantly lower in old than in young females. Such age-differences were not observed with cardiac tissue from male animals. In both old and young males, pinacidil (100 microM) induced outward currents. The difference between current density of pinacidil-sensitive component in females, but not males, was statistically significant. Western blotting analysis revealed higher levels of Kir6.2 and SUR2A proteins in cardiac membrane fraction from young than old females. The present study demonstrates that in females, but not males, aging is associated with decrease in number of cardiac K(ATP) channels which is due to decrease in levels of the SUR2A subunit.     

Functional modulation of the ATP-sensitive potassium channel by extracellular signal-regulated kinase-mediated phosphorylation

ATP-sensitive potassium (K(ATP)) channels play an important role in controlling insulin secretion and vascular tone as well as protecting neurons under metabolic stress. We have previously demonstrated that stimulation of the K(ATP) channel by nitric oxide (NO) requires activation of Ras- and extracellular signal-regulated kinase (ERK) of the mitogen-activated protein kinase (MAPK) family. However, the mechanistic link between ERK and the K(atp) channel remained unknown. To investigate how ERK modulates the function of K(ATP) channels, we performed single-channel recordings in combination with site-directed mutagenesis. The Kir6.2/SUR1 channel, a neuronal K(ATP) channel isoform, was expressed in human embryonic kidney (HEK) 293 cells by transient transfection. Direct application of the activated ERK2 to the cytoplasmic surface of excised, inside-out patches markedly enhanced the single-channel activity of Kir6.2/SUR1 channels. The normalized open probability (NPo) and opening frequency were significantly increased, whereas the mean closed duration was reduced. The single-channel conductance level was not affected. The ERK2-induced stimulation of Kir6.2/SUR1 channels was prevented by heat-inactivation of the enzyme. Furthermore, alanine substitutions of T341 and S385 to disrupt the potential ERK phosphorylation sites present in the Kir6.2 subunit significantly abrogated the stimulatory effects of ERK2, while aspartate substitutions of T341 and S385 to mimic the (negative) charge effect of phosphorylation rendered a small yet significant reduction in the ATP sensitivity of the channel. Taken together, here we report for the first time that ERK2/MAPK activates neuronal-type K(ATP) channels, and this stimulation requires ERK phosphorylation of the Kir6.2 subunit at T341 and S385 residues. The ERK2-induced K(ATP) channel stimulation can be accounted for by changes in channel gating that destabilize the closed states and by reduction in the ATP sensitivity. As Kir6.2 is the pore-forming subunit of K(ATP) channels, ERK2-mediated phosphorylation may represent a common mechanism for K(ATP) channel regulation in different tissues.     

Dual regulation of the ATP-sensitive potassium channel by activation of cGMP-dependent protein kinase

Adenosine triphosphate (ATP)-sensitive potassium (K(ATP)) channels couple cellular metabolic status to membrane electrical activity. In this study, we performed patch-clamp recordings to investigate how cyclic guanosine monophosphate (cGMP)-dependent protein kinase (PKG) regulates the function of K(ATP) channels, using both transfected human SH-SY5Y neuroblastoma cells and embryonic kidney (HEK) 293 cells. In intact SH-SY5Y cells, the single-channel currents of Kir6.2/sulfonylurea receptor (SUR) 1 channels, a neuronal-type K(ATP) isoform, were enhanced by zaprinast, a cGMP-specific phosphodiesterase inhibitor; this enhancement was abolished by inhibition of PKG, suggesting a stimulatory role of cGMP/PKG signaling in regulating the function of neuronal K(ATP) channels. Similar effects of cGMP accumulation were confirmed in intact HEK293 cells expressing Kir6.2/SUR1 channels. In contrast, direct application of purified PKG suppressed rather than activated Kir6.2/SUR1 channels in excised, inside-out patches, while tetrameric Kir6.2LRKR368/369/370/371AAAA channels expressed without the SUR subunit were not modulated by zaprinast or purified PKG. Lastly, reconstitution of the soluble guanylyl cyclase/cGMP/PKG signaling pathway by generation of nitric oxide led to Kir6.2/SUR1 channel activation in both cell types. Taken together, here, we report novel findings that PKG exerts dual functional regulation of neuronal K(ATP) channels in a SUR subunit-dependent manner, which may provide new means of therapeutic intervention for manipulating neuronal excitability and/or survival.     

Functional effects of naturally occurring KCNJ11 mutations causing neonatal diabetes on cloned cardiac KATP channels

ATP-sensitive K⁺ (K_ATP) channels are hetero-octamers of inwardly rectifying K⁺ channel (Kir6.2) and sulphonylurea receptor subunits (SUR1 in pancreatic β-cells, SUR2A in heart). Heterozygous gain-of-function mutations in Kir6.2 cause neonatal diabetes, which may be accompanied by epilepsy and developmental delay. However, despite the importance of K_ATP channels in the heart, patients have no obvious cardiac problems. We examined the effects of adenine nucleotides on K_ATP channels containing wild-type or mutant (Q52R, R201H) Kir6.2 plus either SUR1 or SUR2A. In the absence of Mg²⁺, both mutations reduced ATP inhibition of SUR1- and SUR2A-containing channels to similar extents, but when Mg²⁺ was present ATP blocked mutant channels containing SUR1 much less than SUR2A channels. Mg-nucleotide activation of SUR1, but not SUR2A, channels was markedly increased by the R201H mutation. Both mutations also increased resting whole-cell K_ATP currents through heterozygous SUR1-containing channels to a greater extent than for heterozygous SUR2A-containing channels. The greater ATP inhibition of mutant Kir6.2/SUR2A than of Kir6.2/SUR1 can explain why gain-of-function Kir6.2 mutations manifest effects in brain and β-cells but not in the heart.     

Activation of the Na+/K+-ATPase by insulin and glucose as a putative negative feedback mechanism in pancreatic beta-cells

Pancreatic beta-cells of sulfonylurea receptor type 1 knock-out (SUR1(-/-)) mice exhibit an oscillating membrane potential (V (m)) demonstrating that hyper-polarisation occurs despite the lack of K(ATP) channels. We hypothesize that glucose activates the Na(+)/K(+)-ATPase thus increasing a hyper-polarising current. Elevating glucose in SUR1(-/-) beta-cells resulted in a transient fall in V (m) and [Ca(2+)](c) independent of sarcoplasmic and endoplasmic reticulum Ca(2+)-activated ATPase (SERCA) activation. This was not affected by K(+) channel blockade but inhibited by ATP depletion and by ouabain. Increasing glucose also reduced [Na(+)](c), an effect reversed by ouabain. Exogenously applied insulin decreased [Na(+)](c) and hyper-polarised V (m). Inhibiting insulin signalling in SUR1(-/-) beta-cells blunted the glucose-induced decrease of [Ca(2+)](c). Tolbutamide (1 mmol/l) disclosed the SERCA-independent effect of glucose on [Ca(2+)](c) in wild-type beta-cells. The data show that in SUR1(-/-) beta-cells, glucose activates the Na(+)/K(+)-ATPase presumably by increasing [ATP](c). Insulin can also stimulate the pump and potentiate the effect of glucose. Pathways involving the pump may thus serve as potential drug targets in certain metabolic disorders.     

Functional effects of KCNJ11 mutations causing neonatal diabetes: enhanced activation by MgATP

Recent studies have shown that heterozygous mutations in KCNJ11, which encodes Kir6.2, the pore-forming subunit of the ATP-sensitive potassium (K(ATP)) channel, cause permanent neonatal diabetes either alone (R201C, R201H) or in association with developmental delay, muscle weakness and epilepsy (V59G,V59M). Functional analysis in the absence of Mg2+, to isolate the inhibitory effects of ATP on Kir6.2, showed that both types of mutation reduce channel inhibition by ATP. However, in pancreatic beta-cells, K(ATP) channel activity is governed by the balance between ATP inhibition via Kir6.2 and Mg-nucleotide stimulation mediated by an auxiliary subunit, the sulphonylurea receptor SUR1. We therefore studied the MgATP sensitivity of KCNJ11 mutant K(ATP) channels expressed in Xenopus oocytes. In contrast to wild-type channels, Mg2+ dramatically reduced the ATP sensitivity of heterozygous R201C, R201H, V59M and V59G channels. This effect was predominantly mediated via the nucleotide-binding domains of SUR1 and resulted from an enhanced stimulatory action of MgATP. Our results therefore demonstrate that KCNJ11 mutations increase the current magnitude of heterozygous K(ATP) channels in two ways: by increasing MgATP activation and by decreasing ATP inhibition. They further show that the fraction of unblocked K(ATP) current at physiological MgATP concentrations correlates with the severity of the clinical phenotype.     

The Kir6.2-F333I mutation differentially modulates KATP channels composed of SUR1 or SUR2 subunits

Mutations in Kir6.2, the pore-forming subunit of the K_ATP channel, that reduce the ability of ATP to block the channel cause neonatal diabetes. The stimulatory effect of MgATP mediated by the regulatory sulphonylurea receptor (SUR) subunit of the channel may also be modified. We compared the effect of the Kir6.2-F333I mutation on K_ATP channels containing SUR1, SUR2A or SUR2B. The open probability of Kir6.2/SUR1 channels, or a C-terminally truncated form of Kir6.2 expressed in the absence of SUR, was unaffected by the mutation. However, that of Kir6.2/SUR2A and Kir6.2/SUR2B channels was increased. In the absence of Mg²⁺, ATP inhibition of all Kir6.2-F333I/SUR channel types was reduced, although SUR1-containing channels were reduced more than SUR2-containing channels. These results suggest F333 is involved in differential coupling of Kir6.2 to SUR1 and SUR2. When Mg²⁺ was present, ATP blocked SUR2A channels but activated SUR2B and SUR1 channels. Activation by MgGDP (or MgADP) was similar for wild-type and mutant channels and was independent of SUR. This indicates Mg-nucleotide binding to SUR and the transduction of binding into opening of the Kir6.2 pore are unaffected by the mutation. The data further suggest that MgATP hydrolysis by the nucleotide-binding domains of SUR1 and SUR2B, but not SUR2A, is enhanced by the F333I mutation in Kir6.2. Taken together, our data suggest the region of the C terminus within which F333 lies is involved in more than one type of functional interaction with SUR, and that F333 interacts differentially with SUR1 and SUR2.     

心肌肌细胞膜及线粒体膜ATP敏感钾通道与心肌缺血预处理

The potential cardioprotection can be involved during ischemic preconditioning in heart which mechanisms remain unknown yet. It is reported that sarcolemmal and mitochondrial KA‘rp channels mediate cardioprotection, and they play distinct myoprotective roles during ischemic preconditioning. This paper reviews the present progress in myocardial sarcolemmal and mitochondrial KATP channels in ischemic preconditioning.     

A mutation in the ATP-binding site of the Kir6.2 subunit of the KATP channel alters coupling with the SUR2A subunit

Mutations in the pore-forming subunit of the ATP-sensitive K⁺ (K_ATP) channel Kir6.2 cause neonatal diabetes. Understanding the molecular mechanism of action of these mutations has provided valuable insight into the relationship between the structure and function of the K_ATP channel. When Kir6.2 containing a mutation (F333I) in the putative ATP-binding site is coexpressed with the cardiac type of regulatory K_ATP channel subunit, SUR2A, the channel sensitivity to ATP inhibition is reduced and the intrinsic open probability ( P _o ) is increased. However, the extent of macroscopic current activation by MgADP was unaffected. Here we examine rundown and MgADP activation of wild-type and Kir6.2-F333I/SUR2A channels using single-channel recording, noise analysis and spectral analysis. We also compare the effect of mutating the adjacent residue, G334, on rundown and MgADP activation. All three approaches indicated that rundown of Kir6.2-F333I/SUR2A channels is due to a reduction in the number of active channels in the patch and that MgADP reactivation involves recruitment of inactive channels. In contrast, rundown and MgADP reactivation of wild-type and Kir6.2-G334D/SUR2A channels, and of Kir6.2-F333I/SUR1 channels, involve a gradual change in P _o . Our results suggest that F333 in Kir6.2 interacts functionally with SUR2A to modulate channel rundown and MgADP activation. This interaction is fairly specific as it is not disturbed when the adjacent residue (G334) is mutated. It is also not a consequence of the enhanced P _o of Kir6.2-F333I/SUR2A channels, as it is not found for other mutant channels with high P _o (Kir6.2-I296L/SUR2A).     

Allosteric modulation of the mouse kir6.2 channel by intracellular H+ and ATP

The ATP-sensitive K⁺ (K_ATP) channels are regulated by intracellular H⁺ in addition to ATP, ADP, and phospholipids. Here we show evidence for the interaction of H⁺ with ATP in regulating a cloned K_ATP channel, i.e. Kir6.2 expressed with and without the SUR1 subunit. Channel sensitivity to ATP decreases at acidic pH, while the pH sensitivity also drops in the presence of ATP. These effects are more evident in the presence of the SUR1 subunit. In the Kir6.2 + SUR1, the pH sensitivity is reduced by about 0.4 pH units with 100 μM ATP and 0.6 pH units with 1 m m ATP, while a decrease in pH from 7.4 to 6.8 lowers the ATP sensitivity by about fourfold. The Kir6.2 + SUR1 currents are strongly activated at pH 5.9-6.5 even in the presence of 1 m m ATP. The modulations appear to take place at His175 and Lys185 that are involved in proton and ATP sensing, respectively. Mutation of His175 completely eliminates the pH effect on the ATP sensitivity. Similarly, the K185E mutant-channel loses the ATP-dependent modulation of the pH sensitivity. Thus, allosteric modulations of the cloned K_ATP channel by ATP and H⁺ are demonstrated. Such a regulation allows protons to activate directly the K_ATP channels and release channel inhibition by intracellular ATP; the pH effect is further enhanced with a decrease in ATP concentration as seen in several pathophysiological conditions.     

Pinacidil prevents membrane depolarisation and intracellular Ca2+ loading in single cardiomyocytes exposed to severe metabolic stress

Recently, it has been proposed that, besides sarcolemmal K(ATP) channels, the activation of mitochondrial K(ATP) channels may also contribute to the cardioprotective action of potassium channel openers. In this respect, use of drugs that target both mitochondrial and sarcolemmal K(ATP) channels, such as pinacidil, may be a promising therapeutic strategy against metabolic injury of the heart. Therefore, the main objective of the present study was to determine whether pinacidil could maintain the value of resting membrane potential and intracellular Ca2+ homeostasis in cardiac cells exposed to metabolic stress. Experiments were performed on isolated ventricular cardiomyocytes. The membrane potential was monitored during experiments using whole cell patch clamp electrophysiology and the intracellular Ca2+ concentration was measured by a digital epifluorescence imaging. Chemical hypoxia-reoxygenation was induced by application and removal of the mitochondrial poison 2,4 dinitrophenol (DNP). Under hypoxia-reoxygenation, membrane depolarisation and intracellular Ca2+ loading was induced by Ca2+ influx during hypoxia and release of Ca2+ from intracellular stores during reoxygenation. The K(ATP) channel activator, pinacidil, prevented intracellular Ca2+ loading and membrane depolarisation, irrespective of whether the channel opener was applied throughout the duration of hypoxia-reoxygenation or transiently during the hypoxic or reoxygenation stage. Thus, the present study provides evidence that pinacidil, a non-selective K(ATP) channel opener, can handle membrane potential and intracellular Ca2+ homeostasis in cardiomyocytes under hypoxia-reoxygenation irrespective of the stage of the metabolic insult. This provides further evidence, at the single cell level, that targeting K(ATP) channels may be a valuable approach to protect the myocardium against metabolic challenge.     

Mechanism of action of a sulphonylurea receptor SUR1 mutation (F132L) that causes DEND syndrome

Activating mutations in the genes encoding the ATP-sensitive potassium (K(ATP)) channel subunits Kir6.2 and SUR1 are a common cause of neonatal diabetes. Here, we analyse the molecular mechanism of action of the heterozygous mutation F132L, which lies in the first set of transmembrane helices (TMD0) of SUR1. This mutation causes severe developmental delay, epilepsy and permanent neonatal diabetes (DEND syndrome). We show that the F132L mutation reduces the ATP sensitivity of K(ATP) channels indirectly, by altering the intrinsic gating of the channel. Thus, the open probability is markedly increased when Kir6.2 is co-expressed with mutant TMD0 alone or with mutant SUR1. The F132L mutation disrupts the physical interaction between Kir6.2 and TMD0, but does not alter the plasmalemma channel density. Our results explain how a mutation in an accessory subunit can produce enhanced activity of the K(ATP) channel pore (formed by Kir6.2). They also provide further evidence that interactions between TMD0 of SUR1 and Kir6.2 are critical for K(ATP) channel gating and identify a residue crucial for this interaction at both physical and functional levels.     

A heterozygous activating mutation in the sulphonylurea receptor SUR1 (ABCC8) causes neonatal diabetes

Neonatal diabetes is a genetically heterogeneous disorder with nine different genetic aetiologies reported to date. Heterozygous activating mutations in the KCNJ11 gene encoding Kir6.2, the pore-forming subunit of the ATP-sensitive potassium (K(ATP)) channel, are the most common cause of permanent neonatal diabetes. The sulphonylurea receptor (SUR) SUR1 serves as the regulatory subunit of the K(ATP) channel in pancreatic beta cells. We therefore hypothesized that activating mutations in the ABCC8 gene, which encodes SUR1, might cause neonatal diabetes. We identified a novel heterozygous mutation, F132L, in the ABCC8 gene of a patient with severe developmental delay, epilepsy and neonatal diabetes (DEND syndrome). This mutation had arisen de novo and was not present in 150 control chromosomes. Residue F132 shows evolutionary conservation across species and is located in the first set of transmembrane helices (TMD0) of SUR1, which is proposed to interact with Kir6.2. Functional studies of recombinant K(ATP) channels demonstrated that F132L markedly reduces the sensitivity of the K(ATP) channel to inhibition by MgATP and this increases the whole-cell K(ATP) current. The functional consequence of this ABCC8 mutation mirrors that of KCNJ11 mutations causing neonatal diabetes and provides new insights into the interaction of Kir6.2 and SUR1. As SUR1 is expressed in neurones as well as in beta cells, this mutation can account for both neonatal diabetes and the neurological phenotype. Our results demonstrate that SUR1 mutations constitute a new genetic aetiology for neonatal diabetes and that they act by reducing the K(ATP) channel's ATP sensitivity.     

ATP-sensitive potassium channels: structures, functions, and pathophysiology

ATP-sensitive potassium channels (KATP channels) play important roles in various tissues by coupling cell metabolic status to electrical activity. Recently, molecular biological and electrophysiological techniques have revealed the molecular basis of the KATP channels to be a complex of the Kir6.0 subunit, a member of the inwardly rectifying K+ channel subfamily Kir6.0, and the sulfonylurea receptor (SUR) subunit, a member of ATP-binding cassette (ABC) superfamily; the functional diversity of the various KATP channels is being determined by a combination of the Kir6.0 subunit (Kir6.1 or Kir6.2) and the SUR subunit (SUR1 or SUR2) comprising it. Recent studies of the KATP channels have suggested mechanisms of KATP channel regulation and pathophysiology and also a new model in which ABC proteins regulate the functional expression of ion channels.     

Iptakalim protects against hypoxic brain injury through multiple pathways associated with ATP-sensitive potassium channels

The rapid and irreversible brain injury produced by anoxia when stroke occurs is well known. Cumulative evidence suggests that the activation of neuronal ATP-sensitive potassium (KATP) channels may have inherent protective effects during cerebral hypoxia, yet little information regarding the therapeutic effects of KATP channel openers is available. We hypothesized that pretreatment with a KATP channel opener might protect against brain injury induced by cerebral hypoxia. In this study, adult Wistar rats were treated with iptakalim, a new KATP channel opener, which is selective for SUR2 type KATP channels, by intragastric administration at doses of 2, 4, or 8 mg/kg/day for 7 days before being exposed to simulated high altitude equivalent to 8000 m in a decompression chamber for 8 h leading to hypoxic brain injury. By light and electron microscopic images, we observed that hypobaric hypoxia-induced brain injury could be prevented by pretreatment with iptakalim. It was also observed that the permeability of the blood-brain barrier, water content, Na+ and Ca2+ concentration, and activities of Na+,K+-ATPase, Ca2+-ATPase and Mg2+-ATPase in rat cerebral cortex were increased and the gene expression of the occludin or aquaporin-4 was down- or upregulated respectively, which could also be prevented by the pretreatment with iptakalim at doses of 2, 4, or 8 mg/kg in a dose-dependent manner. Furthermore, we found that in an oxygen-and-glucose-deprived model in ECV304 cells and rat cortical astrocytes, pretreatment with iptakalim significantly increased survived cell rates and decreased lactate dehydrogenate release, which were significantly antagonized by glibenclamide, a K(ATP) channel blocker. We conclude that iptakalim is a promising drug that may protect against brain injury induced by acute hypobaric hypoxia through multiple pathways associated with SUR2-type K(ATP) channels, suggesting a new therapeutic strategy for stroke treatment.     

Characterisation of sulphonylurea and ATP-regulated K+ channels in rat pancreatic A-cells

We have monitored whole-cell and single channel ATP-sensitive K+ (KATP) currents in isolated rat glucagon-secreting pancreatic A-cells. Tolbutamide produced a concentration-dependent decrease in the whole-cell KATP conductance (Ki = 6 microM) and initiated action potential firing. The K+ channel opener diazoxide, but not cromakalim or pinacidil, inhibited electrical activity and increased the whole-cell K+ conductance fourfold. ATP applied to the intracellular face of the membrane inhibited KATP channel activity with a Ki of 17 microM, an effect that could be counteracted by Mg-ADP and Mg-GDP. GTP and UTP did not affect KATP channel activity. Phosphatidylinositol 4,5-bisphosphate activated KATP channels inhibited by ATP after a delay of 90 s. In situ hybridisation demonstrated the expression of the mRNA encoding KATP channel subunits Kir6.2 and SUR1 but not Kir6.1 and SUR2. We conclude that rat pancreatic A-cells express KATP channels with the nucleotide-, sulphonylurea- and K+ channel-opener sensitivities expected for a channel formed by Kir6.2 and SUR1 subunits.     

Human KATP channelopathies: diseases of metabolic homeostasis

Assembly of an inward rectifier K⁺ channel pore (Kir6.1/Kir6.2) and an adenosine triphosphate (ATP)-binding regulatory subunit (SUR1/SUR2A/SUR2B) forms ATP-sensitive K⁺ (K_ATP) channel heteromultimers, widely distributed in metabolically active tissues throughout the body. K_ATP channels are metabolism-gated biosensors functioning as molecular rheostats that adjust membrane potential-dependent functions to match cellular energetic demands. Vital in the adaptive response to (patho)physiological stress, K_ATP channels serve a homeostatic role ranging from glucose regulation to cardioprotection. Accordingly, genetic variation in K_ATP channel subunits has been linked to the etiology of life-threatening human diseases. In particular, pathogenic mutations in K_ATP channels have been identified in insulin secretion disorders, namely, congenital hyperinsulinism and neonatal diabetes. Moreover, K_ATP channel defects underlie the triad of developmental delay, epilepsy, and neonatal diabetes (DEND syndrome). K_ATP channelopathies implicated in patients with mechanical and/or electrical heart disease include dilated cardiomyopathy (with ventricular arrhythmia; CMD1O) and adrenergic atrial fibrillation. A common Kir6.2 E23K polymorphism has been associated with late-onset diabetes and as a risk factor for maladaptive cardiac remodeling in the community-at-large and abnormal cardiopulmonary exercise stress performance in patients with heart failure. The overall mutation frequency within K_ATP channel genes and the spectrum of genotype–phenotype relationships remain to be established, while predicting consequences of a deficit in channel function is becoming increasingly feasible through systems biology approaches. Thus, advances in molecular medicine in the emerging field of human K_ATP channelopathies offer new opportunities for targeted individualized screening, early diagnosis, and tailored therapy.     

ATP敏感性钾离子(KATP)通道的mRNA在大鼠颈动脉体的表达

目的 探讨因缺氧引起颈动脉体K^ 流减少而导致颈动脉体神经活性增加的分子学机制。方法 利用逆转录聚合酶链反应(RT-PCR)评价大鼠颈动脉体的ATP敏感性钾离子(KATP)通道的mRNA表达。结果 在大鼠颈动脉体中内向整流性钾离子通道亚家族Kir6．1的mRNA是存在的，而Kir6．2和磺酰脲受体(SUR1 and SUR2)则未见。结论 Kir6．1 mRNA的存在提示KATP通道在颈动脉体感受缺氧应答时可能起重要作用。     

Analysis of the differential modulation of sulphonylurea block of β-cell and cardiac ATP-sensitive K+ (KATP) channels by Mg-nucleotides

Sulphonylureas stimulate insulin secretion by binding with high-affinity to the sulphonylurea receptor (SUR) subunit of the ATP-sensitive potassium (K_ATP) channel and thereby closing the channel pore (formed by four Kir6.2 subunits). In the absence of added nucleotides, the maximal block is around 60–80 %, indicating that sulphonylureas act as partial antagonists. Intracellular MgADP modulated sulphonylurea block, enhancing inhibition of Kir6.2/SUR1 (β-cell type) and decreasing that of Kir6.2/SUR2A (cardiac-type) channels. We examined the molecular basis of the different response of channels containing SUR1 and SUR2A, by recording currents from inside-out patches excised from Xenopus oocytes heterologously expressing wild-type or chimeric channels. We used the benzamido derivative meglitinide as this drug blocks Kir6.2/SUR1 and Kir6.2/SUR2A currents, reversibly and with similar potency. Our results indicate that transfer of the region containing transmembrane helices (TMs) 8–11 and the following 65 residues of SUR1 into SUR2A largely confers a SUR1-like response to MgADP and meglitinide, whereas the reverse chimera (SUR128) largely endows SUR1 with a SUR2A-type response. This effect was not specific for meglitinide, as tolbutamide was also unable to prevent MgADP activation of Kir6.2/SUR128 currents. The data favour the idea that meglitinide binding to SUR1 impairs either MgADP binding or the transduction pathway between the NBDs and Kir6.2, and that TMs 8–11 are involved in this modulatory response. The results provide a basis for understanding how β-cell K_ATP channels show enhanced sulphonylurea inhibition under physiological conditions, whereas cardiac K_ATP channels exhibit reduced block in intact cells, especially during metabolic inhibition.