Cooling positronium to ultralow velocities with a chirped laser pulse train.

When laser radiation is skilfully applied, atoms and molecules can be cooled1-3, allowing the precise measurements and control of quantum systems. This is essential for the fundamental studies of physics as well as practical applications such as precision spectroscopy4-7, ultracold gases with quantum statistical properties8-10 and quantum computing. In laser cooling, atoms are slowed to otherwise unattainable velocities through repeated cycles of laser photon absorption and spontaneous emission in random directions. Simple systems can serve as rigorous testing grounds for fundamental physics-one such case is the purely leptonic positronium11,12, an exotic atom comprising an electron and its antiparticle, the positron. Laser cooling of positronium, however, has hitherto remained unrealized. Here we demonstrate the one-dimensional laser cooling of positronium. An innovative laser system emitting a train of broadband pulses with successively increasing central frequencies was used to overcome major challenges posed by the short positronium lifetime and the effects of Doppler broadening and recoil. One-dimensional chirp cooling was used to cool a portion of the dilute positronium gas to a velocity distribution of approximately 1 K in 100 ns. A major advancement in the field of low-temperature fundamental physics of antimatter, this study on a purely leptonic system complements work on antihydrogen13, a hadron-containing exotic atom. The successful application of laser cooling to positronium affords unique opportunities to rigorously test bound-state quantum electrodynamics and to potentially realize Bose-Einstein condensation14-18 in this matter-antimatter system.

Molecular basis of a new ovine model for human 3M syndrome-2.

BACKGROUND: Brachygnathia, cardiomegaly and renal hypoplasia syndrome (BCRHS, OMIA 001595-9940 ) is a previously reported recessively inherited disorder in Australian Poll Merino/Merino sheep. Affected lambs are stillborn with various congenital defects as reflected in the name of the disease, as well as short stature, a short and broad cranium, a small thoracic cavity, thin ribs and brachysternum. The BCRHS phenotype shows similarity to certain human short stature syndromes, in particular the human 3M syndrome-2. Here we report the identification of a likely disease-causing variant and propose an ovine model for human 3M syndrome-2. RESULTS: Eight positional candidate genes were identified among the 39 genes in the approximately 1 Mb interval to which the disease was mapped previously. Obscurin like cytoskeletal adaptor 1 (OBSL1) was selected as a strong positional candidate gene based on gene function and the resulting phenotypes observed in humans with mutations in this gene. Whole genome sequencing of an affected lamb (BCRHS3) identified a likely causal variant ENSOARG00000020239:g.220472248delC within OBSL1. Sanger sequencing of seven affected, six obligate carrier, two phenotypically unaffected animals from the original flock and one unrelated control animal validated the variant. A genotyping assay was developed to genotype 583 animals from the original flock, giving an estimated allele frequency of 5%. CONCLUSIONS: The identification of a likely disease-causing variant resulting in a frameshift (p.(Val573Trpfs*119)) in the OBSL1 protein has enabled improved breeding management of the implicated flock. The opportunity for an ovine model for human 3M syndrome and ensuing therapeutic research is promising given the availability of carrier ram semen for BCRHS.

Does the shoe really fit? Characterising ill-fitting footwear among community-dwelling older adults attending geriatric services: an observational cross-sectional study.

BACKGROUND: Falls in older people are common and can result in loss of confidence, fear of falling, restriction in activity and loss of independence. Causes of falls are multi-factorial. There is a paucity of research assessing the footwear characteristics among older people who are at high risk of falls, internationally and in the Irish setting. The aim of this study was to examine the proportion of older adults attending a geriatric day hospital in Ireland who were wearing incorrectly sized shoes. METHODS: A consecutive sample of 111 older adults aged 60 years and over attending a geriatric day hospital in a large Irish teaching hospital was recruited. Demographic data including age, mobility, medications, co-habitation status, footwear worn at home and falls history were recorded. Shoe size and foot length were measured in millimetres using an internal shoe gauge and SATRA shoe size stick, respectively. Participants' self-reported shoe size was recorded. Footwear was assessed using the Footwear Assessment Form (FAF). A Timed Up and Go (TUG) score was recorded. Functional independence was assessed using the Nottingham Extended Activities of Daily Living (NEADL) Scale. The primary outcome of interest in this study was selected as having footwear within the suggested range (10 to 15 mm) on at least one foot. Participants who met this definition were compared to those with ill-fitting footwear on both feet using Chi-square tests, T-tests or Mann-Whitney U tests. RESULTS: The mean difference between shoe length and foot length was 18.6 mm (SD: 9.6 mm). Overall, 72% of participants were wearing footwear that did not fit correctly on both feet, 90% had shoes with smooth, partly worn or fully worn sole treading and 67% reported wearing slippers at home. Participant age, TUG score and NEADL score were not associated with ill-fitting footwear. CONCLUSIONS: Wearing incorrectly fitting shoes and shoes with unsafe features was common among older adults attending geriatric day services in this study. A large number of participants reported wearing slippers at home.

An ABCA12 missense variant in a Shorthorn calf with ichthyosis fetalis.

Two clinical forms of ichthyosis in cattle have been reported, ichthyosis fetalis and congenital ichthyosis. Ichthyosis poses animal welfare and economic issues and the more severe form, ichthyosis fetalis, is lethal. A Shorthorn calf with ichthyosis fetalis was investigated and a likely causal missense variant on chromosome 2 in the ABCA12 gene (NM_001191294.2:c.6776T>C) was identified by whole genome sequencing. Mutations in the ABCA12 gene are known to cause ichthyosis fetalis in cattle and Harlequin ichthyosis in humans. Sanger sequencing of the affected calf and the dam confirmed the variant was homozygous in the affected calf and heterozygous in the dam. Further genotyping of 130 Shorthorn animals from the same property revealed an estimated allele frequency of 3.8%. The presented findings enable genetic testing for breeding and diagnostics.

Inferring the recent ancestry of myostatin alleles affecting muscle mass in cattle.

Double muscling is an inherited condition in cattle characterised by large increases in muscle mass. Mutations in the myostatin (MSTN) gene, responsible for double muscling, were targeted in this study to estimate the time since the most recent common ancestor (TMRCA) for Q204X (p.Gln204*), E226X (p.Glu226*), 821del11 (c.821del11), E291X (p.Glu291*), C313Y (p.Cys313Tyr) and the more phenotypically moderate F94L (p.Phe94Leu) mutation. Genetic variability was examined in eight regions upstream and downstream of the MSTN locus. The molecular distance of the homozygous region associated with each MSTN allele was used to estimate the TMRCA. Long homozygous segments were associated with the MSTN alleles (mostly > 2 Mb), compared to short segments (130 kb) for cattle wild type at the double muscling and F94L sites. Estimates of time indicated that each MSTN allele had a recent common ancestor (<400 years ago). The results from this study, and the increasing frequency of these MSTN alleles in some cattle breeds, demonstrate recent positive selection.

Redox-optimized ROS balance: a unifying hypothesis.

While it is generally accepted that mitochondrial reactive oxygen species (ROS) balance depends on the both rate of single electron reduction of O2 to superoxide (O2.-) by the electron transport chain and the rate of scavenging by intracellular antioxidant pathways, considerable controversy exists regarding the conditions leading to oxidative stress in intact cells versus isolated mitochondria. Here, we postulate that mitochondria have been evolutionarily optimized to maximize energy output while keeping ROS overflow to a minimum by operating in an intermediate redox state. We show that at the extremes of reduction or oxidation of the redox couples involved in electron transport (NADH/NAD+) or ROS scavenging (NADPH/NADP+, GSH/GSSG), respectively, ROS balance is lost. This results in a net overflow of ROS that increases as one moves farther away from the optimal redox potential. At more reduced mitochondrial redox potentials, ROS production exceeds scavenging, while under more oxidizing conditions (e.g., at higher workloads) antioxidant defenses can be compromised and eventually overwhelmed. Experimental support for this hypothesis is provided in both cardiomyocytes and in isolated mitochondria from guinea pig hearts. The model reconciles, within a single framework, observations that isolated mitochondria tend to display increased oxidative stress at high reduction potentials (and high mitochondrial membrane potential, Psim), whereas intact cardiac cells can display oxidative stress either when mitochondria become more uncoupled (i.e., low Psim) or when mitochondria are maximally reduced (as in ischemia or hypoxia). The continuum described by the model has the potential to account for many disparate experimental observations and also provides a rationale for graded physiological ROS signaling at redox potentials near the minimum.

Oscillations of membrane current and excitability driven by metabolic oscillations in heart cells.

Periodic changes in membrane ionic current linked to intrinsic oscillations of energy metabolism were identified in guinea pig cardiomyocytes. Metabolic stress initiated cyclical activation of adenosine triphosphate-sensitive potassium current and concomitant suppression of depolarization-evoked intracellular calcium transients. The oscillations in membrane current and excitation-contraction coupling were linked to oscillations in the oxidation state of pyridine nucleotides but were not driven by pacemaker currents or alterations in the concentration of cytosolic calcium. Interventions that altered the rate of glucose metabolism modulated the oscillations, suggesting that the rhythms originated at the level of glycolysis. The energy-driven oscillations in potassium currents produced cyclical changes in the cardiac action potential and thus may contribute to the genesis of arrhythmias during metabolic compromise.

Phosphorylation-independent modulation of L-type calcium channels by magnesium-nucleotide complexes.

Free magnesium ions and magnesium-nucleotide complexes can exert opposite effects on many fundamental cellular processes. Although increases in the intracellular concentration of magnesium ions inhibit the L-type calcium current in heart cells, magnesium-adenosine triphosphate complexes (MgATP) would be expected to increase the current by promoting channel phosphorylation. Rapid increases in the intracellular concentration of MgATP induced by flash photolysis of caged magnesium or caged ATP resulted in enhanced calcium current. The increase in calcium current was not prevented by blocking phosphorylation, revealing a previously unrecognized direct regulatory action of the magnesium-nucleotide complex.

Suspected hereditary cervicothoracic vertebral subluxation with cervical myopathy in Poll Merino sheep.

BACKGROUND: Cervicothoracic vertebral subluxation in sheep presents as a postural and locomotor disorder, and has been described in several breeds in Australia and overseas. Cervical myopathy may also be present in these cases. CASE REPORT: A New South Wales sheep producer reported a postural and locomotor disorder with a low prevalence in his Poll Merino stud flock, affecting neonate, weaner and adult sheep. Animals with postural abnormalities, variable degrees of ataxia and proprioceptive deficits involving both fore and hind limbs were described. Abnormalities of the cervicothoracic vertebral column were identified grossly during necropsy, with misalignment and consequent narrowing of the posterior cervical spinal canal. Lesions ranging from pallor (cellular degeneration) to white streaky lesions with pinpoint haemorrhage (necrosis) were identified in the cervicothoracic paravertebral musculature of affected animals. Boney abnormalities were further characterised by imaging studies. Pedigree analysis of the very extensive breeding and disease incident records available for this flock suggested that the disease was inherited. A similar case recognised in a separate New South Wales Poll Merino flock is also described. CONCLUSION: This report describes an entity of cervicothoracic vertebral subluxation in two Poll Merino sheep flocks, with cervical myopathy also identified in one, with preliminary evidence in the primary case that there is likely to be a hereditary basis. The two cases outlined in this report resemble the findings of several historical investigations into ovine flock postural disorders in Australia and beyond.

Molecular evidence to suggest pigeon-type Chlamydia psittaci in association with an equine foal loss.

Chlamydia psittaci is an important avian pathogen with spillover from infected wild and domesticated birds also posing a risk to human health. We recently reported a case of C. psittaci equine placentitis associated with further spillover to humans. Molecular typing of this case revealed it belonged to the 6BC clade of C. psittaci, a globally distributed highly virulent set of strains, typically linked to infection spillover from parrots. Equine chlamydiosis associated with C. psittaci infection has previously been reported elsewhere in countries where parrots are not endemic, however, raising questions over the identity of infecting C. psittaci strains and the potential infection reservoirs. In this study, we describe the detection and molecular characterization of C. psittaci in a case of equine abortion in southern Queensland. Equine placenta and fresh liver and lung tissue from the necropsied foetus were positive by C. psittaci-specific qPCR. Chlamydia psittaci-specific multilocus sequence typing and ompA genotyping were used to further characterize the detected equine strains and an additional strain obtained from a dove from a different geographic region presenting with psittacosis. Molecular typing of this case revealed that the infecting equine strains were closely related to the C0sittaci detected in dove, all belonging to an evolutionary lineage of C. psittaci strains typically associated with infections of pigeons globally. This finding suggests a broader diversity of C. psittaci strains may be detected in horses and in association with reproductive loss, highlighting the need for an expansion of surveillance studies globally to understand the epidemiology of equine chlamydiosis and the associated zoonotic risk.

Equine chlamydiosis-An emerging infectious disease requiring a one health surveillance approach.

Psittacosis is a rare but potentially fatal zoonosis caused by Chlamydia psittaci, an organism that is typically associated with bird contact. However C. psittaci is capable of infecting other non-avian hosts, such as horses, sheep, cattle and goats. Stud staff and veterinarians have significant exposure to parturient animals and reproductive materials in their routine work. To investigate the zoonotic potential associated with the emergence of C. psittaci as an abortifacient agent in horses, we established a programme of joint human and animal surveillance in a sentinel horse-breeding region in Australia. This programme comprised cross-notification of equine cases to public health agencies, and active follow-up of known human contacts, including stud workers, foaling staff, veterinarians and laboratory staff. We identified no confirmed cases of acute psittacosis despite intensive surveillance and testing of heavily exposed contacts; however, further work in the area is needed.

Myocardial K(ATP) channels in preconditioning.

We are on the brink of harnessing the cell's natural defenses against ischemia and reperfusion injury after years of research into the destructive and protective mechanisms involved. Since the discovery of ischemic preconditioning, the surface receptors and signal transduction pathways underlying this phenomenon have been clarified, but many questions remain about the downstream targets that ultimately protect the cell. ATP-sensitive K(+) (K(ATP)) channels are thought to play a role in protection, but their mechanism of action has been unclear. Accumulating evidence now suggests that the location of the K(ATP) channels relevant to cytoprotection may be on the mitochondrial inner membrane instead of on the sarcolemma of the cardiac cell. This review discusses recent findings and unanswered questions about the role of K(ATP) channels in preconditioning and protection.

Enhanced Ca(2+)-activated Na(+)-Ca(2+) exchange activity in canine pacing-induced heart failure.

Defective excitation-contraction coupling in heart failure is generally associated with both a reduction in sarcoplasmic reticulum (SR) Ca(2+) uptake and a greater dependence on transsarcolemmal Na(+)-Ca(2+) exchange (NCX) for Ca(2+) removal. Although a relative increase in NCX is expected when SR function is impaired, few and contradictory studies have addressed whether there is an absolute increase in NCX activity. The present study examines in detail NCX density and function in left ventricular midmyocardial myocytes isolated from normal or tachycardic pacing-induced failing canine hearts. No change of NCX current density was evident in myocytes from failing hearts when intracellular Ca(2+) ([Ca(2+)](i)) was buffered to 200 nmol/L. However, when [Ca(2+)](i) was minimally buffered with 50 micromol/L indo-1, Ca(2+) extrusion via NCX during caffeine application was doubled in failing versus normal cells. In other voltage-clamp experiments in which SR uptake was blocked with thapsigargin, both reverse-mode and forward-mode NCX currents and Ca(2+) transport were increased >2-fold in failing cells. These results suggest that, in addition to a relative increase in NCX function as a consequence of defective SR Ca(2+) uptake, there is an absolute increase in NCX function that depends on [Ca(2+)](i) in the failing heart.

Mitochondrial ATP-sensitive potassium channels inhibit apoptosis induced by oxidative stress in cardiac cells.

Mitochondria can either enhance or suppress cell death. Cytochrome c release from mitochondria and depolarization of the mitochondrial membrane potential (DeltaPsi) are crucial events in triggering apoptosis. In contrast, activation of mitochondrial ATP-sensitive potassium (mitoK(ATP)) channels prevents lethal ischemic injury in vivo, implicating these channels as key players in the process of ischemic preconditioning. We probed the relationship between mitoK(ATP) channels and apoptosis in cultured neonatal rat cardiac ventricular myocytes. Incubation with 200 micromol/L hydrogen peroxide induced TUNEL positivity, cytochrome c translocation, caspase-3 activation, poly(ADP-ribose) polymerase cleavage, and dissipation of DeltaPsi. Pharmacological opening of mitoK(ATP) channels by diazoxide (100 micromol/L) preserved mitochondrial integrity and suppressed all markers of apoptosis. Diazoxide prevented DeltaPsi depolarization in a concentration-dependent manner (EC(50) approximately 40 micromol/L, with saturation by 100 micromol/L), as shown by both flow cytometry and quantitative image analysis of cells stained with fluorescent DeltaPsi indicators. These cytoprotective effects of diazoxide were reproduced by pinacidil, another mitoK(ATP) agonist, and blocked by the mitoK(ATP) channel antagonist 5-hydroxydecanoate (500 micromol/L). Our findings identify a novel mitochondrial pathway that is protective against apoptosis. The results also pinpoint mitoK(ATP) channels as logical therapeutic targets in diseases of enhanced apoptosis and oxidative stress.

Mitochondrial ATP-sensitive potassium channels attenuate matrix Ca(2+) overload during simulated ischemia and reperfusion: possible mechanism of cardioprotection.

Mitochondrial ATP-sensitive potassium (mitoK(ATP)) channels play a key role in ischemic preconditioning of the heart. However, the mechanism of cardioprotection remains controversial. We measured rhod-2 fluorescence in adult rabbit ventricular cardiomyocytes as an index of mitochondrial matrix Ca(2+) concentration ([Ca(2+)](m)), using time-lapse confocal microscopy. To simulate ischemia and reperfusion (I/R), cells were exposed to metabolic inhibition (50 minutes) followed by washout with control solution. Rhod-2 fluorescence gradually increased during simulated ischemia and rose even further with reperfusion. The mitoK(ATP) channel opener diazoxide attenuated the accumulation of [Ca(2+)](m) during simulated I/R (EC(50)=18 micromol/L). These effects of diazoxide were blocked by the mitoK(ATP) channel antagonist 5-hydroxydecanoate (5HD). In contrast, inhibitors of the mitochondrial permeability transition (MPT), cyclosporin A and bongkrekic acid, did not alter [Ca(2+)](m) accumulation during ischemia, but markedly suppressed the surge in rhod-2 fluorescence during reperfusion. Measurements of mitochondrial membrane potential, DeltaPsi(m), in permeabilized myocytes revealed that diazoxide depolarized DeltaPsi(m) (by 12% at 10 micromol/L, P<0.01) in a 5HD-inhibitable manner. Our data support the hypothesis that attenuation of mitochondrial Ca(2+) overload, as a consequence of partial mitochondrial membrane depolarization by mitoK(ATP) channels, underlies cardioprotection. Furthermore, mitoK(ATP) channels and the MPT differentially affect mitochondrial calcium homeostasis: mitoK(ATP) channels suppress calcium accumulation during I/R, while the MPT comes into play only upon reperfusion.

Decreased sarcoplasmic reticulum calcium content is responsible for defective excitation-contraction coupling in canine heart failure.

BACKGROUND: Altered excitation-contraction (E-C) coupling in canine pacing-induced heart failure involves decreased sarcoplasmic reticulum (SR) Ca uptake and enhanced Na/Ca exchange, which could be expected to decrease SR Ca content (Ca(SR)) and may explain the reduced intracellular Ca (Ca(i)) transient. Studies in other failure models have suggested that the intrinsic coupling between L-type Ca current (I:(Ca,L)) and SR Ca release is reduced without a change in SR Ca load. The present study investigates whether Ca(SR) and/or coupling is altered in midmyocardial myocytes from failing canine hearts (F). METHODS AND RESULTS: Myocytes were indo-1-loaded via patch pipette (37 degrees C), and Ca(i) transients were elicited with voltage-clamp steps applied at various frequencies. I(Ca,L) density was not significantly decreased in F, but steady-state Ca(i) transients were reduced to 20% to 40% of normal myocytes (N). Ca(SR), measured by integrating Na/Ca exchange currents during caffeine-induced release, was profoundly decreased in F, to 15% to 25% of N. When Ca(SR) was normalized in F by preloading in 5 mmol/L external Ca before a test pulse at 2 mmol/L Ca, a normal-amplitude Ca(i) transient was elicited. E-C coupling gain was dependent on Ca(SR) but was affected similarly in both groups, indicating that intrinsic coupling is unaltered in F. CONCLUSIONS: A decrease in Ca(SR) is sufficient to explain the diminished Ca(i) transients in F, without a change in the effectiveness of coupling. Therefore, therapeutic approaches that increase Ca(SR) may be able to fully correct the Ca handling deficit in heart failure.

Selective pharmacological agents implicate mitochondrial but not sarcolemmal K(ATP) channels in ischemic cardioprotection.

BACKGROUND: Pharmacological evidence has implicated ATP-sensitive K(+) (K(ATP)) channels as the effectors of cardioprotection, but the relative roles of mitochondrial (mitoK(ATP)) and sarcolemmal (surfaceK(ATP)) channels remain controversial. METHODS AND RESULTS: We examined the effects of the K(ATP) channel blocker HMR1098 and the K(ATP) channel opener P-1075 on surfaceK(ATP) and mitoK(ATP) channels in rabbit ventricular myocytes. HMR1098 (30 micromol/L) inhibited the surfaceK(ATP) current activated by metabolic inhibition, whereas the drug did not blunt diazoxide (100 micromol/L)-induced flavoprotein oxidation, an index of mitoK(ATP) channel activity. P-1075 (30 micromol/L) did not increase flavoprotein oxidation but did elicit a robust surfaceK(ATP) current that was completely inhibited by HMR1098. These results indicate that HMR1098 selectively inhibits surfaceK(ATP) channels, whereas P-1075 selectively activates surface K(ATP) channels. In a cellular model of simulated ischemia, the mitoK(ATP) channel opener diazoxide (100 micromol/L), but not P-1075, blunted cellular injury. The cardioprotection afforded by diazoxide or by preconditioning was prevented by the mitoK(ATP) channel blocker 5-hydroxydecanoate (500 micromol/L) but not by the surfaceK(ATP) channel blocker HMR1098 (30 micromol/L). CONCLUSIONS: The cellular effects of mitochondria- or surface-selective agents provide further support for the emerging consensus that mitoK(ATP) channels rather than surfaceK(ATP) channels are the likely effectors of cardioprotection.

Adenosine primes the opening of mitochondrial ATP-sensitive potassium channels: a key step in ischemic preconditioning?

BACKGROUND: Adenosine can initiate ischemic preconditioning, and mitochondrial ATP-sensitive potassium (K(ATP)) channels have emerged as the likely effectors. We sought to determine the mechanistic interactions between these 2 observations. METHODS AND RESULTS: The mitochondrial flavoprotein oxidation induced by diazoxide (100 micromol/L) was used to quantify mitochondrial K(ATP) channel activity in intact rabbit ventricular myocytes. Adenosine (100 micromol/L) increased mitochondrial K(ATP) channel activity and abbreviated the latency to mitochondrial K(ATP) channel opening. These potentiating effects were entirely prevented by the adenosine receptor antagonist 8-(p-sulfophenyl)-theophylline (100 micromol/L) or by the protein kinase C inhibitor polymyxin B (50 micromol/L). The effects of adenosine and diazoxide reflected mitochondrial K(ATP) channel activation, because they could be blocked by the mitochondrial K(ATP) channel blocker 5-hydroxydecanoate (500 micromol/L). In a cellular model of simulated ischemia, adenosine mitigated cell injury; this cardioprotective effect was blocked by 5-hydroxydecanoate but not by the surface-selective K(ATP) channel blocker HMR1098. Moreover, adenosine augmented the cardioprotective effect of diazoxide. A quantitative model of mitochondrial K(ATP) channel gating reproduced the major experimental findings. CONCLUSIONS: Our results support the hypothesis that adenosine receptor activation primes the opening of mitochondrial K(ATP) channels in a protein kinase C-dependent manner. The findings provide tangible links among various key elements in the preconditioning cascade.

Activation of mitochondrial ATP-dependent potassium channels by nitric oxide.

BACKGROUND: Nitric oxide (NO) has been implicated as a mediator of "second-window" ischemic preconditioning, and mitochondrial ATP-dependent K(+) (mitoK(ATP)) channels are the likely effectors. The links between NO and mitoK(ATP) channels are unknown. METHODS AND RESULTS: We measured mitochondrial redox potential as an index of mitoK(ATP) channel opening in rabbit ventricular myocytes. The NO donor S-nitroso-N-acetyl-DL-penicillamine (SNAP, 0.1 to 1 mmol/L) oxidized the mitochondrial matrix dose-dependently without activating sarcolemmal K(ATP) channels. SNAP-induced oxidation was blocked by the selective mitoK(ATP) channel blocker 5-hydroxydecanoate and by the NO scavenger 2-(4-carboxyphenyl)-4,4', 5,5'-tetramethylimidazole-1-oxyl-3-oxide. SNAP-induced mitochondrial oxidation was detectable either by photomultiplier tube recordings of flavoprotein fluorescence or by confocal imaging. SNAP also enhanced the oxidative effects of diazoxide when both agents were applied together. Exposure to 1 mmol/L 8Br-cGMP failed to mimic the effects of SNAP. CONCLUSIONS: NO directly activates mitoK(ATP) channels and potentiates the ability of diazoxide to open these channels. These results provide novel mechanistic links between NO-induced cardioprotection and mitoK(ATP) channels.

Nicorandil, a potent cardioprotective agent, acts by opening mitochondrial ATP-dependent potassium channels.

OBJECTIVES: To determine the mechanism of cardioprotection afforded by nicorandil, an orally efficacious antianginal drug, we examined its effects on ATP-dependent potassium (K(ATP)) channels. BACKGROUND: Nicorandil can mimic ischemic preconditioning, while mitochondrial K(ATP) (mitoK(ATP)) channels rather than sarcolemmal K(ATP) (surfaceK(ATP)) channels have emerged as the likely effectors. METHODS: Flavoprotein fluorescence and membrane current in intact rabbit ventricular myocytes were measured simultaneously to assay mitoK(ATP) channel and surface K(ATP) channel activities, respectively. In a cell-pelleting model of ischemia, cells permeable to trypan blue were counted as killed by 60 and 120 min of ischemia. RESULTS: Nicorandil (100 micromol/liter) increased flavoprotein oxidation but not membrane current; a 10-fold higher concentration recruits both mitoK(ATP) and surfaceK(ATP) channels. Pooled dose-response data confirm that nicorandil concentrations as low as 10 micromol/liter turn on mitoK(ATP) channels, while surfaceK(ATP) current requires exposure to millimolar concentrations. Nicorandil blunted the rate of cell death in a pelleting model of ischemia; this cardioprotective effect was prevented by the mitoK(ATP) channel blocker 5-hydroxydecanoate but was unaffected by the surfaceK(ATP) channel blocker HMR1098. CONCLUSIONS: Nicorandil exerts a direct cardioprotective effect on heart muscle cells, an effect mediated by selective activation of mitoK(ATP) channels.