Effects of activated carbon on reductive dechlorination of PCBs by organohalide respiring bacteria indigenous to sediments.

Polychlorinated biphenyls (PCBs) have accumulated in aquatic sediments due to their inherent chemical stability and their presence poses a risk due to their potential toxicity in humans and animals. Granular activated carbon (GAC) has been applied to PCB contaminated sediment sites to reduce the aqueous concentration by sequestration thus reducing the PCB exposure and toxicity to both benthic and aquatic organisms. However, it is not known how the reduction of PCB bioavailability by adsorption to GAC affects bacterial transformation of PCBs by indigenous organohalide respiring bacteria. In this study, the impact of GAC on anaerobic dechlorination by putative organohalide respiring bacteria indigenous to sediment from Baltimore Harbor was examined. It was shown that the average Cl/biphenyl after dehalogenation of Aroclor 1260 was similar between treatments with and without GAC amendment. However, GAC caused a substantial shift in the congener distribution whereby a smaller fraction of highly chlorinated congeners was more extensively dechlorinated to mono- through tri-chlorinated congeners compared to the formation of tri- through penta-chlorinated congeners in unamended sediment. The results combined with comparative sequence analysis of 16S rRNA gene sequences suggest that GAC caused a community shift to putative organohalide respiring phylotypes that coincided with more extensive dechlorination of ortho and unflanked chlorines. This shift in activity by GAC shown here for the first time has the potential to promote greater degradation in situ by promoting accumulation of less chlorinated congeners that are generally more susceptible to complete mineralization by aerobic PCB degrading bacteria.

A randomized trial of high-dose vitamin D2 in relapsing-remitting multiple sclerosis.

OBJECTIVE: Higher latitude, lower ultraviolet exposure, and lower serum 25-hydroxyvitamin D (25OHD) correlate with higher multiple sclerosis (MS) prevalence, relapse rate, and mortality. We therefore evaluated the effects of high-dose vitamin D2 (D2) in MS. METHODS: Adults with clinically active relapsing-remitting MS (RRMS) were randomized to 6 months' double-blind placebo-controlled high-dose vitamin D2, 6,000 IU capsules, dose adjusted empirically aiming for a serum 25OHD 130-175 nM. All received daily low-dose (1,000 IU) D2 to prevent deficiency. Brain MRIs were performed at baseline, 4, 5, and 6 months. Primary endpoints were the cumulative number of new gadolinium-enhancing lesions and change in the total volume of T2 lesions. Secondary endpoints were Expanded Disability Status Scale (EDSS) score and relapses. RESULTS: Twenty-three people were randomized, of whom 19 were on established interferon or glatiramer acetate (Copaxone) treatment. Median 25OHD rose from 54 to 69 nM (low-dose D2) vs 59 to 120 nM (high-dose D2) (p = 0.002). No significant treatment differences were detected in the primary MRI endpoints. Exit EDSS, after adjustment for entry EDSS, was higher following high-dose D2 than following low-dose D2 (p = 0.05). There were 4 relapses with high-dose D2 vs none with low-dose D2 (p = 0.04). CONCLUSION: We did not find a therapeutic advantage in RRMS for high-dose D2 over low-dose D2 supplementation. CLASSIFICATION OF EVIDENCE: This study provides Class I evidence that high-dose vitamin D2 (targeting 25OHD 130-175 nM), compared to low-dose supplementation (1,000 IU/d), was not effective in reducing MRI lesions in patients with RRMS.

Laminin 332 deposition is diminished in irradiated skin in an animal model of combined radiation and wound skin injury.

Skin exposure to ionizing radiation affects the normal wound healing process and greatly impacts the prognosis of affected individuals. We investigated the effect of ionizing radiation on wound healing in a rat model of combined radiation and wound skin injury. Using a soft X-ray beam, a single dose of ionizing radiation (10-40 Gy) was delivered to the skin without significant exposure to internal organs. At 1 h postirradiation, two skin wounds were made on the back of each rat. Control and experimental animals were euthanized at 3, 7, 14, 21 and 30 days postirradiation. The wound areas were measured, and tissue samples were evaluated for laminin 332 and matrix metalloproteinase (MMP) 2 expression. Our results clearly demonstrate that radiation exposure significantly delayed wound healing in a dose-related manner. Evaluation of irradiated and wounded skin showed decreased deposition of laminin 332 protein in the epidermal basement membrane together with an elevated expression of all three laminin 332 genes within 3 days postirradiation. The elevated laminin 332 gene expression was paralleled by an elevated gene and protein expression of MMP2, suggesting that the reduced amount of laminin 332 in irradiated skin is due to an imbalance between laminin 332 secretion and its accelerated processing by elevated tissue metalloproteinases. Western blot analysis of cultured rat keratinocytes showed decreased laminin 332 deposition by irradiated cells, and incubation of irradiated keratinocytes with MMP inhibitor significantly increased the amount of deposited laminin 332. Furthermore, irradiated keratinocytes exhibited a longer time to close an artificial wound, and this delay was partially corrected by seeding keratinocytes on laminin 332-coated plates. These data strongly suggest that laminin 332 deposition is inhibited by ionizing radiation and, in combination with slower keratinocyte migration, can contribute to the delayed wound healing of irradiated skin.

A new HLA-A*31 null allele, A*3114N.

A new HLA-A*31 null allele results from addition of an extra C near the beginning of exon 4 after a string of seven Cs.

Concentrations of polychlorinated biphenyl congeners in cultivated oysters (Crassostrea gigas) in western Taiwan.

Spatial and temporal variations of polychlorinated biphenyl (PCB) concentrations in cultivated oysters from five aquaculture areas along the western coast of Taiwan were investigated. Poor correlations between total PCB concentrations (ng/g dry weight [dw]) and physiologic parameters of oysters (shell length, width, thickness, and lipid content) were found. Total PCB concentrations ranged from 3.4 to 94 cng/g dw. The highest value was found in oysters from the Lu'ermen aquaculture area, which receives wastewater from a sodium hydroxide and pentachlorophenol factory. Furthermore, principal component analysis confirmed that the PCB congener pattern in this area was distinct from others and that two additional pollution sources might exist in the Tainan and Hsinchu areas. Oysters with PCB concentrations > 1 SD above the geometric mean were found mainly in Tainan (stations TN5 to TN9) with only one increased concentration in the Yunlin (station YL2) and Hsinchu (station HC4) aquaculture areas. Except for four confined stations, TN3 to TN6, where total PCB concentrations were higher (p = 0.028) in the warmer (May and July) than in the colder season (November and March), seasonal variation of total PCB concentrations in oysters was not significant. Geometric mean and geometric mean plus 1 SD of total PCB concentrations in this study are lower than those in South Korea and the United States (Mussel Watch).

HLA class I null alleles and new alleles affect unrelated bone marrow donor searches.

Unrecognized HLA null alleles or new alleles may affect the outcome of bone marrow transplants using unrelated donors. Some reports suggest that null alleles occur in the range of 0.003-0.07% (1, 2), which has led some transplant programs to stop performing serologic typing. We describe nine cases involving expression variants or new alleles. Three cases involved expression variants, including two null alleles and A*24020102L. One of the null alleles was a new variant of A*02. Seven cases involved new alleles. In five cases, there where discrepancies between HLA typing by serology and PCR-SSP. These included the three expression variants, one new B40 allele that typed serologically as B41 and one new B*07 allele that typed serologically as B42. Eight of these cases were found in the course of typing bone marrow transplant patients or potential unrelated donors since May of 2001 (total tested, 710 patients, 1914 donors). Thus, the incidence of null alleles was two in 2,624 (0.08%). Sequence-based typing (SBT) was performed on 676 of these samples. The decision to perform SBT was influenced by finding a serologic typing discrepancy in two cases. In one of those cases, SBT would probably have been performed at a later time, prior to final selection of a donor. Thus, the incidence of new alleles was between 4 and 6 of 676 (0.59-0.89%). We conclude that new HLA alleles and null alleles are uncommon but not extremely rare, and they continue to affect a significant number of unrelated donor searches.

Comparing polybrominated diphenyl ether and polychlorinated biphenyl bioaccumulation in a food web in Grand Traverse Bay, Lake Michigan.

Levels of polybrominated diphenyl ethers (BDEs) in Great Lakes salmonids and ambient air have been recently reported, but few studies worldwide have examined the accumulation of BDEs within aquatic food webs. Here we report some of the first measurements of six BDE congeners that are common components of the pentaBDE commercial mixture within an entire Lake Michigan food web. BDEs were detected in all samples and the dominant BDE congener was 2,2',4,4'-tetrabromodiphenyl ether (BDE 47). BDE 47 levels were consistently greater than those of the 2,2',4,4',5-pentabromodiphenyl ether (BDE 99), despite similar levels of these two compounds in commercial mixtures, suggesting differences in the bioavailability of the BDE congeners or differences in their ability to be metabolized. Additionally, congener composition was significantly different among deepwater sculpin, bloater chub, and lake trout, indicating differences in exposure or differences in biotransformation capacities. Total BDE concentrations in this food web were positively correlated (r = 0.94) with levels of PCBs previously measured in these samples (Stapleton et al. 2001a). Levels of BDE 47 and PCB 153, compounds with similar physicochemical properties, were compared to examine the relative exposure and bioaccumulation of these two classes of chemicals that have different environmental loading histories. Food web magnification factors calculated for these two congeners were 3.2 and 4.0 for BDE 47 and PCB 153, respectively, indicating a comparable potential for biomagnification in food webs.

cDNA cloning, purification, properties, and function of a beta-1,3-glucan recognition protein from a pyralid moth, Plodia interpunctella.

Microorganisms possess distinctive biochemical or molecular patterns on their cell surfaces, such as those formed by the lipopolysaccharides, lipoteichoic acids, and/or peptidoglycans of bacteria and the beta-1,3-glucans of fungi. Pattern recognition proteins that bind to these surface moieties have been implicated in the activation of the innate immune response in insects and other invertebrates. We report the purification and cloning of a cDNA for a 53-kDa beta-1,3-glucan recognition protein (betaGRP) from the Indianmeal moth, Plodia interpunctella (Hübner) (Lepidoptera: Pyralidae). BetaGRP cDNA contains an open reading frame that encodes 488 amino acids, of which the first 17 residues comprise the secretion signal peptide. The calculated molecular mass of the 471-residue mature protein is 53,311 Da. The protein consists of a carboxyl-terminal domain that is similar to other recognition proteins from invertebrates, beta-1,3-glucanases from bacteria, and a beta-1,3-glucanase from the sea urchin, Strongylocentrotus purpuratus. The amino-terminus of betaGRP shares sequence similarity with other invertebrate recognition molecules and the beta-1,3-glucanase from S. purpuratus. Affinity purification of a 53-kDa protein and subsequent sequencing of a peptide produced by tryptic cleavage confirmed the presence of the betaGRP in P. interpunctella larval hemolymph. RT-PCR analysis indicates that betaGRP is constitutively expressed in all life-stages, with no detectable induction following exposure of wandering larvae to microbial elicitors. Northern blot analysis indicates that the 1.8-kb betaGRP transcript is transcribed within the fat body. Recombinant betaGRP retains beta-1,3-glucan-binding activity, binds to lipopolysaccharide and lipoteichoic acid in vitro, causes aggregation of microorganisms, and activates the prophenoloxidase cascade in the presence of soluble beta-1,3-glucan. These data support the hypothesis that the 53-kDa betaGRP functions to recognize pathogen surface molecules as nonself and subsequently activates insect innate immune responses.

Trigeminal and polyradiculoneuritis in a dog presenting with masticatory muscle atrophy and Horner's syndrome.

A 9-year-old, spayed female, Airedale Terrier was euthanatized and necropsied after a progressive clinical course that included Horner's syndrome of the left eye and unilateral atrophy of the masticatory muscles. Although gross lesions were limited, a polyradiculoneuritis and ganglionitis that was most severe in the trigeminal nerves and ganglia were confirmed histologically. The inflammatory infiltrate consisted predominantly of macrophages and B and T lymphocytes that were phenotypically confirmed by immunostaining. Horner's syndrome was the result of damage to postganglionic sympathetic fibers that were incorporated in segments of the inflamed trigeminal nerve and its ophthalmic branch. Histologically, the character and distribution of the inflammation was similar to previously described syndromes of suspected immune-mediated etiology in humans and animals.

Recent declines in PAH, PCB, and toxaphene levels in the northern Great Lakes as determined from high resolution sediment cores.

Sediment cores were collected from two sites in Grand Traverse Bay, Lake Michigan in May 1998, dated using 210Pb geochronology, and analyzed for polychlorinated biphenyl (PCB) congeners, polycyclic aromatic hydrocarbons (PAHs), and toxaphene. The extraordinarily high sediment focusing and accumulation rates in these cores relative to other Great Lakes sediments allowed quantification of high-resolution temporal trends in the burial of hydrophobic organic contaminants. The focus-corrected accumulation rate of total PCBs (sum of 105 congeners) in 1998 was 0.50 ng/cm2-year at both sites. Toxaphene and total PAH (t-PAH; sum of 33 compounds) surficial accumulations varied at each site and ranged from 0.08 to 0.41 ng/cm2-year for toxaphene and 25 to 52 ng/cm2-yr for t-PAHs at the two sites. The maximum t-PAH accumulation rate was in sediment dated from 1942, and PAH accumulation decreased from 1942 to 1980 with a first-order rate of decline 0.017 yr(-1). Both toxaphene and t-PCB accumulations peaked in sediment deposited in 1972, afterwhich their accumulations decreased with nearly identical rates of decline (0.027 yr(-1) and 0.028 yr(-1), respectively).

Accumulation of atmospheric and sedimentary PCBs and toxaphene in a Lake Michigan food web.

Seston, sediment, settling organic matter, and food web members were collected from Grand Traverse Bay, Lake Michigan, between April 1997 and September 1998 to examine PCB and toxaphene biomagnification. Stable isotopes of nitrogen and carbon were analyzed in samples and used to establish trophic structure of the food web and to determine the importance of atmospheric versus sedimentary sources in delivering PCBs to the food web. Nitrogen isotopes were confounded by multiple variables in this system, particularly seasonal variation, and did not display a simple pattern of enrichment among trophic levels. However, delta13C displayed little seasonal variation and was positively correlated with PCB concentrations among food web members (r2 = 0.69). Plots of delta13C vs PCBs separate food web members into three distinct groupings comprised of invertebrates, primary forage fish, and predatory fish. Stable isotope values of the primary organic sources indicate that the atmosphere, and not the sediment, is the most likely source of PCBs to the food web of Lake Michigan. Additionally, we suggest that seston may be important in delivering PCBs to pelagic food web members and species that receive a majority of their nutrition through pelagic sources. In contrast, settling particles are implicated in delivering PCBs to benthic organisms and Mysis relicta.

Sarcolemmal and mitochondrial K(atp)channels mediate cardioprotection in chronically hypoxic hearts.

X. Kong, J. S. Tweddell, G. J. Gross and J. E. Baker. Sarcolemmal and Mitochondrial K(ATP)Channels Mediate Cardioprotection in Chronically Hypoxic Hearts. Journal of Molecular and Cellular Cardiology (2001) 33, 1041-1045. Hypoxia from birth increases the resistance of the isolated neonatal heart to ischemia. We determined if increased resistance to ischemia was due to activation of sarcolemmal or mitochondrial K(ATP)channels. Rabbits (n=8/group) were raised from birth in a normoxic (F(I)O(2)=0.21) or hypoxic (F(I)O(2)=0.12) environment for 8-10 days and the heart perfused with Krebs-Henseleit bicarbonate buffer. A mitochondrial-selective K(ATP)channel blocker 5-hydroxydecanoate (5-HD) (300 micromol/l) or a sarcolemmal-selective K(ATP)channel blocker HMR 1098 (30 micromol/l) were added alone or in combination for 20 min prior to a global ischemic period of 30 min, followed by 35 min reperfusion. Recovery of ventricular developed pressure was higher in chronically hypoxic than normoxic hearts. 5-HD and HMR 1098 partially reduced the cardioprotective effect of chronic hypoxia, but had no effect in normoxic hearts. The combination of 5-HD and HMR 1098 abolished the cardioprotective effect of chronic hypoxia. We conclude that both sarcolemmal and mitochondrial K(ATP)channels contribute to cardioprotection in the chronically hypoxic heart.

Heat shock protein 90 mediates the balance of nitric oxide and superoxide anion from endothelial nitric-oxide synthase.

The balance of nitric oxide (.NO) and superoxide anion (O(2)) plays an important role in vascular biology. The association of heat shock protein 90 (Hsp90) with endothelial nitric-oxide synthase (eNOS) is a critical step in the mechanisms by which eNOS generates.NO. As eNOS is capable of generating both.NO and O(2), we hypothesized that Hsp90 might also mediate eNOS-dependent O(2) production. To test this hypothesis, bovine coronary endothelial cells (BCEC) were pretreated with geldanamycin (GA, 10 microg/ml; 17.8 microm) and then stimulated with the calcium ionophore, (5 microm). GA significantly decreased -stimulated eNOS-dependent nitrite production (p < 0.001, n = 4) and significantly increased -stimulated eNOS-dependent O(2) production (p < 0.001, n = 8). increased phospho-eNOS(Ser-1179) levels by >1.6-fold over vehicle (V)-treated levels. Pretreatment with GA by itself or with increased phospho-eNOS levels. In unstimulated V-treated BCEC cultures low amounts of Hsp90 were found to associate with eNOS. Pretreatment with GA and/or increased the association of Hsp90 with eNOS. These data show that Hsp90 is essential for eNOS-dependent.NO production and that inhibition of ATP-dependent conformational changes in Hsp90 uncouples eNOS activity and increases eNOS-dependent O(2) production.

Protein tyrosine phosphatase CD148-mediated inhibition of T-cell receptor signal transduction is associated with reduced LAT and phospholipase Cgamma1 phosphorylation.

In this study, we investigate the role of the receptor-like protein tyrosine phosphatase CD148 in T-cell activation. Overexpression of CD148 in the Jurkat T-cell line inhibited activation of the transcription factor nuclear factor of activated T cells following T-cell receptor (TCR) stimulation but not following stimulation through a heterologously expressed G protein-coupled receptor, the human muscarinic receptor subtype 1. Using a tetracycline-inducible expression system, we show that the TCR-mediated activation of both the Ras and calcium pathways was inhibited by expression of CD148 at levels that approximate those found in activated primary T cells. These effects were dependent on the phosphatase activity of CD148. Analysis of TCR-induced protein tyrosine phosphorylation demonstrated that most phosphoproteins were unaffected by CD148 expression. However, phospholipase Cgamma1 (PLCgamma1) and LAT were strikingly hypophosphorylated in CD148-expressing cells following TCR stimulation, whereas the phosphorylation levels of Slp-76 and Itk were modestly reduced. Based on these results, we propose that CD148 negatively regulates TCR signaling by interfering with the phosphorylation and function of PLCgamma1 and LAT.

Nitric oxide activates the sarcolemmal K(ATP) channel in normoxic and chronically hypoxic hearts by a cyclic GMP-dependent mechanism.

Chronic myocardial hypoxia results in elevated nitric oxide (NO) production and increased current through the sarcolemmal K(ATP) channel. We hypothesized these two processes are related and determined whether NO alters the electrophysiology of Purkinje fibers obtained from rabbits (n=12/group) raised in a normoxic (F(I)O2=0.21) or hypoxic (F(I)O2=0.12) environment from birth to 9 days of age. Action potential duration (APD)(90) was shorter (112+/-3 ms v 126+/-3 ms) and maximum diastolic potential (MDP) was more negative (-84+/-2 mV v-80+/-1 mV) in hypoxic hearts compared with normoxic controls. In normoxic hearts the NO donors, S-nitrosoglutathione (GSNO) 50 microM and spermine NONOate (50 microM) shortened APD(90) and increased MDP to levels present in chronically hypoxic hearts. This effect was completely abolished by the K(ATP) channel blocker glibenclamide (3 microM) and by a nitric oxide trap, Carboxy-PTIO (100 microM). The NO carrier glutathione (50 microM) and decomposed spermine NONOate had no effect on APD(90) or MDP. GSNO had no effect in hypoxic hearts; however, when GSNO was combined with glibenclamide APD(90) increased, and MDP decreased to normoxic values. 8-Bromo cGMP (100 microM) shortened APD(90) and increased MDP to levels present in chronically hypoxic hearts. This effect was abolished by glibenclamide. A soluble guanylyl cyclase inhibitor, ODQ (10 microM), had no effect on action potentials in normoxic hearts but in hypoxic hearts resulted in an increase in APD(90) to levels present in normoxic hearts and a decrease in MDP. The effect of ODQ could not be reversed by GSNO. We conclude that NO activates the sarcolemmal K(ATP) channel in normoxic and chronically hypoxic hearts by a cyclic GMP-dependent mechanism.

Ammonia exchange between the atmosphere and the surface waters at two locations in the Chesapeake Bay.

Excess phytoplankton production, which contributes to hypoxic conditions, is nitrogen limited in the Chesapeake Bay during the summer months. Therefore, understanding the flux of ammonia by direct deposition to the biologically active surface layer is critical to understanding the nutrient dynamics of the bay. This paper presents the results of a 2-yr study measuring gaseous ammonia (NH3) and aerosol ammonium (NH4+) in Baltimore and Solomons, MD, from which direct atmospheric loading of total ammonia (Nt = NH3 + NH4+) to the Chesapeake Bay is estimated. Mean atmospheric concentrations of total ammonia for Baltimore and Solomons were 2.7 +/- 1.7 and 1.0 +/- 0.8 microg of N m(-3), respectively. Monte Carlo estimates of gross dry deposition ranged from <100 to 4900 microg of N m(-2) d(-1). However, based upon water quality parameters, Monte Carlo estimates of gross volatilization of NH3 were calculated to range from <100 to 7700 microg of N m(-2) d(-1). The resulting net air-sea exchange flux varied seasonally from a net deposition into the water during the winter to a net volatilizing into the atmosphere during the summer. A total of 60% of the paired air-water samples had flux estimates that were not significantly different than equilibrium at the 90% confidence interval. The gross deposition, gross volatilization, and net air-sea fluxes were greater and more variable in Baltimore relative to the rural site. Atmospheric ammonia concentrations decrease during the winter at the rural site. However, the net exchange is still into the water due to an exponential decrease in [NH3]eq with temperature. These results indicate that the nitrogen-limited Chesapeake Bay can act as a source of ammonia to the local atmosphere.

Metabolism of PCBs by the deepwater sculpin (Myoxocephalus thompsoni).

Methylsulfonyl-PCBs (MeSO2-PCBs) are hydrophobic organic contaminants that bioaccumulate in the environment similar to their parent molecules, PCBs. Previously, MeSO2-PCBs have primarily been identified in tissues of birds, humans, and other mammals. However, evidence now supports formation of these metabolites in deepwater sculpin, Myoxocephalus thompsoni, a benthic forage fish predominant in the Great Lakes. The ability of deepwater sculpin to form MeSO2-PCBs is unprecedented for a freshwater fish species and presents a novel biochemical pathway for organochlorine metabolism. Additionally, this appears to be a unique PCB metabolic pathway resulting in a reduction of as much as 10% in the sculpin PCB burden, which is further transformed into another class of organic contaminants in the Great Lakes ecosystem.

Increased mitochondrial K(ATP) channel activity during chronic myocardial hypoxia: is cardioprotection mediated by improved bioenergetics?

Increased resistance to myocardial ischemia in chronically hypoxic immature rabbit hearts is associated with activation of ATP-sensitive K(+) (K(ATP)) channels. We determined whether chronic hypoxia from birth alters the function of the mitochondrial K(ATP) channel. The K(ATP) channel opener bimakalim (1 micromol/L) increased postischemic recovery of left ventricular developed pressure in isolated normoxic (FIO(2)=0.21) hearts to values (42+/-4% to 67+/-5% ) not different from those of hypoxic controls but did not alter postischemic recovery of developed pressure in isolated chronically hypoxic (FIO(2)=0.12) hearts (69+/-5% to 72+/-5%). Conversely, the K(ATP) channel blockers glibenclamide (1 micromol/L) and 5-hydroxydecanoate (5-HD, 300 micromol/L) attenuated the cardioprotective effect of hypoxia but had no effect on postischemic recovery of function in normoxic hearts. ATP synthesis rates in hypoxic heart mitochondria (3.92+/-0.23 micromol ATP. min(-1). mg mitochondrial protein(-1)) were significantly greater than rates in normoxic hearts (2.95+/-0.08 micromol ATP. min(-1). mg mitochondrial protein(-1)). Bimakalim (1 micromol/L) decreased the rate of ATP synthesis in normoxic heart mitochondria consistent with mitochondrial K(ATP) channel activation and mitochondrial depolarization. The effect of bimakalim on ATP synthesis was antagonized by the K(ATP) channel blockers glibenclamide (1 micromol/L) and 5-HD (300 micromol/L) in normoxic heart mitochondria, whereas glibenclamide and 5-HD alone had no effect. In hypoxic heart mitochondria, the rate of ATP synthesis was not affected by bimakalim but was attenuated by glibenclamide and 5-HD. We conclude that mitochondrial K(ATP) channels are activated in chronically hypoxic rabbit hearts and implicate activation of this channel in the improved mitochondrial bioenergetics and cardioprotection observed.

Thermodynamics and kinetics of a molecular motor ensemble.

If, contrary to conventional models of muscle, it is assumed that molecular forces equilibrate among rather than within molecular motors, an equation of state and an expression for energy output can be obtained for a near-equilibrium, coworking ensemble of molecular motors. These equations predict clear, testable relationships between motor structure, motor biochemistry, and ensemble motor function, and we discuss these relationships in the context of various experimental studies. In this model, net work by molecular motors is performed with the relaxation of a near-equilibrium intermediate step in a motor-catalyzed reaction. The free energy available for work is localized to this step, and the rate at which this free energy is transferred to work is accelerated by the free energy of a motor-catalyzed reaction. This thermodynamic model implicitly deals with a motile cell system as a dynamic network (not a rigid lattice) of molecular motors within which the mechanochemistry of one motor influences and is influenced by the mechanochemistry of other motors in the ensemble.

A thermodynamic muscle model and a chemical basis for A.V. Hill's muscle equation.

Direct measurements of a relationship between force and actin-myosin biochemistry in muscle suggest that molecular forces in active muscle rapidly equilibrate among. not within, individual myosin crossbridges [Baker et al. (1999) Biophys J 77: 2657 2664]. This observation suggests a thermodynamic model of muscle contraction in which muscle, not an individual myosin crossbridge, is treated as a near-equilibrium system. The general approach can be applied to any ensemble of molecular motors that undergo a physicochemical step against a constant external potential. In this paper we apply the model to a simple two-state crossbridge scheme like that proposed by A.F. Huxley (1957) [Prog Biophys 7: 255 317], and we immediately obtain A.V. Hill's muscle equation. We show that this equation accurately describes steady-state muscle mechanics, biochemistry and energetics. This thermodynamic model provides a novel description of force-dependent actin-myosin kinetics in muscle and provides precise chemical expressions for myosin cooperativity, myosinduty ratios, the number of working strokes per ATP hydrolyzed, muscle efficiency. and energy transfer.