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PubMed1970

Divalent cation activation of deoxycholate-solubilized and -inactivated membrane reduced nicotinamide adenine dinucleotide oxidase of Bacillus megaterium KM.

After treating Bacillus megaterium KM membranes with 0.2% sodium deoxycholate, most of the membrane reduced nicotinamide adenine dinucleotide (NADH) oxidase was inactivated, and all of the membrane NADH-2,6 dichlorophenol indophenol oxidoreductase was solubilized. Dilution of the deoxycholate-treated membranes in the presence of divalent cations restored almost all of the original membrane NADH oxidase. The effectiveness of the divalent cation activation decreased in the order Ba(2+) > Ca(2+) > Mg(2+) > Mn(2+). After centrifugation, the deoxycholate-treated membranes at 100,000 x g for 1 hr, all of the NADH oxidase that was activated by a divalent cation was soluble. Cation-activated oxidase, however, was insoluble. The results show that 0.2% deoxycholate at least partially solubilizes the total electron chain from NADH to O(2) in an inactive from which can be reactivated by divalent cations with the formation of active, insoluble NADH oxidase.

Bacillus megaterium↗

PubMed1976

Divalent cations and the action potential of leech Retzius cells.

The effects of Sr, Ba, Mn, La and Co on the action potential of the leech Retzius cell were examined using intracellular recording techniques. A previous paper showed that these cells could fire Ca-dependent action potentials in Na-free solution provided TEA was present (Kleinhaus and Prichard, 1975). Under the same conditions Sr 1.5--20 mM was capable of substituting as a current carrier. Ba 2--25mM added to normal Ringer prolonged the duration and increased the amplitude of the action potential of the Retzius cell, and supported action potentials without requiring TEA in Na-free solutions. The overshoots of the Sr- and Ba-dependent action potentials varied with a slope of 40 mM and 75 mV, respectively per 10-fold change in divalent cation concentration. Mn and La selectively blocked that portion of the action potential resulting from an inward movement of Ca, Sr or Ba without affecting the Na-dependent depolarization. The actions of Ca 1 mM on Sr-dependent action potentials were compatible with reversible competitive antagonism. In conclusion the findings: 1. support the proposition that outward K current must be blocked in order for divalent cations to dominate the Retzius cell's behavior during excitation. 2. characterize the divalent cation conductance channel as pharmacologically distinct from the Na conductance channel in the Retzius cell and similar to those described in several other excitable membranes. 3. suggest that the current carrying divalent cations probably flow through the same channel.

Action Potentials↗

PubMed2006

Uncompetitive inhibition of Xenopus laevis aldehyde dehydrogenase 1A1 by divalent cations.

Aldehyde dehydrogenases (ALDHs) convert aldehydes into their corresponding carboxylic acids. ALDH1A1, also known as ALDH class 1 (ALDH1) or retinaldehyde dehydrogenase (RALDH1), prefers retinal to acetaldehyde as a substrate. To investigate the effects of divalent cations on the dehydrogenase activity of Xenopus laevis ALDH1A1, the formation of acetate and retinoic acid from acetaldehyde and retinal, respectively, was investigated in the presence of Ca2+, Mg2+, Mn2+ or Zn2+. All divalent cations tested inhibited the oxidation of acetaldehyde and retinal by ALDH1A1. When acetaldehyde was used as a substrate, the 50% inhibitory concentrations (IC50) were 10, 24, 35 and 220 microM for Zn2+, Mn2+, Mg2+ and Ca2+, respectively. Kinetic studies of ALDH1A1 dehydrogenase activity in the presence or absence of each cation revealed that the inhibition mode by cations was uncompetitive against acetaldehyde, retinal, and NAD+, and that their inhibitory potencies were greater against acetaldehyde than retinal. It was concluded that the divalent cations inhibited X. laevis ALDH1A1 activity in a substrate-dependent manner by affecting a step of the dehydrogenase reaction that occurred after the formation of the ternary complex of the enzyme, substrate, and coenzyme.

Acetaldehyde↗

PubMed1999

Simultaneous separation of common mono- and divalent cations on silica gel modified with aluminium by non-suppressed ion chromatography with conductimetric detection and nitric acid-15-crown-5 as eluent.

The modification of silica gel with aluminium by a coating method was very effective for the preparation of silica-based stationary phases which acted as a cation exchanger under strongly acidic conditions. However, the separation of common mono- and divalent cations (Li+, Na+, NH4+, K+, Mg2+ and Ca2+) on an aluminium-adsorbing silica (Al-Silica) column was moderate by a conductimetric detection ion chromatography (IC) with strongly acidic eluents. Then, the addition of various crown ethers (12-crown-4, 15-crown-5 and 18-crown-6) in acidic eluent was carried out. As a result, it was found that 15-crown-5 was most effective for the improvement of peak resolution. Excellent separation of these cations was achieved in 20 min by elution with 2 mM nitric acid-2 mM 15-crown-5. The proposed IC was successfully applied to the determination of major cations in various natural waters.

Aluminum↗

PubMed2002

Effect of divalent cations and cytosine protonation on thermodynamic properties of intermolecular DNA double and triple helices.

The contribution of divalent cations and cytosine protonation to conformation and stability of duplex and triplex formation were intensively investigated and characterized by ultraviolet (UV), circular dichroism (CD), differential scanning calorimetry (DSC), and electrophoresis mobility shift assay (EMSA). CD spectra showed that the divalent cations investigated would not significantly distort nucleotide geometry, while UV and DSC melting experiments revealed that the cation binding abilities to duplexes and triplexes were clearly dependent on the types of cations under near physiological conditions. The calorimetric enthalpies were generally underestimated relative to the corresponding van't Hoff enthalpies for Hoogsteen and Watson-Crick transitions, but free energy changes derived from the DSC measurements were in good agreement with those derived from the UV measurements. The adjacent placing of the C(+) x G.C triplets in triplexes lowered the stabilities of not only Hoogsteen base-pairing but also Watson-Crick base-pairing. The protonation contribution of the given cytosine residues might depend on the local and global structure of the protonated cytosine complex. A rigid structural targeted-strand would favor the protonation of cytosine residues. The apparent pK(a) values for parallel duplex and triplex investigated were determined to be 6.4 and 7.6, respectively, which are considerably heightened by 2.1 and 3.3 pH unit as compared to the intrinsic pK(a) value of the free cytosine residues.

Calorimetry, Differential Scanning↗

PubMed1989

Regional distribution of somatostatin receptor affinity states in rat brain: effects of divalent cations and GTP.

In the present work, we have characterized by film radioautography the effects of divalent cations and guanine nucleotide on specific receptor for somatostatin (SRIF) using 125I-TyrO-DTrp8-SRIF14 (125I-ToD8-SRIF) as a ligand. The experiments were performed on coronal 20-microM-thick sections cut at the level of the amygdala, thus allowing to study binding sites in several regions enriched in binding sites (frontal cortex, hippocampus CA1 and dentate gyrus, habenula, basolateral nucleus of the amygdala). In a preliminary set of experiments using brain cortical membranes it was found that 3 mM Mg2+ ions doubled the specific binding of 125I-ToD8-SRIF. However, Mg2+ enhanced equally by a factor of 3 affinities of high- and low-affinity binding sites as evidenced by SMS 201.995 displacement curves without modifying the ratio between high and low affinity sites. In radioautographic studies while SRIF14 and SRIF28 elicited monophasic displacement curves, SMS 201.995 displaced 125I-ToD8-SRIF binding in a biphasic manner in all regions tested but the baso-lateral nucleus of the amygdala. Radioautographic distribution of 125I-ToD8-SRIF binding sites was identical whether the sections were incubated with MgCl2 or with MnCl2 and almost undetectable in the absence of ions. In all structures investigated increasing concentrations of GTP totally inhibited 125I-ToD8-SRIF binding with an IC50 value of 3 microM. In conclusion, our results demonstrate that 125I-ToD8-SRIF-binding sites in brain occur on two different affinity states as assessed by a displacement curve using endogenous ligands and SMS 201.995. According to the comparable effects of divalent cations and GTP, the two subtypes of 125I-ToD8-SRIF-binding sites discriminated by SMS 201.995 are likely to correspond to interconvertible forms of the same receptor coupled to a G protein-transducing system.

Animals↗

PubMed1982

The effects of monovalent and divalent cations on the alpha-adrenoceptor of intact human platelets.

1 We have examined the effects of monovalent and divalent cations and purine nucleotides on the binding of agonists and antagonists to the alpha-adrenoceptor of intact human platelets. 2 Replacement of Na+ (150mM) by NH4+ (150mM) in the incubation medium significantly reduced the binding affinity of [3H]-dihydroergocryptine (P less than 0.05) but did not alter the binding capacity. The competitive binding affinity of adrenaline and noradrenaline was unaltered. 3 The addition of Ca2+ (1mM) or Mg2+ (1mM) to the platelet suspension significantly reduced the platelet alpha-adrenoceptor capacity as indicated by either [3H]-dihydroergocryptine (P less than 0.05) or [3H]-yohimbine (P less than 0.01; Ca2+ only). 4 The addition of Ca2+ (1mM) or Mg2+ (1mM) had no effect on the binding affinity of [3H]-dihydroergocryptine but significantly reduced that of [3H]-yohimbine (P less than 0.05). The competitive affinity of adrenaline and noradrenaline determined by inhibition of [3H]-dihydroergocryptine binding, was unchanged in the presence of either cation. 5 Addition of the purine nucleotides ADP, ATP, GDP or GTP (final concentration 10 microM), either alone or in the presence of 1 mM Ca2+ or 1 mM Mg2+, had no effect on the binding of [3H]-dihydroergocryptine or on the competitive affinity of adrenaline or noradrenaline. 6 We conclude that the alpha-adrenoceptor of intact human platelet displays the binding characteristics of the alpha 2L form of the receptor previously identified in the platelet lysate preparation.

Adult↗

PubMed1988

Photolabile chelators for the rapid photorelease of divalent cations.

The properties of a recently synthesized photolabile chelator for divalent cations are described, the affinity of which for Ca2+ changes by some 5 orders of magnitude on illumination. The compound 1-(2-nitro-4,5-dimethoxyphenyl)-N,N,N',N'-tetrakis[(oxycarbonyl)me thyl]-1,2-ethanediamine (DM-nitrophen) binds Ca2+ (Kd approximately 5.0 x 10(-9) M) and Mg2+ (Kd approximately 2.5 x 10(-6) M) with relatively high affinities. On exposure of the DM-nitrophen-Ca2+ complex to UV light in the 350-nm range, the chelator is cleaved yielding iminodiacetic products with a much lower affinity for Ca (Kd approximately 3 x 10(-3) M) and the free [Ca2+] increases. The quantum yield for Ca2+ release is 0.18. In experiments with chemically skinned skeletal muscle fibers, a fully relaxed fiber equilibrated with DM-nitrophen-Ca2+ complex produced maximal contraction after a single flash from a frequency-doubled ruby laser (347 nm). Half-maximal tension was achieved in approximately 40 ms, some 5 times faster than that obtained after a rapid solution change from a Ca2+-free to a Ca2+-containing solution. In experiments with resealed human erythrocyte ghosts, irradiation of ghosts containing the DM-nitrophen-Ca2+ complex activates a Ca2+-dependent K+ efflux pathway, which is not observed in the absence of illumination. DM-nitrophen is sufficiently stable and photolabile to be used as a caged Ca (or caged Mg) for the rapid photoinitiation of divalent cation-dependent processes over a wide concentration range with a significant increase in temporal resolution over conventional mixing methods.

Acetates↗

PubMed1999

Antibacterial activities of new synthetic divalent cation chelators.

A series of new synthetic ligand compounds which chelate divalent cations was examined for the antibacterial activities of the compounds. Only 2 of 14 synthetic chelators, 9-trans-anthryl-1, 4, 8, 11-tetraaza-tetradecane (No. 6) and bis(2-pyridyl)methylamine (No. 13) showed antibacterial activities, whereas none of the diamines, hydrophilic triamines nor tetramines showed antibacterial activities. Chelators No. 6 and No. 13 inhibited the growth of both Gram-negative and -positive bacteria at doses of 25-200 microg/ml, comparable to those of common antibiotics such as polymixin B, fosfomycin and macrolides. Ethylenediaminetetraacetate (EDTA) potentiated these antibacterial activities, whereas an inhibitory effect of Mg2+ on the MICs of these chelators was observed. Moreover, these chelators enhanced the leakage of periplasmic beta-lactamase. It is therefore suggested that chelators No. 6 and No. 13 disrupt both the membranes and cytoplasmic functions of bacteria, resulting in cell death.

Anthracenes↗

PubMed1982

Divalent cation modulation of fibronectin binding to heparin and to DNA.

Fibronectin is an adhesive glycoprotein that binds to heparin and to DNA. The binding of tryptic fragments of human plasma fibronectin to these ligands is found to be highly dependent on the concentration of divalent cations. We have identified 3 types of binding to heparin. 1) Calcium-sensitive binding is inhibited by CaCl2, but not by MgCl2 or by MnCl2. The NH2-terminal 31,000-dalton fragment (fragment 23) has this type of binding, which is half-maximally inhibited by 3 to 4 mM CaCl2. 2) Divalent cation-sensitive binding is exhibited by a 75,000-dalton fragment (fragment 13); its binding is inhibited by all 3 divalent cations. 3) Divalent cation-insensitive binding is characteristic of a 95,000-dalton fragment (fragment 10) and larger fragments. These 3 fragments (fragments 10, 13, and 23) are not disulfide-bonded to other fragments. Specific tryptic fragments of fibronectin also bind readily to native DNA in the presence of EDTA, but the binding of all fragments is abolished by the presence of 10 mM CaCl2 or MgCl2. Our results indicate that the binding of specific domains of fibronectin to heparin or to DNA can be modulated by divalent cations.

Calcium↗

PubMed1965

EFFECT OF ETHAMBUTOL ON NUCLEIC ACID METABOLISM IN MYCOBACTERIUM SMEGMATIS AND ITS REVERSAL BY POLYAMINES AND DIVALENT CATIONS.

Forbes, M. (Lederle Laboratories Division, Pearl River, N.Y.), N. A. Kuck, and E. A. Peets. Effect of ethambutol on nucleic acid metabolism in Mycobacterium smegmatis and its reversal by polyamines and divalent cations. J. Bacteriol. 89:1299-1305. 1965.-Mycobacterium smegmatis, harvested from cultures inhibited by ethambutol and then suspended in drug-free medium, exhibited a prolonged lag before growth resumed. Polyamines and magnesium ions shortened this lag. Polyamines and magnesium added to the culture increased the minimal inhibitory concentration of the drug and reversed the inhibitory effect of the drug, even when added after the drug had already inhibited growth. When ethambutol was added to a culture in its exponential phase of growth, synthesis of protein and deoxyribonucleic acid (DNA), as measured by incorporation of S(35) and P(32), continued for 3 hr at a rate slightly less than in the control cells and then essentially ceased. Synthesis of ribonucleic acid (RNA) was depressed, but it proceeded even after protein synthesis had ceased. Even though the synthesis of RNA continued, the net RNA decreased, and inhibited cells became deficient in RNA. Polyamines and divalent cations, which reverse the inhibitory effect of the drug, have been reported to be involved in nucleic acid turnover. These considerations suggested that ethambutol may exert its inhibitory effect by interfering with a function of cellular polyamines and divalent cations in RNA metabolism.

Amines↗

PubMed2002

Rapid and reversible block of N-type calcium channels (CaV 2.2) by omega-conotoxin GVIA in the absence of divalent cations.

Omega-conotoxin GVIA (omegaCGVIA) has been reported to be an irreversible blocker of N-type calcium channels (Ca(V) 2.2). However, recent studies have demonstrated that the omegaCGVIA off-rate is correlated with divalent cation concentration, because increasing [Ba2+]o accelerated the recovery from omegaCGVIA block. This predicts that the dissociation of omegaCGVIA from N-channels will be negligible in the absence of divalent cations. Surprisingly, we find that omegaCGVIA block is rapidly reversible in divalent cation-free (0 Ba2+) external solutions in which current was carried by MA+. The recovery followed a single-exponential time course with tau = 31 sec. Isochronic measurements showed that, at 2 min after the removal of toxin, current returned to 86% of control in 0 Ba2+ compared with 19% in 3 mm Ba2+. The off-rate of omegaCGVIA from N-channels was dependent on [Ba2+]o, because, at an intermediate concentration (3 microm Ba2+), N-current recovered with tau = 64 sec, significantly slower than that in 0 Ba2+ but faster than in 3 mm Ba2+. Recovery from omegaCGVIA block was also observed when Cs+ or Na+ carried the current in divalent cation-free conditions. The off-rate was sensitive to [Ba2+]o only during washout, because current recovered slowly in the presence of 3 mm Ba2+, even after it was blocked in 0 Ba2+. Assuming that the toxin is a pore blocker, our findings are consistent with a model in which Ba2+ interacts at a site on the extracellular surface of the channel to regulate omegaCGVIA dissociation from N-channels.

Animals↗

PubMed1989

Divalent cations suppress 3',5'-adenosine monophosphate accumulation by stimulating a pertussis toxin-sensitive guanine nucleotide-binding protein in cultured bovine parathyroid cells.

We used pertussis toxin to study the mechanism(s) by which divalent cations lower cellular cAMP content in bovine parathyroid cells. In cultured parathyroid cells, high extracellular Ca2+ (5 mM) or Mg2+ (5-10 mM) lowers dopamine-stimulated cAMP content by 70-90%. Pertussis toxin (0.5 microgram/ml) totally blocks the inhibitory effects of Ca2+ and Mg2+ on cAMP content. Ba2+ and Sr2+ (5 mM) also lower cAMP content by 80-90%, and this effect is, likewise, blocked by pertussis toxin. Pretreatment with pertussis toxin had no effect on the release of cAMP into the extracellular fluid. The toxin also did not modify phosphodiesterase activity in sonicates of parathyroid cells (42.68 +/- 3.26 vs. 47.00 +/- 2.82 pmol cAMP hydrolyzed/10(6) cells.20 min in control and toxin-treated cells, respectively). Moreover, addition of the phosphodiesterase inhibitor isobutyl-methylxanthine did not modify the inhibition of dopamine-stimulated cAMP accumulation by 5 mM Ca2+ in control cells (85% vs. 86% inhibition, respectively, with and without isobutylmethylxanthine). Pertussis toxin-catalyzed ADP ribosylation in homogenates of control cells demonstrated the presence of two substrates with mol wt of 40K and 41K. Preexposure of cells to pertussis toxin overnight resulted in the complete loss of both substrates on subsequent ADP ribosylation with [32P]NAD. Pertussis toxin pretreatment did not enhance adenylate cyclase activity indirectly via reducing the extracellular Ca2+-induced rise in cytosolic Ca2+, since the cytosolic Ca2+ level at 5 mM Ca2+ was about 60% higher in pertussis toxin-treated than in control cells (531 +/- 85 vs. 326 +/- 35 nM; P less than 0.05). In addition, ionomycin had no significant effect on cellular cAMP levels in control cells despite increasing the cytosolic Ca2+ concentration to levels as high as 1700 nM at 10(-5) M. Thus, changes in cytosolic Ca2+ phosphodiesterase activity, or efflux of cAMP from the cell cannot explain the inhibition of cAMP accumulation by divalent cations or the reversal of this effect by pertussis toxin. Instead, the present data suggest that extracellular divalent cations modulate the formation of cellular cAMP in parathyroid cells by a process involving a pertussis toxin-sensitive guanine nucleotide regulatory protein, presumably inhibition of adenylate cyclase by Gi via a receptor-like mechanism.

1-Methyl-3-isobutylxanthine↗

PubMed2004

Synergistic modulation of cystinyl aminopeptidase by divalent cation chelators.

Membranes of Chinese hamster ovary (CHO-K1) cells were used to study the opposite modulation of enzyme activity and [125I]Ang IV binding to cystinyl aminopeptidase (EC 3.4.11.3) by divalent cation chelators. Whereas ethylene diamine tetraacetic acid (EDTA) or ethylene glycol-bis(2-aminoethylether)-N,N,N',N'-tetraacetic acid (EGTA) alone only slightly affected the enzyme activity, 1,10-phenanthrolin (1,10-PHE) produced a complete and concentration-dependent inhibition. Interestingly EDTA (> or =0.05 mM) or EGTA (> or =0.15 mM) enhanced the inhibitory effect of 1,10-PHE. Two-site analysis of the corresponding inhibition curves revealed that EDTA and EGTA converted enzymes with low sensitivity towards 1,10-PHE into enzymes with high sensitivity. The combined inhibition by EDTA (0.1 mM) and 1,10-PHE (0.1 mM) could be prevented and reversed by addition of Zn2+ (at about 0.04-0.1 mM). In contrast, specific binding of [125I]Ang IV was enhanced in the presence of 1,10-PHE. Binding was only slightly affected by EDTA or EGTA alone. Furthermore, the stimulatory effect of 1,10-PHE was potentiated by EDTA (> or =0.05 mM) as well as EGTA (> or =0.15 mM). In the presence of EDTA (0.1 mM) and 1,10-PHE (0.1 mM), specific [125I]Ang IV binding was completely inhibited by Zn2+ (IC50= 39.7 +/- 6.2 microM). The present data show that divalent cations such as Zn2+ are essential for the enzyme activity of cystinyl aminopeptidase and inhibitory for [125I]Ang IV binding. Modulation of the effects of 1,10-PHE by other chelators such as EDTA or EGTA, suggests that, in addition to the binding site for zinc in the catalytic site, cystinyl aminopeptidase also bears a regulatory divalent cation binding site.

Angiotensin II↗

PubMed1996

Effect of divalent cations on the formation and stability of myosin subfragment 1-ADP-phosphate analog complexes.

Myosin belongs to the family of motor proteins. Its interaction with actin coupled with hydrolysis of ATP is the molecular basis of muscle contraction. The head segment of myosin, called subfragment 1 (S1), contains the distinct binding sites for ATP and actin and responsible for the ATPase activity. The rate-limiting step of the ATP hydrolysis is the dissociation of the S1-MgADP-Pi complex which is accelerated by actin. The substitution of Pi with phosphate analogs (PA), such as vanadate (Vi) or beryllium fluoride (BeF(x)), highly stabilizes the complex. We studied the role of the divalent cations in the ATPase activity and in the formation and decomposition of PA-containing stable complexes by substituting Mg2+ with Fe2+, Mn2+, Ni2+, Co2+, and Ca2+. These metal ions supported the actin activation of S1 ATPase and affected the obtained kinetic parameters, Km and V(max). The ATPase activity of S1 in the absence of actin increased with the increasing ionic radius of the metal (Me) ions. These ions also substituted for Mg2+ in the formation of the stable ternary S1-MeADP-PA complexes, which cannot be generated in the absence of divalent cations. Upon formation of stable ternary complexes, S1 reversibly loses its ability to catalyze the hydrolysis of ATP. The formation of the complexes can be followed by monitoring the disappearance of the ATPase activity. The rate of the complex formation depends on the divalent cation present and decreases in the order Mn > Fe > Ni > Co > Mg and Ca > Mn > Fe > Mg > Co in the Vi- and BeF(x)-containing complexes, respectively. The ATPase activity of S1 is recovered upon addition of actin, which causes the decomposition of the complex. The spontaneous decomposition of the complexes was studied in the presence of ethylenediaminetetraacetic acid (EDTA), which chelates the metal divalent cations released from the complex and prevents its reformation. The rate of decomposition was assessed by monitoring the recovery of the ATPase activity of S1 in the presence of EDTA. The rate of decomposition of the Vi- and BeF(x)-containing complexes follows the order Mn > Fe > Co > Mg > Ni and Ca >> Mn > Fe > Co > Mg, respectively. The rate of decomposition increases with the increasing ionic radius of the metal ions, similarly as observed in the case of ionic radius dependence of the ATPase activity. On the basis of this similarity, it is assumed that the decomposition of the complexes consists of two steps, the first step being the very slow release of PA followed by a rapid dissociation of MeADP from S1. The stability of the complexes has been calculated from the formation and decomposition rates. Except in the case of Mg, the stabilities of the BeF(x) complexes are higher than those containing Vi. The results indicate that the metal cations have a significant role in maintaining the proper structure of the transient state complex in the myosin-catalyzed ATP hydrolysis.

Adenosine Diphosphate↗

PubMed1990

Effect of divalent cations on NADH-dependent and NADPH-dependent cytochrome b5 reduction by hepatic microsomes.

The effect of Ca2+ or Mg2+ on cytochrome b5 reduction by porcine liver microsomes was examined using trypsin-solubilized cytochrome b5 as a substrate. The reduction of exogenous cytochrome b5 by microsomes was low at 1.2 microM cytochrome b5 (3.9 or 2.7 nmol/min/mg protein, respectively, with NADH or NADPH). The addition of CaCl2 greatly enhanced either NADH-dependent or NADPH-dependent cytochrome b5 reduction. At 2 mM CaCl2, the reduction rate was increased to 23- or 18-fold of control, respectively with NADH or NADPH. The concentration for half-maximal effect (EC50) was 0.5 or 0.6 mM in the NADH or NADPH systems, respectively. MgCl2 also stimulated cytochrome b5 reduction with a EC50 value of 1.0 mM in the NADH system or 0.6 mM in the NADPH system. The comparison with the result with KCl indicated that the activation by CaCl2 or MgCl2 is caused mainly by their divalent cation moiety. The Km value for cytochrome b5 was decreased and the Vmax was increased by calcium with either the NADH- or the NADPH-dependent system. NADH-ferricyanide reductase activity was not affected by calcium, but NADPH-ferricyanide reductase activity was stimulated as well as NADPH-cytochrome c reductase activity. In the presence of Triton X-100, divalent cations were inhibitory in NADH-dependent cytochrome b5 reduction, and in contrast, stimulative in NADPH-dependent reaction. These findings suggest that the activation of cytochrome b5 reduction by divalent cations in the NADH system is mainly due to an increasing accessibility of the substrate, and in the NADPH system, in addition to this, a direct effect of divalent cations on NADPH-cytochrome P450 reductase is also involved.

Animals↗

PubMed1996

Mercurial-induced alterations in neuronal divalent cation homeostasis.

Mercurials such as Hg2+ and methylmercury (MeHg) are environmental contaminants. Both are neurotoxic upon chronic and acute exposure, however, these toxic manifestations are distinct. The mechanisms underlying this cytotoxicity remain unknown, but may be related to a disruption in divalent cation homeostasis because both disrupt Ca(2+)-dependent processes in several model systems. These effects include a block in nerve-evoked neurotransmitter release as well as an increase in spontaneous transmitter release. This suggests that mercurials simultaneously decrease Ca2+ influx following nerve stimulation, and increase intracellular Ca2+ concentration ([Ca2+]i) in the nerve terminal. Although these effects appear to be at odds, they can be justified mechanistically. Both Hg2+ and MeHg block voltage-activated Ca2+ channels in the nerve terminal. The mechanism of block by these mercurials is different, since Hg2+ and MeHg are competitive and noncompetitive inhibitors of Ca2+ influx, respectively. The functional consequence in both instances remains decreased Ca2+ influx into the nerve terminal following the invasion of an action potential leading to decreased nerve-evoked release of neurotransmitter. The effects of mercurials on voltage-activated Ca2+ channels are distinct from those which mediate the increases in spontaneous transmitter release. Reducing extracellular Ca2+ concentration ([Ca2+]e) decreased, but did not prevent, the mercurial-induced increases in spontaneous transmitter release, suggesting that both intra- and extracellular sources of Ca2+ contribute to mercurial-induced elevations in [Ca2+]i in a nerve terminals. The effects of MeHg on divalent cation homeostasis have been studied using isolated nerve terminals from the rat brain (synaptosomes) and cells in culture (NG108-15 and isolated cerebellar granule cells) loaded with the Ca(2+)-selective fluorescent indicator fura-2. In synaptosomes, MeHg caused an Ca(2+)e-independent elevation in intrasynaptosomal Zn2+ concentration ([Zn2+]i) as well as an Ca(2+)e-dependent elevation in [Ca2+]i. The elevations in [Zn2+]i and [Ca2+]i were mediated by release of Zn2+ from soluble synaptosomal proteins and increased plasma membrane permeability, respectively. In NG108-15 cells, the effects of MeHg on divalent cation concentrations were more complex. First, MeHg mobilized Ca2+ from an intracellular store sensitive to inositol-1,4,5-tris-phosphate (IP3) which was independent of IP3 generation. Second, MeHg increased the intracellular concentration of an endogenous polyvalent cation, possibly Zn2+. Finally, MeHg caused an increase in the plasma membrane permeability to Ca2+ which was attenuated by high concentrations of the voltage-activated Ca2+ channel blocker nifedipine or by the voltage-activated Na+ channel blocker tetrodotoxin (TTX). While these studies demonstrate mercurials interfere with divalent cation regulation in neuronal systems, the consequences of these effects are not yet known.

Animals↗

PubMed2006

Release of colloidal particles in natural porous media by monovalent and divalent cations.

We study mobilization of colloidal particles from natural porous media, such as soils and groundwater aquifers. Extensive laboratory scale column experiments of particle release from four different subsurface materials are presented. The important characteristics of the release process are (i) its non-exponential kinetics, (ii) the finite supply of colloidal particles and (iii) the strong dependence of the release kinetic on the nature of the adsorbed cations. Particle release depends most sensitively on the relative saturation of the medium with divalent cations. We propose a mathematic model, which captures all these aspects quantitatively, and can be used to describe the coupling between transport of major cations and the release of colloidal particles. The present experimental investigations as well as the developed modeling framework represent an important step towards the understanding of colloid-facilitated transport phenomena in natural porous media.

Cations↗