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PubMed1982

Divalent cations and fast axonal transport in chemically desheathed (Triton X-treated) frog sciatic nerve.

We have studied the ability of divalent cations to restore to normal axonal transport (AXT) which was inhibited by deprivation of Ca2+ and/or Mg2+ ions. The epi- and perineurium of the frog sciatic nerve were damaged by a 30-s wash in Triton X-100 containing frog Ringer's. This treatment did not affect either AXT or nerve levels of Ca2+ and Mg2+, but made the ions more easily extractable with a Ca2+- and Mg2+-free Ringer's solution (CMFR). Inhibition of AXT was achieved by incubating Triton X-100-treated nerves in CMFR + EGTA for 5 h, followed by an additional incubation for 12 h in CMFR or Ringer's devoid of only Ca2+ (CFR). These treatments reduced Ca2+ and Mg2+ contents by 77% and 38% respectively. Addition of Ca2+ (1.1 mM) during the 12-h period stimulated AXT, measured as accumulation of 3H-labelled components in front of a ligature, several fold. Mg2+ could not substitute for Ca2+ but potentiated the stimulating effect of Ca2+. Addition of other divalent cations did not affect AXT (Sr2+ and Ba2+) or potentiated the inhibition caused by Ca2+-deprived medium (Mn2+ and Co2+). ATP and creatine phosphate contents were similar in nerves incubated in Ca2+-deprived medium and in Ca2+-containing Ringer's. Thus, inhibition of AXT in the former situation was not due to a decreased availability of high energy phosphates. Two calcium antagonists, D-600 and nifedipin, which are potent smooth muscle relaxants, effectively blocked AXT. The present results suggest that Ca2+ is specifically required to maintain AXT and that an analogy exists between Ca2+ regulation during smooth muscle contraction and AXT.

Animals↗

PubMed2003

A small molecule alpha4beta1/alpha4beta7 antagonist differentiates between the low-affinity states of alpha4beta1 and alpha4beta7: characterization of divalent cation dependence.

An alpha4beta1/alpha4beta7 dual antagonist, 35S-compound 1, was used as a model ligand to study the effect of divalent cations on the activation state and ligand binding properties of alpha4 integrins. In the presence of 1 mM each Ca2+/Mg2+, 35S-compound 1 bound to several cell lines expressing both alpha4beta1 and alpha4beta7, but 2S-[(1-benzenesulfonyl-pyrrolidine-2S-carbonyl)-amino]-4-[4-methyl-2S-(methyl-[2-[4-(3-o-tolyl-ureido)-phenyl]-acetyl]-amino) pentanoylamino]-butyric acid (BIO7662), a specific alpha4beta1 antagonist, completely inhibited 35S-compound 1 binding, suggesting that alpha4beta1 was responsible for the observed binding. 35S-Compound 1 bound RPMI-8866 cells expressing predominantly alpha4beta7 with a KD of 1.9 nM in the presence of 1 mM Mn2+, and binding was inhibited only 29% by BIO7662, suggesting that the probe is a potent antagonist of activated alpha4beta7. With Ca2+/Mg2+, 35S-compound 1 bound Jurkat cells expressing primarily alpha4beta1 with a KD of 18 nM. In contrast, the binding of 35S-compound 1 to Mn2+-activated Jurkat cells occurred slowly, reaching equilibrium by 60 min, and failed to dissociate within another 60 min. The ability of four alpha4beta1/alpha4beta7 antagonists to block binding of activated alpha4beta1 or alpha4beta7 to vascular cell adhesion molecule-1 or mucosal addressin cell adhesion molecule-1, respectively, or to 35S-compound 1 was measured, and a similar rank order of potency was observed for native ligand and probe. Inhibition of 35S-compound 1 binding to alpha4beta1 in Ca2+/Mg2+ was used to identify nonselective antagonists among these four. These studies demonstrate that alpha4beta1 and alpha4beta7 have distinct binding properties for the same ligand, and binding parameters are dependent on the state of integrin activation in response to different divalent cations.

Binding Sites↗

PubMed1992

Potency of depolarization-induced transmitter release is determined by divalent cation influx in PC 12 cells.

Evoked release of [3H]dopamine ([3H]DA) from pheochromocytoma cells (PC 12) is dependent on extracellular calcium ([Ca2+]ex), but it can take place if calcium ions (Ca2+) are substituted by other divalent ions such as strontium (Sr2+) and barium (Ba2+). The potency of the divalent cations at supporting release varies with the cell type; in PC 12 cells the order of potency is Ba2+ > Sr2+ > Ca2+. The close correlation between depolarization-evoked Ca2+ entry and depolarization-evoked transmitter release prompted us to examine whether the higher evoked transmitter release in the presence of Sr2+ correlates with an increased evoked Sr2+ influx. Influx studies were conducted on PC12 cells using a radioactive tracer (45Ca2+ or 85Sr2+, < 1 microM) in the presence of either Sr2+ (0.5 mM) or Ca2+ (0.5 mM). Depolarization with K Cl (60 mM) increased evoked 45Ca2+ influx 2-fold when Ca2+ was substituted with Sr2+. Similarly, evoked 85Sr2+ influx increased 1.87-fold by substituting Ca2+ for Sr2+. Thus the amount of evoked cation influx is determined by the type of divalent ion which is accessible in the extracellular medium, independently of the radioactive tracer used. Increased evoked transmitter release in the presence of Sr2+ was associated with increased evoked Sr2+ influx. This suggests that the potency of evoked transmitter release is determined predominantly by the influx of divalent cations. Furthermore, the steps subsequent to cation influx in the release process are equally efficient for both cations.

Animals↗

PubMed1978

Effect of divalent cations on acetylcholine release from cardiac parasympathetic nerve endings.

The influence of several divalent cations on acetylcholine (ACh) release from vagus nerve terminals was studied in quiescent guinea-pig auricles. ACh release was induced by stimulus trains subthreshold for excitation of atrial cells. The ACh released evoked a hyperpolarization of the atrial cell membrane. Changes in ACh sensitivity of the atrial cell membrane were tested by application of ACh-containing solutions. ACh release increased with the external Ca concentration. External Sr maintained the ACh release in 'Ca-free' media. However, Sr ions were less effective than Ca ions. External Mg acted as a weak inhibitor of the stimulus evoked ACh release, whereas Co and Mn ions exerted a strong inhibition.

Acetylcholine↗

PubMed2006

Removal of some divalent cations from water by membrane-filtration assisted with alginate.

The removal of divalent metal ions from hard waters or galvanic wastewater by polymer-assisted membrane filtration using alginate was investigated. The ability of this natural polymer to form aggregates and gels in presence of metal ions was studied, in order to carry out metal removal by ultra or micro-filtration. Alginate titrations have shown the presence of amine groups in addition to carboxylates onto the polymer backbone. The binding properties of alginate with divalent cations have been studied, showing an increasing affinity for Ca2+ over Mg2+ as polymer concentration increases, and the relative affinity Pb2+ > or = Cu2+ > Zn2+ > Ni2+. The softening of hard natural waters was achieved successfully and easily, but needs an optimal alginate concentration approximately 4 x 10(-2) M. The alginate powder can be directly added to hard waters. Except for Ni2+, metal-removal was efficient. Polymer regeneration has shown that Cu2+-complexes are labiles.

Alginates↗

PubMed2001

Effects of some divalent cations on nitrergic relaxations in the mouse corpus cavernosum.

Acute effects of some divalent cations (Cd2+, Ni2+, Co2+, Zn2+, Mn2+ and Sn2+) were investigated on neurogenic and endothelium-dependent relaxations in the isolated mouse corpus cavernosum. Neither neurogenic nor endothelium-dependent relaxation was affected by cations at the concentrations used (up to 100 microM), except Cd2+. Although Cd2+ (20 and 40 microM) did not cause any significant alteration in the acetylcholine- (ACh) or sodium nitroprusside- (SNP) induced relaxation, it inhibited electrical field stimulation- (EFS) produced relaxation significantly. Zn2+ and selenium could not reverse this inhibitory action. Cd2+ did block the EFS-evoked guanethidine-sensitive contraction in the presence of N(G)-nitro-L-arginine. Elevation of external Ca2+ content significantly reduced the inhibitions due to Cd2+ on the EFS-induced relaxation and on the EFS-evoked guanethidine-sensitive contraction. In the Ca2+-omitted medium, EFS-induced relaxation disappeared, while acetylcholine-elicited relaxation resisted. Verapamil was ineffective on the relaxation produced by EFS or acetylcholine. However, it significantly diminished phenylephrine-induced contractions. These findings suggest that unlike other cations at the concentrations used in the present study, Cd2+ may have an effect on an external Ca2+-dependent mechanism at the neuronal level, and this effect may be responsible for its acute inhibitory action on the neurogenic relaxation in the mouse corpus cavernosum.

Animals↗

PubMed1993

Stabilization of platelet-fibrinogen interactions: modulation by divalent cations.

The binding of fibrinogen to its GPIIb-IIIa receptor is divalent-cation dependent. In addition to Ca+2 and Mg+2, Mn+2 has been shown to modulate adhesive protein interactions with integrins. This study examined the effect of Mn+2 on fibrinogen interactions with intact platelets. Compared with that of control platelets in buffer containing 1 mmol/L Mg+2, fibrinogen binding to adenosine diphosphate- or thrombin-stimulated platelets decreased 23% +/- 12% and 15% +/- 9% (mean +/- SD, n = 4), respectively, after addition of 1 mmol/L Mn+2. No change in binding affinity was noted, but the stability of platelet-fibrinogen interactions was diminished markedly. Ethylenediaminetetraacetic acid dissociated 68% +/- 8% of fibrinogen bound to ADP-treated platelets (p < 0.05) during a 60-minute incubation with fibrinogen and 1 mmol/L Mn+2, compared with 40% +/- 13% of fibrinogen bound to control platelets and 29% +/- 8% of fibrinogen bound in the presence of Ca+2 (mean +/- SD, n = 6). Mn+2 also diminished the stabilization of fibrinogen interaction with thrombin-stimulated platelets and inhibited the recovery of bound fibrinogen with the Triton X-100 (Union Carbide Corp., Danbury, Conn.) insoluble cytoskeleton. Only 31% +/- 10% of fibrinogen bound to thrombin-stimulated platelets for 60 minutes in the presence of Mn+2 associated with the cytoskeleton (p < 0.05), compared with 61% +/- 14% and 75% +/- 20% of fibrinogen bound to control platelets incubated with and without Ca+2, respectively. Mn+2 further inhibited large adenosine diphosphate- or thrombin-induced platelet aggregate formation and reduced the ability of platelets to retract fibrin clots. These data suggest that Mn+2 alters GPIIb-IIIa function relative to native fibrinogen and support a role for the stabilization of platelet-fibrinogen interactions in platelet aggregation and clot retraction.

Adenosine Diphosphate↗

PubMed2002

Determination of total acidity and of divalent cations by ion chromatography with n-hexadecylphosphocholine as the stationary phase.

An ion chromatographic (IC) method is reported for simultaneous determination of total acidity (H+), Ba2+, Ca2+, and Mg2+ in aqueous samples. A standard ODS silica column modified by coating with n-hexadecylphosphocholine was used as the separation column. Water alone was used as the eluent, with conductivity detection of the sample ions. An excess of sodium iodide was added to each sample so that both H+ and divalent cations were always eluted with iodide as the counterion. The elution order was Ba2+, Mg2+, Ca2+, and H+ with H+ being eluted much later than the divalent cations. Acetic acid and several other weak acids could also be separated because all the protons were transposed from acetic acid (HAc) to HI by the sodium iodide. Detection limits for 100 microl injection, S/N=3 were in the low micromolar range for the divalent cations and approximately 0.3 mM for H+/I-. This method was used successfully for simultaneous determination of total acidity, magnesium and calcium in HCl-type of hot-spring water.

Acids↗

PubMed1996

Divalent cations and heparin/heparan sulfate cooperate to control assembly and activity of the fibroblast growth factor receptor complex.

Polypeptides of the fibroblast growth factor (FGF) family are ubiquitous bioregulators within tissues whose activity is controlled by heparan sulfates within the pericellular matrix. FGF and the ectodomain of their transmembrane tyrosine kinase receptors (FGFR) exhibit heparin-binding domains that when juxtaposed in a FGF middle dotFGFR complex can accommodate a single, potentially bivalent, decameric polysaccharide chain in a ternary complex. Here we show that the interaction of heparin with FGF ligands is not affected by divalent cations. In contrast, the high affinity interaction (apparent Kd = 10 nM) of heparin with FGFR requires Ca2+ or Mg2+ at physiological concentrations. Divalent cations maintain FGFR in a heparan sulfate-dependent state in respect to FGF binding and an FGF- and heparan sulfate-dependent state in respect to autophosphorylation. A model is proposed where divalent cations and heparan sulfate cooperate to maintain FGFR in a conformation that restricts trans-phosphorylation between intracellular kinase domains. The restriction is overcome by FGF or constitutively as a common consequence of diverse mutations in FGFR associated with skeletal and craniofacial abnormalities.

Binding Sites↗

PubMed1990

Effects of divalent cations on the frequency of spontaneous action potentials from the lateral line organ of Xenopus laevis.

The effect of superfusion of the internal surface of the skin of Xenopus laevis with saline containing Co2+, Ca2+, Mg2+, or Ba2+, on the frequency of spontaneous action potentials of the lateral line nerve, was studied to investigate the role of extracellular Ca2+ in spontaneous neural activity. Addition of divalent cations to frog saline, either singly or as a mixture of two different ions, produced concentration-dependent suppression of spontaneous rate. The rank order of potency for suppression by each ion, perfused alone, was Co2+ greater than Ca2+ greater than Mg2+ greater than Ba2+. Suppression by combinations of Mg2+ and Co2+, or of Ca2+ and Co2+, was approximated by the sum of the suppressive effects of each cation. Ca2+ was more suppressive than Mg2+ when each of these ions was paired with the same amount of Co2+, while Ca2+ was approximately as suppressive as Co2+ when similarly paired with Mg2+. One interpretation of the suppression by Ca2+ invokes the hypothesis that divalent cations suppress spontaneous activity by charge screening of voltage-sensitive Na+ channels on afferent dendrites and that release of neurotransmitter by the influx of extracellular Ca2+ through voltage-sensitive Ca2+ channels of hair cells may not be the sole mechanism for generation of spontaneous activity in the lateral line. These results quantify the relative suppressive potency of common divalent cations in the lateral line, and serve as a caveat to investigators who interpret a blockade of action potentials by high concentrations of Co2+ or Mg2+ as sufficient evidence for dependence of neurotransmission upon extracellular Ca2+, particularly in acousticolateralis systems.

Action Potentials↗

PubMed1990

Divalent cation binding to reduced and octanoyl acyl-carrier protein.

Mn(II) EPR binding studies with reduced acyl-carrier protein (ACP-SH) strongly suggest the presence of two relatively high-affinity manganese-binding sites (average Kd/site approximately 80 microM) at physiological pH. Lowering the pH or titrating with sodium chloride reduces the average number of bound divalent cations and decreases the binding affinity. This is consistent with the idea that anionic ligand(s), e.g. the carboxylate of glutamic or aspartic acid, on the protein are involved in manganese ion coordination. At pH values above 8.0, binding affinity is also reduced, whereas the average number of bound metal ions increases to about five at pH 8.5. By interacting weakly with divalent cations (average Kd/site approximately 1 mM), octanoyl acyl-carrier protein (OcoACP) exhibits dramatically different metal-ion-binding properties compared to ACP-SH. Calcium and magnesium can compete in either ACP species for manganese binding. Photochemically-induced dynamic nuclear polarisation 1H-NMR experiments strongly suggest that ACP-SH and OcoACP undergo at pH-induced conformational change between pH 5.5 and pH 7.0, and that divalent cations stabilize the protein against such pH-induced structural perturbations.

Acyl Carrier Protein↗

PubMed2006

Removal of divalent cations induces structural transitions in red clover necrotic mosaic virus, revealing a potential mechanism for RNA release.

The structure of Red clover necrotic mosaic virus (RCNMV), an icosahedral plant virus, was resolved to 8.5 A by cryoelectron microscopy. The virion capsid has prominent surface protrusions and subunits with a clearly defined shell and protruding domains. The structures of both the individual capsid protein (CP) subunits and the entire virion capsid are consistent with other species in the Tombusviridae family. Within the RCNMV capsid, there is a clearly defined inner cage formed by complexes of genomic RNA and the amino termini of CP subunits. An RCNMV virion has approximately 390 +/- 30 Ca2+ ions bound to the capsid and 420 +/- 25 Mg2+ ions thought to be in the interior of the capsid. Depletion of both Ca2+ and Mg2+ ions from RCNMV leads to significant structural changes, including (i) formation of 11- to 13-A-diameter channels that extend through the capsid and (ii) significant reorganization within the interior of the capsid. Genomic RNA within native capsids containing both Ca2+ and Mg2+ ions is extremely resistant to nucleases, but depletion of both of these cations results in nuclease sensitivity, as measured by a significant reduction in RCNMV infectivity. These results indicate that divalent cations play a central role in capsid dynamics and suggest a mechanism for the release of viral RNA in low-divalent-cation environments such as those found within the cytoplasm of a cell.

Amino Acid Sequence↗

PubMed1994

Contractile responses of longitudinal muscle strip to 5-HT and influences of divalent cations in the guinea-pig isolated colon.

The contractile effects of 5-hydroxytryptamine (5-HT) and influences of several kinds of divalent cations were investigated on longitudinal muscle strips of the guinea-pig isolated distal colon. 5-HT (10 nM-10 microM) produced phasic contractions which were partially inhibited by atropine (1 microM) and markedly inhibited by tetrodotoxin (1 microM), indicating that 5-HT acts mainly on the myenteric plexus and releases transmitters to cause contraction of the longitudinal muscle. The contractile response to 5-HT (3 microM) was almost completely inhibited by spantide (10 microM), a substance P antagonist, in the presence of atropine (1 microM), while spantide alone did not block 5-HT-induced contraction. Of several divalent cations including Cd2+, Co2+, Mg2+, Mn2+, Ni2+, Sr2+ and Zn2+, Cd2+ ions (10 mu-100 microM), which block L- and N-type Ca2+ channels, were most effective inhibitor of the 5-HT-induced contractions. While Sr2+ and Co2+ at a concentration of 100 microM did not have a significant effect. The order effectiveness of inhibition was Cd2+ >> Mn2+ > Mg2+ = Ni2+ = Zn2+. Bay K 8644 (1 microM), a L-type Ca2+ channel activator, did not influence the contractile response of the longitudinal muscle strip to 5-HT (3 microM). The present results suggest that 5-HT may mainly act on N-type Ca2+ channels in the myenteric neurones and cause the release of at least acetylcholine and substance P to induce contractions of the longitudinal muscle in the guinea-pig distal colon.

Acetylcholine↗

PubMed1996

Divalent cations regulate the organization of integrins alpha v beta 3 and alpha v beta 5 on the cell surface.

Extracellular divalent cations are important regulators of integrin ligand binding activity. In this study we evaluated how divalent cations affect the organization of integrins into focal adhesion sites. Integrins alpha v beta 3 and alpha v beta 5 were compared because they share a high degree of structural homology and because both integrins mediate cell adhesion to vitronectin. On MG-63 osteosarcoma cells, we found that both the extent and pattern of integrin organization was regulated by the type of extracellular divalent ion. Integrin alpha v beta 3 organized in focal contacts when Mn2+ or Mg2+ was present, but not in Ca2+. In contrast, alpha v beta 5 organized in focal contacts only when Ca2+ or Mg2+ was present. Integrin alpha v beta 5 clustered in a centrally located punctate field on the ventral surface of the cell in the presence of Mn2+. These observations reveal a previously unappreciated role for divalent ions in regulating the organization of integrins into focal adhesion sites.

Antigens, CD↗

PubMed1982

Alterations in spontaneous transmitter release by divalent cations after treatment of the neuromuscular junction with beta-bungarotoxin.

1. Spontaneous transmitter release was studied at the frog sartorius neuromuscular junction in the presence of a variety of cations before and after treatment with the specific presynaptic neurotoxin, beta-bungarotoxin (beta-BuTX). 2. Treatment with beta-BuTX produced a maintained increase in spontaneous release, as indicated by the miniature end-plate potential (m.e.p.p.) frequency. It was demonstrated that the m.e.p.p. frequency remained dependent on the extracellular calcium concentration. 3. A 30 mM increase in extracellular sodium chloride produced a reversible increase in frequency only after beta-BuTX treatment, indicating that beta-BuTX had increased the permeability of the presynaptic terminal. 4. Furthermore, several divalent cations other than calcium were shown to either maintain or greatly increase the m.e.p.p. frequency after beta-BuTX treatment (before toxin treatment replacement of calcium by these divalent cations produced only small changes in frequency). The relative effectiveness of the divalent cations tested in increasing spontaneous transmitter release after toxin treatment was Co2+ congruent to Ni2+ greater than Mg2+ greater than Ca2+ congruent to Sr2+ greater than Mn2+. The effect of cobalt, which increased the m.e.p.p. frequency 6.5 times after toxin treatment, was studied in detail.

Acetylcholine↗

PubMed1985

Sickle erythrocyte adherence to vascular endothelium. Morphologic correlates and the requirement for divalent cations and collagen-binding plasma proteins.

Increased adherence of sickle erythrocytes to vascular endothelium has been suggested by Hebbel and his colleagues to play a role in vasocclusive events of sickle cell disease. To define the role of cell membrane changes and plasma factors in cell adherence, a micropipette technique previously developed by us to obtain a direct, quantitative measure of cell adherence was used to evaluate the adhesivity of different morphologic classes of sickle cells to endothelial cells in various suspending media. Irregularly shaped, deformable sickle cells were four- to fivefold more adherent than discoid sickle cells, whereas rigid irreversibly sickled cells were least adherent. Sickle erythrocytes adhered to endothelial cells when suspended in autologous citrated or heparinized plasma but were totally nonadherent when suspended in autologous EDTA plasma. Removal of the divalent cation chelator and addition of calcium to EDTA plasma restored its ability to promote adhesion, implying an absolute requirement for divalent cations in sickle cell adherence. Sickle cells also did not adhere to endothelial cells in protein-free media containing divalent cations, suggesting an additional requirement for plasma proteins. Removal of collagen-binding proteins from citrated sickle plasma resulted in a three- to fivefold reduction in its ability to promote cell adhesion, suggesting an important role for these plasma proteins in sickle cell adherence. The results of this study imply that sickle cell adherence to vascular endothelium is a complex process in which temporal changes in the numbers of cells identified to be most adhesive and the plasma concentration of protein(s) involved in the adhesive process determine the extent of in vivo sickle cell adherence.

Anemia, Sickle Cell↗

PubMed1987

The phosphate-pyrophosphate exchange and hydrolytic reactions of the membrane-bound pyrophosphatase of Rhodospirillum rubrum: effects of pH and divalent cations.

The relation that exist between the Pi-PPi exchange reaction and pyrophosphate hydrolysis by the membrane-bound pyrophosphatase of chromatophores of Rhodospirillum rubrum was studied. The two reactions have a markedly different requirement for pH. The optimal pH for hydrolysis was 6.5 Mg2+ or Pi for the enzyme; Mn2+ and Co2+ support the Pi-PPi exchange reaction partially (50%), but the reaction is slower than with Mg2+; other divalent cations like Zn2+ or Ca2+ do not support the exchange reaction. In the hydrolytic reaction, Zn2+, at low concentration, substitutes for Mg2+ as substrate, and Co2+ also substitutes in limited amount (50%). Other cations (Ca2+, Cu2+, Fe2+, etc.) do not act as substrates in complex with PPi. The Zn2+ at high concentrations inhibited the hydrolytic reaction, probably due to uncomplexed free Zn2+. In the presence of high concentration of substrate for the hydrolysis (Mg-PPi) the divalent cations are inhibitory in the following order: Zn2+ greater than Mn2+ greater than Ca2+ greater than or equal to Co2+ greater than Fe2+ greater than Cu2+ greater than Mg2+. The data in this work suggest that H+ and divalent cations in their free form induced changes in the kinetic properties of the enzyme.

Cations, Divalent↗

PubMed2004

Divalent cations regulate acidity within the lumen and tubulovesicle compartment of gastric parietal cells.

BACKGROUND & AIMS: Until recently, it has not been possible to evaluate factors that regulate the acidity of the microenvironment within the tubulovesicles and luminal (TV/L) spaces of the gastric gland. The goal of this study was to develop a method for monitoring the mechanisms that regulate acidity in the TV/L compartment. METHODS: Isolated rabbit gastric glands (intact or permeabilized with S. aureus alpha-toxin) were loaded with a recently characterized fluorescent dye, LysoSensor Yellow-Blue DND 160 (Molecular Probes, Eugene, OR), which localizes to highly acidic compartments and can be used to monitor acidity ratiometrically. RESULTS: In resting glands, the pH of the TV/L compartment was approximately 3.4. Moderate alkalizations ( approximately 0.5 to 1.0 pH unit alkalization) were observed during exposure to inhibitors of the apical H(+)/K(+) ATPase (omeprazole and SCH28080), thereby unmasking a stable, low-level leak of H(+) ions from the TV/L compartment. Similar changes were observed in alpha-toxin permeabilized glands following depletion of ATP in the cytoplasm. In intact and permeabilized glands, we used the cell-permeant, divalent cation chelator, tetrakis-(2-pyridylmethyl)ethylenediamine (TPEN) to probe the effects of lowering divalent cation content of the TV/L compartment. Exposure to relatively low concentrations (20 micromol/L, 50 micromol/L) of TPEN reversibly promoted H(+) leakage. At these concentrations, simultaneous inhibition using SCH28080 led to marked enhancement of the rate of alkalization. CONCLUSIONS: The effects of low-dose TPEN suggests that acidity within the TV/L compartment of the gastric gland may be regulated, at least in part, by its content of divalent cations such as Zn(2+), for which TPEN has high affinity.

Animals↗