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PubMed2004

Block by extracellular divalent cations of Drosophila big brain channels expressed in Xenopus oocytes.

Drosophila Big Brain (BIB) is a transmembrane protein encoded by the neurogenic gene big brain (bib), which is important for early development of the fly nervous system. BIB expressed in Xenopus oocytes is a monovalent cation channel modulated by tyrosine kinase signaling. Results here demonstrate that the BIB conductance shows voltage- and dose-dependent block by extracellular divalent cations Ca(2+) and Ba(2+) but not by Mg(2+) in wild-type channels. Site-directed mutagenesis of negatively charged glutamate (Glu(274)) and aspartate (Asp(253)) residues had no effect on divalent cation block. However, mutation of a conserved glutamate at position 71 (Glu(71)) in the first transmembrane domain (M1) altered channel properties. Mutation of Glu(71) to Asp introduced a new sensitivity to block by extracellular Mg(2+); substitutions with asparagine or glutamine decreased whole-cell conductance; and substitution with lysine compromised plasma membrane expression. Block by divalent cations is important in other ion channels for voltage-dependent function, enhanced signal resolution, and feedback regulation. Our data show that the wild-type BIB conductance is attenuated by external Ca(2+), suggesting that endogenous divalent cation block might be relevant for enhancing signal resolution or voltage dependence for the native signaling process in neuronal cell fate determination.

Animals↗

PubMed1980

Regulation of macrophage lysosomal enzyme secretion: role of arachidonate metabolites, divalent cations and cyclic AMP.

We have investigated the role in macrophage lysosomal enzyme release of arachidonate metabolites, extracellular divalent cations and cyclic AMP (cAMP) which modulate secretion in other cell types. Lysosomal enzyme secretion induced by zymosan was accompanied by release of malondialdehyde (MDA), which is derived from arachidonic acid via prostaglandin synthase. Blockade of MDA formation, by aspirin or indomethacin, was associated with only a small inhibitory effect on lysosomal enzyme release by zymosan: arachidonate metabolites thus play only a minor role in mediating macrophage lysosomal enzyme release. Zymosan-induced secretion of lysosomal enzymes from macrophages did not require extracellular magnesium or calcium although release was enhanced by magnesium and inhibited by calcium. These effects may be related to an influence of the ions on phagocytosis. Elevation of intracellular divalent cation concentrations, by ionophore A23187, induced release of lysosomal enzymes but this was a result of cell lysis. Adenylate cyclase stimulants and dibutyryl cAMP produced slight inhibition of zymosan-induced lysosomal enzyme release. Aminophylline and papaverine caused more marked inhibition but their effects may be due to actions independent of phosphodiesterase inhibition. Our data indicate that arachidonate metabolites and cAMP do not play a major role in regulating zymosan-induced enzyme release from macrophages. Extracellular calcium and magnesium may modulate secretion but the role of intracellular divalent cations remains to be established. We conclude that macrophage lysosomal enzyme secretion is controlled by regulatory mechanisms different from those which control similar degranulation processes in other cell types.

Animals↗

PubMed1999

Control of enzymatic degradation of hyaluronan by divalent cations.

Enzymatic degradation of hyaluronan (HA) by testicular hyaluronidase (HAase, hyaluronate 4-glucanohydrolase) requires inclusion of mono- or divalent cations in the reaction mixture. Most divalent cations activated HAase with equal potency; however, Cu2+ suppressed degradation, and Ca2+ showed a concentration-dependent regulation of size of the oligosaccharide products. Careful selection of HAase assay parameters is critical for discovery of novel HAase inhibitors and for preparation of controlled-size oligosaccharide fragments.

Calcium↗

PubMed2002

Hydrodynamic properties and quaternary structure of the 90 kDa heat-shock protein: effects of divalent cations.

The 90 kDa heat-shock protein (Hsp90) is one of the major stress proteins whose overall structure remains unknown. In this study, we investigated the influence of divalent cations Mg(2+) and Ca(2+) on the hydrodynamic properties and quaternary structure of Hsp90. Using analytical ultracentrifugation, size-exclusion chromatography, and polyacrylamide gel electrophoresis, we showed that native Hsp90 was mostly dimeric. The Hsp90 dimer had a sedimentation coefficient, s(w,20) degrees, of 6.10 +/- 0.03 S, which slightly deviated from the hydrodynamics of a globular protein. Using chemical cross-linking and analytical ultracentrifugation, we showed that Mg(2+) and Ca(2+) induced a tertiary conformational change of Hsp90, leading to a self-association process. In the presence of divalent cations, Hsp90 existed as a mixture of monomers, dimers, and tetramers at equilibrium. Finally, to identify Hsp90 domains involved in this divalent cation-dependent self-association, we studied the oligomerization state of the N-terminal (positions 1-221) of Hsp90, the influence of an N-terminal specific ligand, geldanamycin (GA), and the effect of C-terminal truncation on the ability of Hsp90 to oligomerize in the presence of divalent cations. We previously showed that GA inhibits Hsp90 heat-induced oligomerization [Garnier, C., Protasevich, I., Gilli, R., Tsvetkov, P., Lobachov, V., Peyrot, V., Briand, C., and Makarov, A. (1998) Biochem. Biophys. Res. Commun. 249, 197-201], but now we observed that GA does not influence divalent cation-dependent oligomerization of Hsp90, suggesting another mechanism. This mechanism involved the C-terminal part of the protein since C-terminally truncated Hsp90 did not oligomerize in the presence of divalent cations.

Animals↗

PubMed1993

Divalent cations block the cyclic nucleotide-gated channel of olfactory receptor neurons.

1. The effects of external divalent cations on odor-dependent, cyclic AMP-activated single-channel currents from olfactory receptor neurons of the tiger salamander (Ambystoma tigrinum) were studied in inside-out membrane patches taken from dendritic regions of freshly isolated sensory cells. 2. Channels were reversibly activated by 100 microM cyclic AMP. In the absence of divalent cations, the channel had a linear current-voltage relation giving a conductance of 45 pS. With increasing concentrations of either Ca2+ or Mg2+ in the external solution, the channel displayed a rapid flickering behavior. At higher concentrations of divalent cations, the transitions were too rapid to be fully resolved and appeared as a reduction in mean unitary single-channel current amplitude. 3. This effect was voltage dependent, and on analysis was shown to be due to an open channel block by divalent ions. In the case of Mg2+, the block increased steadily with hyperpolarization. In contrast, for Ca2+ the block first increased with hyperpolarization and then decreased with further hyperpolarization beyond -70 mV, providing evidence for Ca2+ permeation of this channel. 4. This block is similar to that seen in voltage-gated calcium channels. Additionally, the cyclic nucleotide-gated channel shows some pharmacological similarities with L-type calcium channels, including a novel block of the cyclic nucleotide channel by nifedipine (50 microM). 5. Our results indicate that the sensory generator current simultaneously depends on the presence of the second messenger and on the membrane potential of the olfactory neuron.

Ambystoma↗

PubMed1989

Effects of divalent cations and saccharides on Vibrio metschnikovii cytolysin-induced hemolysis of rabbit erythrocytes.

Divalent cations and polysaccharides such as inulin and dextran reversibly inhibited hemolysis of rabbit erythrocytes caused by Vibrio metschnikovii cytolysin. On the basis of the 50% inhibitory doses, the cations were divided into two groups, group I (Cd2+, Cu2+, Ni2+, Sn2+, and Zn2+) and group II (Ba2+, Ca2+, Co2+, Mg2+, Mn2+, and Sr2+). Neither divalent cations nor polysaccharides interfered with the binding of toxins to the erythrocyte membrane. Group I cations disturbed tetramer formation of cytolysin on the cytolysin-lysed erythrocyte membrane, although group II cations and dextran did not affect the process. Erythrocytes treated with cytolysin in the presence of group II cations or dextran lysed after transfer to toxin- and inhibitor (group II cations or dextran)-free buffer at both 37 and 4 degrees C. However, erythrocytes treated in the presence of group I cations lysed at 37 degrees C but not at 4 degrees C, indicating that group I cations block the temperature-dependent lesion (tetramer)-forming step subsequent to the binding of cytolysin to the erythrocytes. The cytolysin-treated erythrocytes swelled in a colloid osmotic manner, and the swelling was preceded by the binding and the lesion-forming steps. It is also suggested that the lysis of the erythrocytes proceeds in a temperature-independent manner and that the cytolysin does not bind to the erythrocytes at 4 degrees C. These findings suggest that the sequence of V. metschnikovii cytolysin-induced hemolysis is defined by three steps: (i) a temperature-dependent binding step, (ii) a temperature-dependent lesion-forming step, and (iii) a temperature-independent lysis step.

Animals↗

PubMed1991

Effect of external Cd2+ and other divalent cations on carbachol-activated non-selective cation channels in guinea-pig ileum.

1. Inhibitory actions of the external divalent cations, Cd2+, Ni2+, Mn2+ and Co2+ on carbachol (CCh)-induced inward current (Ins,ACh) were investigated in caesium aspartate-loaded single longitudinal smooth muscle cells of guinea-pig ileum, using a rapid solution-switching device, under voltage-clamped conditions. 2. Cd2+ (20-1000 microM), when added to the external solution, reduced Ins,ACh rapidly and reversibly. This effect occurred dose dependently, the relationship being adequately described by a Michaelis-Menten equation with a Hill coefficient (n) of 1.0 and a dissociation constant (Kd) of 98 microM. 3. The inhibitory action of Cd2+ was associated neither with agonist concentration (CCh) nor with changes in reversal potential, and was voltage independent. 4. The appearance and removal of the effect of Cd2+ were both rapid (a few hundred milliseconds), in sharp contrast with a relatively slow time course of atropine or CCh action (of the order of seconds). 5. Other divalent cations (Ni2+, Mn2+, Co2+, Mg2+) applied externally also suppressed Ins,ACh, but less potently. The sequence of apparent potency was Cd2+ (98 microM) greater than or equal to Ni2+ (131 microM) much greater than Co2+ (700 microM) greater than or equal to Mn2+ (1000 microM) much greater than Mg2+ (approximately 10 mM). 6. External Ca2+ increased Ins,ACh dose dependently and antagonized the inhibitory effect of Cd2+. However, this effect may not be a simple competition with Cd2+. 7. These results show that external divalent cations strongly modulate Ins,ACh channels, possibly through direct interaction with the channel protein.

Animals↗

PubMed1998

Characterization of the monovalent and divalent cation requirements for the xenobiotic carboxylic acid: CoA ligases of bovine liver mitochondria.

The XL-I, XL-II and XL-III forms of xenobiotic/medium-chain fatty acid: CoA ligase were found to be inactive toward benzoate in the absence of either monovalent or divalent cations. The absolute requirement for monovalent cation was satisfied by either K+, Rb+, or NH4+. Na+ only supported a very low rate. Varying the nature of the anion had only a minor effect. For XL-I and XI-II, the optimum concentration of K+ was 50 mM; higher (physiologic) concentrations led to a decrease in activity. K+ did not inhibit XL-III. The absolute requirement for divalent cation was satisfied by Mg2+ or Mn2+, or to a lesser extent by Co2+ or Fe2+. For the XL-I and XL-II, excess uncomplexed Mg2+ or Mn2+ decreased the rate; the optimum concentration of Mn2+ was approximately the same as the concentration of ATP in the assay, and the optimum concentration of Mg2+ was approximately double the concentration of ATP in the assay. This is consistent with the concept that the divalent cation is required to complex with ATP and with the known stability constants for the ATP complexes of these two divalent cations. XL-III was not inhibited by uncomplexed divalent cations. Uncomplexed ATP was a moderate inhibitor of XL-I and XL-II, and a weak inhibitor of XL-III. The data indicate that in vivo benzoate conjugation is K+ and Mg2+ dependent, and that the cation effects are complex and differ for XL-I and XL-II as compared with XL-III.

Adenosine Triphosphate↗

PubMed1975

The role of calcium and cyclic nucleotides in alpha-amylase release from slices of rat parotid: studies with the divalent cation ionophore A-23187.

The divalent cation ionophore A-23187 caused a Ca-2+-dependent increase in alpha-amylase release from slices of rat parotid gland. The effect of A-23187 on alpha-amylase release was not caused by release of endogenous agonists since l-propranolol, phentolamine, and atropine had no effect. The magnitude of alpha-amylase release caused by A-23187 was small compared to the effect of isoproterenol. In this respect it more closely resembles the action of cholinergic and alpha-adrenergic agonists on alpha-amylase release. A-23187 inhibited the increase in the level of parotid adenosine 3,5'-monophosphate caused by isoprotrenol. The inhibitory effect required incubation of the slices with the ionophore before the addition of isoproterenol. The ionophore also caused a Ca-2+-dependent increase in the level of guanosine 3',5'-monophosphate (cyclic GMP). Theophylline enhanced the effect of A-23187 on the level of cyclic GMP. These results emphasize the role of Ca-2+ in the regulation of parotid cyclic nucleotide levels. Since the effects of the ionophore depended on the presence of Ca-2+, it is possible that some of the effects of agonists on parotid gland physiology are secondary to an action on intracellular Ca-2+ distribution.

Amylases↗

PubMed2004

Interaction of thin wetting films of lecithin with some divalent cations.

The impact of some divalent cations on the structure of a model membrane system, comprising wetting lipid films, is assayed in this work. The results from impedimetry suggest prominent structural changes upon the addition of the discussed ions to the electrolyte solution contacting the film. These changes are manifested by the increase of resistivity of the films as well as by the decrease of capacitance dispersion. In accordance with other data in the literature, manganese (Mn(2+)) turned out to have an effect greater than those of Mg(2+) and Ca(2+).

Calcium↗

PubMed1996

Effect of divalent cations on the contractile response of rat aorta to depolarization before and after nifedipine treatment.

The influence of the divalent cations, Ca2+, Mg2+ and Ba2+, on the contractile response of the rat aorta to KCl and on the recovery of this response after nifedipine treatment was analyzed. KCl (80 mmol/l) promoted a two-phase (phasic and tonic) contractile response in Krebs solution but, as expected, no contractile response in Ca(2+)-free medium. In Mg(2+)-free medium, the phasic response to KCl was unaffected but the tonic one decreased slowly, suggesting that a long incubation time in the absence of Mg2+ (65 min) promotes a loss of or a change in the intracellular distribution of this ion that modifies Ca2+ entry through L channels or Ca2+ handling. Ba2+ (1.8 mmol/l) contracted the rat aorta in the absence or presence of Ca2+ but, when Ca2+ was not present, Ba2+ modified the contractile process so that a new addition of KCl did not reproduce the contractile response. After nifedipine treatment, no phasic response to KCl was observed and only a slow response appeared that increased as long as the KCl was present (45 min). After washing, KCl induced a phasic response similar to the standard response, but a smaller tonic one. Divalent cations accelerated the recovery of the channels involved in the tonic contraction. However, the recovery of the Ca2+ channels related to the phasic contraction was independent of divalent cations.

Analysis of Variance↗

PubMed1994

Inositol (1,4,5)-trisphosphate (InsP3)-gated Ca channels from cerebellum: conduction properties for divalent cations and regulation by intraluminal calcium.

The conduction properties of inositol (1,4,5)-trisphosphate (InsP3)-gated calcium (Ca) channels (InsP3R) from canine cerebellum for divalent cations and the regulation of the channels by intraluminal Ca were studied using channels reconstituted into planar lipid bilayers. Analysis of single-channel recordings performed with different divalent cations present at 55 mM on the trans (intraluminal) side of the membrane revealed that the current amplitude at 0 mV and the single-channel slope conductance fell in the sequence: Ba (2.2 pA, 85 pS) > Sr (2.0 pA, 77 pS) > Ca (1.4 pA, 53 pS) > Mg (1.1 pA, 42 pS). The mean open time of the InsP3R recorded with Ca (2.9 ms) was significantly shorter than with other divalent cations (approximately 5.5 ms). The "anomalous mole fraction effect" was not observed in mixtures of divalent cations (Mg and Ba), suggesting that these channels are single-ion pores. Measurements of InsP3R activity at different intraluminal Ca levels demonstrated that Ca in the submillimolar range did not potentiate channel activity, and that very high levels of intraluminal Ca (> or = 10 mM) decreased channel open probability 5-10-fold. When InsP3R were measured with Ba as a current carrier in the presence of 110 mM cis potassium, a PBa/PK of 6.3 was estimated from the extrapolated value for the reversal potential. When the unitary current through the InsP3R at 0 mV was measured as a function of the permeant ion (Ba) concentration, the half-maximal current occurred at 10 mM trans Ba. The following conclusions are drawn from these data: (a) the conduction properties of InsP3R are similar to the properties of the ryanodine receptor, another intracellular Ca channel, and differ dramatically from the properties of voltage-gated Ca channels of the plasma membrane. (b) The estimated size of the Ca current through the InsP3R under physiological conditions is 0.5 pA, approximately four times less than the Ca current through the ryanodine receptor. (c) The potentiation of InsP3R by intraluminal Ca in the submillimolar range remains controversial. (d) A quantitative model that explains the inhibitory effects of high trans Ca on InsP3R activity was developed and the kinetic parameters of InsP3R gating were determined.

Animals↗

PubMed1993

Distinct divalent cation requirements for integrin-mediated CD4+ T lymphocyte adhesion to ICAM-1, fibronectin, VCAM-1, and invasin.

Integrins are a large family of cell surface receptors that mediate the adhesion of cells to other cells and to components of the extracellular matrix. Various divalent cations, particularly Ca2+ and Mn2+, have been shown to modulate the functional activity of many different integrins expressed on a wide variety of cell types. In this study, we have characterized the divalent cation requirements for the adhesion of human peripheral CD4+ T cells to four distinct integrin ligands: the alpha 4 beta 1 and alpha 5 beta 1 ligand fibronectin, the alpha 4 beta 1 ligand VCAM-1, the LFA-1 ligand ICAM-1, and the alpha 4 beta 1 bacterial ligand invasin. We find that there are distinct divalent cation requirements for T cell adhesion to each of these ligands: 1) Mg2+/EGTA treatment selectively up-regulates T cell adhesion to ICAM-1; 2) Mn2+ coordinately up-regulates adhesion to ICAM-1, fibronectin, and VCAM-1, with a peak response at 100 microM Mn2+; 3) Ca2+ can selectively support adhesion to VCAM-1 induced by activation and inhibit Mn(2+)-dependent adhesion to ICAM-1; and 4) binding to invasin is maximal in the presence of Ca2+, Mg2+, or Mn2+. Furthermore, divalent cation modifications do not fully up-regulate T cell adhesion to fibronectin, VCAM-1, and ICAM-1, because additional cell activation with phorbol ester treatment can further enhance adhesion in the presence of Mn2+. These results suggest that modification of divalent cations may provide a mechanism by which an individual integrin receptor/ligand interaction can be specifically and selectively regulated.

Adhesins, Bacterial↗

PubMed2004

Pre- and post-synaptic effects of manipulating surface charge with divalent cations at the photoreceptor synapse.

Persistence of horizontal cell (HC) light responses in extracellular solutions containing low Ca2+ plus divalent cations to block Ca2+ currents (ICa) has been attributed to Ca2+-independent neurotransmission. Using a retinal slice preparation to record both ICa and light responses, we demonstrate that persistence of HC responses in low [Ca2+]o can instead be explained by a paradoxical increase of Ca2+ influx into photoreceptor terminals arising from surface charge-mediated shifts in ICa activation. Consistent with this explanation, application of Zn2+ or Ni2+ caused a hyperpolarizing block of HC light responses that was relieved by lowering [Ca2+]o. The same concentrations of Zn2+ and Ni2+ reduced the amplitude of ICa at the rod dark potential and this reduction was relieved by a hyperpolarizing shift in voltage dependence induced by lowering [Ca2+]o. Block of ICa by Mg2+, which has weak surface charge effects, was not relieved by low [Ca2+]o. Recovery of HC responses in low [Ca2+]o was assisted by enhancement of rod light responses. To bypass light stimulation, OFF bipolar cells were stimulated by steps to -40 mV applied to presynaptic rods during simultaneous paired recordings. Consistent with surface charge theory, the post-synaptic current was inhibited by Zn2+ and this inhibition was relieved by lowering [Ca2+]o. Nominally divalent-free media produced inversion of HC light responses even though rod light responses remained hyperpolarizing; HC response inversion can be explained by surface charge-mediated shifts in ICa. In summary, HC light responses modifications induced by low divalent cation solutions can be explained by effects on photoreceptor light responses and membrane surface charge without necessitating Ca2+-independent neurotransmission. Furthermore, these results suggest that surface charge effects accompanying physiological changing divalent cation levels in the synaptic cleft may provide a means for modulating synaptic output from photoreceptors.

Animals↗

PubMed1987

Solubilization of a divalent cation dependent ATPase from dog heart sarcolemma.

Heart sarcolemma has been shown to contain an ATPase hydrolyzing system which is activated by millimolar concentrations of divalent cations such as Ca2+ or Mg2+. Although Ca2+-dependent ATPase is released upon treating sarcolemma with trypsin, a considerable amount of the divalent cation dependent ATPase activity was retained in the membrane. This divalent cation dependent ATPase was solubilized by sonication of the trypsin-treated dog heart sarcolemma with 1% Triton X-100. The solubilized enzyme was subjected to column chromatography on a Sepharose-6B column, followed by ion-exchange chromatography on a DEAE cellulose column. The enzyme preparation was found to be rather labile and thus the purity of the sample could not be accurately assessed. The solubilized ATPase preparations did not show any cross-reactivity with dog heart myosin antiserum or with Na+ + K+ ATPase antiserum. The enzyme was found to be insensitive to inhibitors such as ouabain, verapamil, oligomycin and vanadate. The enzyme preparation did not exhibit any Ca2+-stimulated Mg2+ dependent ATPase activity. Furthermore, the low affinity of the enzyme for Ca2+ (Ka = 0.3 mM) rules out the possibility of its involvement in the Ca2+ pump mechanism located in the plasma membrane of the cardiac cell.

Adenosine Triphosphatases↗

PubMed1991

High affinity divalent cation exchange on actin. Association rate measurements support the simple competitive model.

Each actin molecule has one high affinity site which binds a divalent cation. It has been proposed that an isomerization of the actin molecule is involved in divalent cation exchange at this site ("isomerization model," Frieden, C. (1982) J. Biol. Chem. 257, 2882-2886); we have maintained that exchange is by a simple competitive mechanism (Estes, J. E., Selden, L. A., and Gershman, L. C. (1987) J. Biol. Chem. 262, 4952-4957). Here, using fluorescent-labeled actin, we measure the apparent rate constant for exchange (kapp) as a function of the ratio of free Ca2+ and Mg2+ concentrations, ([Ca]/[Mg]), and show that both models are consistent with the data. The major parameter controlling this relationship in the simple competitive exchange model, the ratio of the association rate constants for Ca2+ and Mg2+ to actin (kCa/kMg), is found to have a value of about 90. We have verified this parameter by direct measurements of kCa and kMg, finding that kCa = 1.9 x 10(7) M-1 s-1 and kMg = 2.3 x 10(5) M-1 s-1, consistent with the characteristics of the Ca2+ and Mg2+ aquo ions. The corresponding parameter derived from the isomerization model is not verifiable. We conclude that high affinity divalent cation exchange on actin proceeds by a simple competitive mechanism.

Actins↗

PubMed2006

Divalent cations as modulators of neuronal excitability: emphasis on copper and zinc.

Based on indirect evidence, a role for synaptically released copper and zinc as modulators of neuronal activity has been proposed. To test this proposal directly, we studied the effect of copper, zinc, and other divalent cations on voltage-dependent currents in dissociated toad olfactory neurons and on their firing rate induced by small depolarizing currents. Divalent cations in the nanomolar range sped up the activation kinetics and increased the amplitude of the inward sodium current. In the micromolar range, they caused a dose dependent inhibition of the inward Na+ and Ca2+ currents (INa and ICa) and reduced de amplitude of the Ca2+-dependent K+ outward current (ICa-K). On the other hand, the firing rate of olfactory neurons increased when exposed to nanomolar concentration of divalent cations and decreased when exposed to micromolar concentrations. This biphasic effect of divalent cations on neuronal excitability may be explained by the interaction of these ions with high and low affinity sites in voltage-gated channels. Our results support the idea that these ions are normal modulators of neuronal excitability.

Action Potentials↗

PubMed1994

Temperature-dependent modification of divalent cation flux in the rat parotid gland basolateral membrane.

Divalent cation (Mn2+, Ca2+) entry into rat parotid acinar cells is stimulated by the release of Ca2+ from the internal agonist-sensitive Ca2+ pool via a mechanism which is not yet defined. This study examines the effect of temperature on Mn2+ influx into internal Ca2+ pool-depleted acini (depl-acini, as a result of carbachol stimulation of acini in a Ca(2+)-free medium for 10 min) and passive 45Ca2+ influx in basolateral membrane vesicles (BLMV). Mn2+ entry into depl-acini was decreased when the incubation temperature was lowered from 37 to 4 degrees C. At 4 degrees C, Mn2+ entry appeared to be inactivated since it was not increased by raising extracellular [Mn2+] from 50 microM up to 1 mM. The Arrhenius plot of depletion-activated Mn2+ entry between 37 and 8 degrees C was nonlinear, with a change in the slope at about 21 degrees C. The activation energy (Ea) increased from 10 kcal/mol (Q10 = 1.7) at 21-37 degrees C to 25 kcal/mol (Q10 = 3.0) at 21-8 degrees C. Under the same conditions, Mn2+ entry into basal (unstimulated) cells and ionomycin (5 microM) permeabilized depl-acini exhibit a linear decrease, with Ea of 7.8 kcal/mol (Q10 = 1.5) and 6.2 kcal/mol (Q10 < 1.5), respectively. These data suggest that depletion-activated Mn2+ entry into parotid acini is regulated by a mechanism which is strongly temperature dependent and distinct from Mn2+ entry into unstimulated acini.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗