Role of membrane-associated CA2+ dependent matrix metalloprotease-2 in the oxidant activation of Ca2+ATPase by tertiary butylhydroperoxide

Treatment of bovine pulmonary artery smooth muscle plasma membrane suspension with the oxidant tert-butylhydroperoxide (t-buOOH) increases Ca²⁺⁺ATPase activity. The smooth muscle plasma membrane possesses a Ca²⁺⁺ dependent protease activity in the gelatin containing zymogram having an apparent molecular mass of 72 kDa. The 72 kDa protease activity was found to be inhibited by EGTA and the tissue inhibitor of metalloprotease-2 (TIMP-2). Since 72 kDa is the molecular mass of MMP-2 and since in our present study the 72 kDa protease in the gelatin containing zymogram is inhibited by matrix metalloprotease inhibitors, EGTA and TIMP-2, it may be suggested that the 72 kDa protease is the MMP-2. In addition to the increasing Ca²⁺⁺ATPase activity, t-buOOH also enhances the activity of the membrane associated Ca²⁺⁺ dependent protease that degrades ¹⁴C-gelatin. The oxidant triggered protease activity and the Ca²⁺⁺ATPase activity were found to be prevented by the antioxidant vitamin E, and also by the Ca²⁺⁺ dependent matrix metalloprotease inhibitors: EGTA and TIMP-2. Adding MMP-2 to the smooth muscle plasma membrane suspension caused an increase in Ca²⁺⁺ATPase activity and pretreatment with TIMP-2 prevents the increase in Ca²⁺⁺ATPase activity. Combined treatment of the smooth muscle plasma membrane with low doses of MMP-2 and t-buOOH augments further the Ca²⁺⁺ATPase activity caused by the respective doses of either t-buOOH or MMP-2. Pretreatment with TIMP-2 prevents the increase in Ca²⁺⁺ATPase activity elicited by the low doses of MMP-2 and/or t-buOOH.     

Plant Ca<Superscript>2+</Superscript>-ATPases

Plant calcium pumps, similarly to animal Ca²⁺ pumps, belong to the superfamily of P-type ATPase comprising also the plasma membrane H⁺-ATPase of fungi and plants, Na⁺/K⁺ ATPase of animals and H⁺/K⁺ ATPase of mammalian gastric mucosa. According to their sensitivity to calmodulin the plant Ca²⁺-ATPases have been divided into two subgroups: type IIA (homologues of animal SERCA) and type IIB (homologues of animal PMCA). Regardless of the similarities in a protein sequence, the plant Ca²⁺ pumps differ from those in animals in their cellular localization, structure and sensitivity to inhibitors. Genomic investigations revealed multiplicity of plant Ca²⁺-ATPases; they are present not only in the plasma membranes and ER but also in membranes of most of the cell compartments, such as vacuole, plastids, nucleus or Golgi apparatus. Studies using yeast mutants made possible the functional and biochemical characterization of individual plant Ca²⁺-ATMPases. Plant calcium pumps play an essential role in signal transduction pathways, they are responsible for the regulation of [Ca²⁺] in both cytoplasm and endomembrane compartments. These Ca²⁺-ATPases appear to be involved in plant adaptation to stress conditions, like salinity, chilling or anoxia.     

Role of MMP-2 in inhibiting Na+ dependent Ca2+ uptake by H2O2 in microsomes isolated from pulmonary smooth muscle

Treatment of microsomes (preferentially enriched with endoplasmic reticulum) isolated from bovine pulmonary artery smooth muscle tissue with H₂O₂ (1 mM) markedly stimulated matrix metalloproteinase activity and also inhibited Na⁺ dependent Ca²⁺ uptake. Electron micrograph revealed that H₂O₂ (1 mM) does not cause any damage to the microsomes. MMP-2 and TIMP-2 were determined to be the ambient protease and corresponding antiprotease of the microsomes. Pretreatment with vitamin E (1 mM) and TIMP-2 (50 g/ml) reversed the effect produced by H₂O₂ (1 mM) on Na⁺ dependent Ca²⁺ uptake in the microsomes. However, H₂O₂ (1 mM) caused changes in MMP-2 activity and Na⁺ dependent Ca²⁺ uptake were not reversed upon pretreatment of the microsomes with a low concentration of 5 g/ml of TIMP-2 which otherwise reversed MMP-2 (1 g/ml) mediated increase in ¹⁴C-gelatin degradation and inhibition of Na⁺ dependent Ca²⁺ uptake. Combined treatment of the microsomes with a low dose of MMP-2 (0.5 g/ml) and H₂O₂ (0.5 mM) inhibited Na⁺ dependent Ca²⁺ uptake in the microsomes compared to the respective low dose of either of them. Direct treatment of TIMP-2 (5 g/ml) with H₂O₂ (1 mM) abolished the inhibitory effect of the inhibitor on ¹⁴C-gelatinolytic activity elicited by 1 g/ml of MMP-2. Thus, one of the mechanisms by which H₂O₂ activates MMP-2 could be due to inactivation of TIMP-2 by the oxidant. The resulting activation of MMP-2 subsequently inhibits Na⁺ dependent Ca²⁺ uptake in the microsomes. (Mol Cell Biochem 270: 79–87, 2005)     

Hypoxic Modulation of Ca<Superscript>2+</Superscript> Signaling in Human Venous and Arterial Endothelial Cells

Our understanding of vascular endothelial cell physiology is based on studies of endothelial cells cultured from various vascular beds of different species for varying periods of time. Systematic analysis of the properties of endothelial cells from different parts of the vasculature is lacking. Here, we compare Ca²⁺ homeostasis in primary cultures of endothelial cells from human internal mammary artery and saphenous vein and how this is modified by hypoxia, an inevitable consequence of bypass grafting (2.5% O₂, 24 h). Basal [Ca²⁺] _i and store depletion-mediated Ca²⁺ entry were significantly different between the two cell types, yet agonist (ATP)–mediated mobilization from endoplasmic reticulum stores was similar. Hypoxia potentiated agonist-evoked responses in arterial, but not venous, cells but augmented store depletion-mediated Ca²⁺ entry only in venous cells. Clearly, Ca²⁺ signaling and its remodeling by hypoxia are strikingly different in arterial vs. venous endothelial cells. Our data have important implications for the interpretation of data obtained from endothelial cells of varying sources.     

L-type Ca<Superscript>2+</Superscript> channel and Ca<Superscript>2+</Superscript>-permeable nonselective cation channel as a Ca<Superscript>2+</Superscript> conducting pathway in myocytes isolated from the cricket lateral oviduct

The Ca²⁺-conducting pathway of myocytes isolated from the cricket lateral oviduct was investigated by means of the whole-cell patch clamp technique. In voltage-clamp configuration, two types of whole cell inward currents were identified. One was voltage-dependent, initially activated at –40 mV and reaching a maximum at 10 mV with the use of 140 mM Cs²⁺-aspartate in the patch pipette and normal saline in the bath solution. Replacement of the external Ca²⁺ with Ba²⁺ slowed the current decay. Increasing the external Ca²⁺ or Ba²⁺ concentration increased the amplitude of the inward current and the current–voltage (I–V) relationship was shifted as expected from a screening effect on negative surface charges. The inward current could be carried by Na⁺ in the absence of extracellular Ca²⁺. Current carried by Na⁺ (I _Na) was almost completely blocked by the dihydropyridine Ca²⁺ channel antagonist, nifedipine, suggesting that the I _Na is through voltage-dependent L-type Ca²⁺ channels. The other inward current is voltage-independent and its I–V relationship was linear between –100 mV to 0 mV with a slight inward rectification at more hyperpolarizing membrane potentials when 140 mM Cs⁺-aspartate and 140 mM Na⁺-gluconate were used in the patch pipette and in the bath solution, respectively. A similar current was observed even when the external Na⁺ was replaced with an equimolar amount of K⁺ or Cs⁺, or 50 mM Ca²⁺ or Ba²⁺. When the osmolarity of the bath solution was reduced by removing mannitol from the bath solution, the inward current became larger at negative potentials. The I–V relationship for the current evoked by the hypotonic solution also showed a linear relationship between –100 mV to 0 mV. Bath application of Gd³⁺ (10 M) decreased the inward current activated by membrane hyperpolarization. These results clearly indicate that the majority of current activated by a membrane hyperpolarization is through a stretch-activated Ca²⁺-permeable nonselective cation channel (NSCC). Here, for the first time, we have identified voltage-dependent L-type Ca²⁺ channel and stretch-activated Ca²⁺-permeable NSCCs from enzymatically isolated muscle cells of the cricket using the whole-cell patch clamp recording technique.Abbreviations I _Ca Ca²⁺ current - I _Na Na⁺ current - I–V current–voltage - NSCC nonselective cation channel Communicated by G. Heldmaier     

Ca<Superscript>2+</Superscript> binding protein-1 inhibits Ca<Superscript>2+</Superscript> currents and exocytosis in bovine chromaffin cells

Summary Calcium binding protein-1 (CaBP1) is a calmodulin like protein shown to modulate Ca²⁺ channel activities. Here, we explored the functions of long and short spliced CaBP1 variants (L- and S-CaBP1) in modulating stimulus-secretion coupling in primary cultured bovine chromaffin cells. L- and S-CaBP1 were cloned from rat brain and fused with yellow fluorescent protein at the C-terminal. When expressed in chromaffin cells, wild-type L- and S-CaBP1s could be found in the cytosol, plasma membrane and a perinuclear region; in contrast, the myristoylation-deficient mutants were not found in the membrane. More than 20 and 70% of Na⁺ and Ca²⁺ currents, respectively, were inhibited by wild-type isoforms but not myristoylation-deficient mutants. The [Ca²⁺] _i response evoked by high K⁺ buffer and the exocytosis elicited by membrane depolarizations were inhibited only by wild-type isoforms. Neuronal Ca²⁺ sensor-1 and CaBP5, both are calmodulin-like proteins, did not affect Na⁺, Ca²⁺ currents, and exocytosis. When expressed in cultured cortical neurons, the [Ca²⁺] _i responses elicited by high-K⁺ depolarization were inhibited by CaBP1 isoforms. In HEK293T cells cotransfected with N-type Ca²⁺ channel and L-CaBP1, the current was reduced and activation curve was shifted positively. These results demonstrate the importance of CaBP1s in modulating the stimulus-secretion coupling in excitable cells. M.-L. Chen and Y.-C. Chen contributed equally to this study     

Role of hydroxyl radical in the oxidant H2O2-mediated Ca2+ release from pulmonary smooth muscle mitochondria

We sought to investigate the mechanism(s) by which the oxidant H₂O₂ stimulates Ca²⁺ release from mitochondria of bovine pulmonary vascular smooth muscle tissue and to test the hypothesis that hydroxyl radical is involved in this phenomenon. Treatment of the smooth muscle tissue with 1 mM H₂O₂ dramatically stimulated hydroxyl radical generation as measured by methane (CH₄) production by GLC using dimethylsulfoxide (DMSO) as the substrate. Pretreatment of the mitochondria with the hydroxyl radical scavanger dimethylthiourea (DMTU) prevented the increase in CH₄ production caused by H₂O₂. In the absence of EGTA, H₂O₂ caused stimulation of Ca²⁺ release from mitochondria occurred with a lag time of about 4 min. Addition of EGTA to Ca²⁺ loaded mitochondria resulted an immediate loss of Ca²⁺ and that has been found to be augmented by H₂O₂. The release of Ca₂₊ by H₂O₂ did not appear to occur with concommitant increase in sucrose entry into, K⁺ release from, and swelling of mitochondria when the Ca²⁺ cycling was prevented by EGTA. These observations suggested that H₂O₂-mediated Ca²⁺ release from bovine pulmonary vascular smooth muscle tissue mitochondria occurred (i) through the involvement of hydroxyl radical; (ii) via specific pathway(s); and (iii) did not appear to happen primarily via nonspecific pore formation.Abbreviations H₂O₂ hydrogen peroxide - OH· hydroxyl radical - t-buOOH tert-butyl hydroperoxide - CH₄ methane - GLC gas liquid chromatography - DMTU dimethylthiourea - EGTA ethylene glycol bis(-aminoethyl ether) - N Ntetraacetic acid - DMSO dimethyl sulfoxide - CH₄ methane - HBPS Hank's buffered physiological saline - HEPES N-2-hydroxyethylpiperazine-N-2-ethane sulfonic acid - MOPS 3-(N-morpholino)propane sulfonic acid - Tris tris (hydroxymethyl aminomethane)     

Regulation of Plant Plasma Membrane H<Superscript>+</Superscript>- and Ca<Superscript>2+</Superscript>-ATPases by Terminal Domains

In the last few years, major progress has been made to elucidate the structure, function, and regulation of P-type plasma membrane H⁺-and Ca²⁺-ATPases. Even though a number of regulatory proteins have been identified, many pieces are still lacking in order to understand the complete regulatory mechanisms of these pumps. In plant plasma membrane H⁺- and Ca²⁺-ATPases, autoinhibitory domains are situated in the C- and N-terminal domains, respectively. A model for a common mechanism of autoinhibition is discussed.     

Intracellular Ca<Superscript>2+</Superscript> Pools and Fluxes in Cardiac Muscle-Derived H9c2 Cells

Relevant Ca²⁺ pools and fluxes in H9c2 cells have been studied using fluorescent indicators and Ca²⁺-mobilizing agents. Vasopressin produced a cytoplasmic Ca²⁺ peak with half-maximal effective concentration of 6 nM, whereas thapsigargin-induced Ca²⁺ increase showed half-maximal effect at 3 nM. Depolarization of the mitochondrial inner membrane by protonophore was also associated with an increase in cytoplasmic Ca²⁺. Ionomycin induced a small and sustained depolarization, while thapsigargin had a small but transient effect. The thapsigargin-sensitive Ca²⁺ pool was also sensitive to ionomycin, whereas the protonophore-sensitive Ca²⁺ pool was not. The vasopressin-induced cytoplasmic Ca²⁺ signal, which caused a reversible discharge of the sarco-endoplasmic reticulum Ca²⁺ pool, was sensed as a mitochondrial Ca²⁺ peak but was unaffected by the permeability transition pore inhibitor cyclosporin A. The mitochondrial Ca²⁺ peak was affected by cyclosporin A when the Ca²⁺ signal was induced by irreversible discharge of the intracellular Ca²⁺ pool, i.e., adding thapsigargin. These observations indicate that the mitochondria interpret the cytoplasmic Ca²⁺ signals generated in the reticular store.     

Mitochondrial Ca<Superscript>2+</Superscript> cycle mediated by the palmitate-activated cyclosporin a-insensitive pore

Earlier we found that in isolated rat liver mitochondria the reversible opening of the mitochondrial cyclosporin A-insensitive pore induced by low concentrations of palmitic acid (Pal) plus Ca²⁺ results in the brief loss of Δψ [Mironova et al., J Bioenerg Biomembr (2004), 36:171–178]. Now we report that Pal and Ca²⁺, increased to 30 and 70 nmol/mg protein respectively, induce a stable and prolonged (10 min) partial depolarization of the mitochondrial membrane, the release of Ca²⁺ and the swelling of mitochondria. Inhibitors of the Ca²⁺ uniporter, ruthenium red and La³⁺, as well as EGTA added in 10 min after the Pal/Ca²⁺-activated pore opening, prevent the release of Ca²⁺ and repolarize the membrane to initial level. Similar effects can be observed in the absence of exogeneous Pal, upon mitochondria accumulating high [Sr²⁺], which leads to the activation of phospholipase A₂ and appearance of endogenous fatty acids. The paper proposes a new model of the mitochondrial Ca²⁺ cycle, in which Ca²⁺ uptake is mediated by the Ca²⁺ uniporter and Ca²⁺ efflux occurs via a short-living Pal/Ca²⁺-activated pore.     

Influence of Ca<Superscript>2+</Superscript> Oscillatory Influx on Membrane Ca<Superscript>2+</Superscript>-ATPase Activity: a Kinetic Model

A kinetic model for the membrane Ca²⁺-ATPase is considered. The catalytic cycle in the model is extended by enzyme auto-inhibition and by oscillatory calcium influx. It is shown that the conductive enzyme activity can be registered as damped or sustained Ca²⁺ pulses similar to observed experimentally. It is shown that frequency variations in Ca²⁺ oscillatory influx induce changes of pulsating enzyme activity. Encoding is observed for the signal frequency into a number of fixed levels of sustained pulses in the enzyme activity. At certain calcium signal frequencies, the calculated Ca²⁺-ATPase conductivity demonstrates chaotic multi-level pulses, similar to those observed experimentally.__________Translated from Biokhimiya, Vol. 70, No. 4, 2005, pp. 539–544.Original Russian Text Copyright © 2005 by Goldstein, Mayevsky, Zakrjevskaya.     

Evolution of mechanisms of Ca<Superscript>2+</Superscript>-signaling. Significance of Ca<Superscript>2+</Superscript>-messenger systems during transition of organisms to multicellularity

The review considers Ca²⁺-messenger systems in primitive multicellulars (sponges and hydrozoa organisms). Analysis is performed of Ca²⁺ participation in regulation of early development of the organisms, their mobility, metamorphosis, chemoreception, and some other functions.     

Nicotine-induced Ca<Superscript>2+</Superscript>-myristoyl Switch of Neuronal Ca<Superscript>2+</Superscript> Sensor VILIP-1 in Hippocampal Neurons: A Possible Crosstalk Mechanism for Nicotinic Receptors

Visinin-like protein (VILIP-1) belongs to the neuronal Ca²⁺ sensor family of EF-hand Ca²⁺-binding proteins that regulate a variety of Ca²⁺-dependent signal transduction processes in neurons. It is an interaction partner of α4β2 nicotinic acetylcholine receptor (nAChR) and increases surface expression level and agonist sensitivity of the receptor in oocytes. Nicotine stimulation of nicotinic receptors has been reported to lead to an increase in intracellular Ca²⁺ concentration by Ca²⁺-permeable nAChRs, which in turn might lead to activation of VILIP-1, by a mechanism described as the Ca²⁺-myristoyl switch. It has been postulated that this will lead to co-localization of the proteins at cell membranes, where VILIP-1 can influence functional activity of α4-containing nAChRs. In order to test this hypothesis we have investigated whether a nicotine-induced and reversible Ca²⁺-myristoyl switch of VILIP-1 exists in primary hippocampal neurons and whether pharmacological agents, such as antagonist specific for distinct nAChRs, can interfere with the Ca²⁺-dependent membrane localization of VILIP-1. Here we report, that only α7- but not α4-containing nAChRs are able to elicit a Ca²⁺-dependent and reversible membrane-translocation of VILIP-1 in interneurons as revealed by employing the specific receptor antagonists dihydro-beta-erythroidine and methylallylaconitine. The nAChRs are associated with processes of synaptic plasticity in hippocampal neurons and they have been implicated in the pathology of CNS disorders, including Alzheimer’s disease and schizophrenia. VILIP-1 might provide a novel functional crosstalk between α4- and α7-containing nAChRs.     

Stimulation of plasma membrane Ca2+-pump ATPase of vascular smooth muscle by cGMP-dependent protein kinase: Functional reconstitution with purified proteins

A 240-kDa protein isolated from porcine aortic smooth muscle as a substrate for cGMP-dependent protein kinase (cGMP kinase) whose phosphorylation was in a close association with stimulation of partially purified plasma membrane Ca2+-pump ATPase by the kinase was later shown to represent splicing variants of type 1 inositol 1,4,5-trisphosphate (IP3) receptor. To further clarify the role played by this protein in the stimulation of Ca2+-pump ATPase, it was attempted in the present study to specifically remove the protein by immunoprecipitation with an antibody specific to type 1 IP3 receptor. Contrary to expectation, stimulation of the ATPase by cGMP kinase was still observed after removal of the IP3 receptor. Furthermore, cGMP kinase stimulated a highly purified preparation of Ca2+-pump ATPase deprived of IP3 receptor when the concentrations of the ATPase were low enough (10-20 nM) to make it retain a monomeric form, while it did not produce stimulation when the concentration of the enzyme was increased to 40 nM at which the enzyme is known to take an oligomeric, fully activated form insensitive to activation by calmodulin. Heat-inactivated cGMP kinase and cGMP kinase without cGMP failed to stimulate the highly purified Ca2+-pump ATPase. In addition, type I but not type I cGMP kinase was found to stimulate the ATPase. The stimulation of Ca2+-pump ATPase by cGMP kinase occurs without any detectable phosphorylation of the ATPase. In conclusion, cGMP kinase can stimulate the plasma membrane Ca2+-pump ATPase when it is in a monomeric form without phosphorylating the Ca2+-pump ATPase and that of the two cGMP kinase isozymes found in the vascular smooth muscle, only type I cGMP kinase participates in the stimulation.     

Purification and characterization of a cytosolic Ca<Superscript>2+</Superscript>-independent phospholipase A<Subscript>2</Subscript> from bovine brain

The Ca²⁺-independent phospholipase A₂ (iPLA₂) subfamily of enzymes is associated with arachidonic acid (AA) release and the subsequent increase in fatty acid turnover. This phenomenon occurs not only during apoptosis but also during inflammation and lymphocyte proliferation. In this study, we purified and characterized a novel type of iPLA₂ from bovine brain. iPLA₂ was purified 4,174-fold from the bovine brain by a sequential process involving DEAE-cellulose anion exchange, phenyl-5PW hydrophobic interaction, heparin-Sepharose affinity, Sephacryl S-300 gel filtration, Mono S cation exchange, Mono Q anion exchange, and Superose 12 gel filtration. A single peak of iPLA₂ activity was eluted at an apparent molecular mass of 155 kDa during the final Superose 12 gel-filtration step. The purified enzyme had an isoelectric point of 5.3 on twodimensional gel electrophoresis (2-DE) and was inhibited by arachidonyl trifluoromethyl ketone (AACOCF₃), Triton X-100, iron, and Ca²⁺. However, it was not inhibited by bromoenol lactone (BEL), an inhibitor of iPLA₂, and adenosine triphosphate (ATP). The spot with the iPLA₂ activity did not match with any known protein sequence, as determined by matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) analysis. Altogether, these data suggest that the purified enzyme is a novel form of cytosolic iPLA₂.     

Ca<Superscript>2+</Superscript> channel currents of cortical neurons from pure and mixed cultures

Voltage-gated Ca²⁺ channels (VGCCs) are key regulators of many neuronal functions, and involved in multiple central nervous system diseases. In the last 30 years, a large number of injury and disease models have been established based on cultured neurons. Culture with serum develops a mixture of neurons and glial cells, while culture without serum develops pure neurons. Both of these neuronal-culture methods are widely used. However, the properties of Ca²⁺ currents in neurons from these two cultures have not been compared. In this study, we cultured rat cortical neurons in serum-containing or -free medium and then recorded the Ca²⁺ channel currents using patch-clamp technique. Our results showed that there were significant differences in the amplitude and activation properties of whole-cell Ca²⁺ channel currents, and of non-L-type Ca²⁺ channel currents between the neurons from these two culture systems. Our data suggested that the difference of whole-cell Ca²⁺ currents may result from the differences in non-L-type currents. Understanding of these properties will considerably advance studies of VGCCs in neurons from pure or mixed culture.     

The role of mitochondrial palmitate/Ca<Superscript>2+</Superscript>-activated pore in palmitate-induced apoptosis

The evidence of possible involvement of the mitochondrial cyclosporin A-insensitive palmitate/Ca²⁺-activated pore in palmitate-induced apoptosis is presented. It has been established that the opening of the palmitate/Ca²⁺-activated pore results in the high-amplitude swelling of mitochondria and the release of the apoptosis-inducing factor from organelles. These processes are accompanied by a short-term slight decrease of membrane potential, which recovers in 1 min. The possible role of the palmitate/Ca²⁺-activated pore in the induction of palmitate-induced apoptosis is discussed.     

Store-Dependent Entry of Ca<Superscript>2+</Superscript> into Sensory Neurons of the Rat

We studied store-dependent (activated by depletion of the endoplasmic reticulum, ER, store) entry of Ca²⁺ from the extracellular medium into neurons of the rat spinal ganglia (small- and medium-sized cells; diameter, 18 to 36 μm). Activation of ryanodine-sensitive receptors of the ER in the studied neurons superfused by Tyrode solutions containing Ca²⁺ or with no Ca²⁺ was provided by application of 10 mM caffeine. The decay phase of caffeine-induced calcium transients in a Ca²⁺-containing solution was significantly longer than that in a Ca²⁺-free solution. This fact allows us to suppose that such a phenomenon is determined by Ca²⁺ entry into the neuron from the extracellular medium activated by caffeine-induced depletion of the ER store. Substitution of Ca²⁺-free extracellular solution by Ca²⁺-containing Tyrode solution, after depletion of the ER stores induced by applications of 100 nM ryanodine, 200 μM ATP, or 1 μM thapsigargin, resulted in increases in the concentration of intracellular Ca²⁺. These observations allow us to postulate that store-dependent Ca²⁺ entry into the studied neurons is activated after depletion not only of the inositol trisphosphate-sensitive ER store but also of the ryanodine-sensitive store. This entry also occurs after blocking of ATPases of the ER by thapsigargin. The kinetic characteristics of the rising phase of store-dependent Ca²⁺ entry induced by depletion of the ER stores under the influence of various agents are dissimilar; this can be related to different mechanisms of activation of such signals and/or to a compartmental organization of the ER. Neirofiziologiya/Neurophysiology, Vol. 37, No. 3, pp. 277–283, May–June, 2005.     

Calmodulin can induce and control damped oscillations in plasma membrane Ca<Superscript>2+</Superscript>-ATPase activity: A kinetic model

Plasma membrane Ca²⁺-ATPase is the pump that extrudes calcium ions from cells using ATP hydrolysis to maintain low Ca²⁺ concentrations in the cell. Calmodulin stimulates Ca²⁺-ATPase by binding to the autoinhibitory enzyme domain, which allows the access of cytoplasmic ATP and Ca²⁺ to the catalytic and transport sites. Our kinetic model predicts damped oscillations of the enzyme activity and interprets the known nonmonotonic kinetic behavior of the enzyme in the presence of calmodulin. For parameters close to experimental data, the kinetic model explains the dependence of the frequency and damping factor of the oscillatory enzyme activity on the calmodulin concentration. The calculated pre-steady-state curves fit well to known experimental data. Kinetic analysis allows us to assign Ca²⁺-ATPase to hysteretic enzymes exhibiting activity oscillations in open systems.