Extracellular regulation of fibroblast multiplication. Quantitative differences in nutrient and serum factor requirements for multiplication of normal and SV40 virus-transformed human lung cells

The principles of Henri-Michaelis-Menten kinetic analysis were applied to directly relate the concentration of serum growth factors and individual nutrients in the culture medium to the multiplication rate of a population of normal (N-HLF) and SV40 virus-transformed (SV-HLF) human lung fibroblasts. When all nutrient concentrations were optimal and in steady state, the concentration of serum factors that was required to support a half-maximal rate of proliferation of both N-HLF and SV-HLF was similar. When the serum factor concentration was optimal and constant, SV-HLF cells exhibited a reduced requirement (p less than 0.001) for 12 of 27 individual nutrients that were examined. Serum factors control the cellular requirement for Ca2+, K+, Mg2+, phosphate ions, and 2-oxocarboxylic acids for multiplication of N-HLF (McKeehan, W. L., and McKeehan, K. A. (1980) Proc. Natl, Acad, Sci, U. S. A. 77, 3417-3421). SV-HLF exhibited a constitutively reduced requirement for Ca2+, K+, and Mg2+ which partially removed the requirement for the 3 ions for multiplication of SV-HLF from the control of serum factors. The results suggest that SV40 virus transformation confers a growth advantage on human lung fibroblasts by alteration of their quantitative requirements for specific nutrients.     

Changes in microsomal Na+, K+-, Mg2+- and Ca2+-ATPase activities during proliferation of Chinese hamster V-79 and human HeLaS-3 cells

Changes in microsomal Na+, K+-, Mg2+- and Ca2+-ATPase activities during cell proliferation were examined in Chinese hamster V-79 (V-79) cells (normal cells) and human HeLaS-3 (HeLaS-3) cells (malignant cells). For V-79 cells, the Mg2+-ATPase activity per cell (pmol Pi/h/cell) in the confluent phase was higher than that in the logarithmically growing (log) phase. The amount of microsomal protein per cell was also high in the confluent phase. Specific activities (mumol Pi/h/mg protein) of Na+, K+-, Mg2+- and Ca2+-ATPase were significantly lower in the confluent phase than in the log phase. For HeLaS-3 cells, an increase in Ca2+-ATPase activity per cell was observed. The amount of microsomal protein per cell did not change between the log and confluent phase. The specific activity of Ca2+-ATPase in the confluent phase was also markedly higher than in the log phase. The relation between changes in ATPase activities and cell proliferation is discussed.     

Extracellular regulation of fibroblast multiplication: A direct kinetic approach to analysis of role of low molecular weight nutrients and serum growth factors

The principles of enzyme kinetic analysis were applied to quantitate the relationships among serum-derived growth factors, nutrients, and the rate of survival and multiplication of human fibroblasts in culture. The survival or multiplication rate of a population of cells plotted against an increasing concentration of a growth factor or nutrient in the medium exhibited a hyperbolic pattern that is characteristic of a dissociable, saturable interaction between cells and the ligands. Parameters equivalent to the K_m and V_max of enzyme kinetics were assigned to nutrients and growth factors. When all nutrient concentrations were optimized and in steady state, serum factors accelerated the rate of multiplication of a normal cell population. The same set of nutrients that supported a maximal rate of multiplication in the presence of serum factors supported the maintenance of non-proliferating cells in the absence of serum factors. Therefore, under this condition, serum factors are required for cell division and play a purely regulatory iole in multiplication of the cell population. The quantitative requirement for 18 nutrients of 29 that were examined was significantly higher (P < 0.001) for cell multiplication in the presence of serum factors than for cell maintenance in the absence of serum factors. This indicated specific nutrients that may be quantitatively important in cell division processes as well as in cell maintenance. The quantitative requirement for Ca²⁺, Mg²⁺, K⁺, Pi, and 2-oxocarboxylic acid for cell multiplication was modified by serum factors and other purified growth factors. The requirement for over 30 other nutrients could not clearly be related to the level of serum factors in the medium. Serum factors also determined the Ca²⁺, K⁺, and 2-oxocarboxylic acid requirement for maintenance of non-proliferating cells. Therefore, when either Ca²⁺, K⁺, or 2-oxocarboxylic acid concentration was limiting, factors in serum played a role as cell “survival or maintenance” factors in addition to their role in cell division as “growth regulatory” factors. However, with equivalent levels of serum factors in the medium, the requirement for Ca²⁺, K⁺, and 2-oxocarboxylic acids was still much higher for multiplication than for maintenance. Kinetic analysis revealed that the concentrations of individual nutrients modify the quantitative requirement for others for cell multiplication in a specific pattern. Thus, specific quantitative relationships among different nutrients in the medium are important in the control of the multiplication rate of the cell population. When all nutrient concentrations were optimal for multiplication of normal cells, the multiplication response of SV40-virus-transformed cells to serum factors was similar to that of normal cells. When serum factors were held constant, transformed cells required significantly less (P < 0.001) of 12 of the 26 nutrients examined. Therefore, the transformed cells only have a growth advantage when the external concentration of specific nutrients limits the multiplication rate of normal cells. Taken together, the results suggest that the control of cell multiplication is intimately related to external concentrations of nutrients. Specific growth regulatory factors may stimulate cell proliferation by modification of the response of normal cells to nutrients. Transforming agents may confer a selective growth advantage on cells by a constitutive alteration of their response to extracellular nutrients.     

The effects of calcium deficiency on the electrical activity of Nitella flexilis.

Calcium chelators such as ethylenediaminetetraacetic acid and sodium citrate produce repetitive activity and prolong the spike of internodal cells of Nitella flexilis. Removal of Ca2+, Mg2+, Na+, and K+ from the outside of the cell by washing the preparation with Tris propionate or Tris chloride hyperpolarizes the cells but does not initiate repetitive activity or increase the duration of the spike appreciably. It was concluded that cell-bound Ca2+ controls the threshold for stimulation and the duration of the spike, and that the removal of Ca2+ from the cell membrane, either by chelation or displacement, changes the normal behaviour of the cell by altering its permeability to some other ion or ions.     

Calcium, magnesium, and growth control in the WI-38 human fibroblast cell

WI-38 and SV40WI-38 cells have been synchronized using centrifugal elutriation. This technique allows for the rapid harvesting of early G1 phase cells from exponentially growing populations of both the normal and transformed cell. Using these cells, as well as WI-38 cells synchronized by serum deprivation, we have examined the effects of extracellular Ca and Mg levels on the progression of cells through G1 phase. A differential sensitivity to both Ca and Mg deprivation is observed between normal and transformed cells. The WI-38 cell requires higher levels of both ions for traversal of G1 phase and for continued proliferation as compared to the transformed cell. The temporal nature of the Ca and Mg requirements for the WI-38 cell has been examined during G1 phase. Ca is strictly required during early and late G1 phase, but not necessarily throughout mid-G1. An early as well as a late G1 Ca requirement is also found in serum-stimulated WI-38 cells. In contrast, the Mg requirement of WI-38 cells does not appear to be temporally well-defined. Mg appears to be a permissive factor, required throughout G1 phase rather than at certain prescribed intervals. On the basis of these data, it seems unlikely that these two cations exert their effects on cell growth entirely through a common competitive mechanism. Ca would appear to be involved in early serum or growth factor-mediated G1 events and later pre-S-phase events, as suggested in previous studies on other cell lines.     

Effect of R plasmid RPI on the nutritional requirements of Escherichia coli in batch culture.

The minimal nutritional requirements of Escherichia coli have been quantitatively determined in batch culture for cells with (R+) and without (R-) the R plasmid RP1. In these conditions R+ cells have a greater requirement than R- for several nutrients, particularly Mg2+, K+, Fe2+ and PO43-. The maximum growth rate in a simple salts medium was the same for R+ and R- cells. At low concentrations of phosphate, the specific growth rate of R+ cells differed from that found for R- cells. The R plasmid was stable in simple salts medium, irrespective of the nutrient ultimately depleted by growth, but, on storage, R+ cells survived for a shorter time than R- cells.     

Effect of thyroid hormone receptors in normal and cancerous cells on the ionic conductivity of bilayer phospholipid membranes

The effect of thyroid hormones receptors isolated from normal and cancer cells on bilayer phospholipid membranes (BPhLM) conductivity, has been studied. The receptor isolated from normal cells in complex X with the hormone selectively induces H+-conductivity of BPhLM generating transmembrane potential equal to 42 mV on the membrane at pH gradient equal to 1. In the presence of K+, Na+, Ca+, Mn2+, Sr2+, Mg2+ the changes of BPhLM are not observed. Neither hormones (T3, T4) nor receptor in free position affect the BPhLM conductivity. Thyroid hormone receptor isolated from mamalignantly transformed cells in a complex with T3 or T4 increases the BPhLM permeability for Ca2+. The transmembrane potential measured at 10fold Ca2+ ion concentration is equal to 16 mV. In the presence of H+, K+, Na+, Mn2+, Sr2+, Mg2+, Ba2+, the resistance of BPhLM doesn't change.     

Cytochemical localization of calcium and Ca2+,Mg2(+)-adenosine triphosphatase in colchicine-altered rat incisor ameloblasts

Colchicine is known to affect secretory, transport, and degradative functions of ameloblasts. The effects of colchicine on membrane-associated calcium and Ca2+,Mg2(+)-ATPase in secretory and maturation ameloblasts were investigated cytochemically. The pyroantimonate (PPA) method was used for localizing calcium and a modified Wachstein-Meisel medium was used to localize Ca2+,Mg2(+)-ATPase. Sections representing secretory and early maturation stages were examined by transmission electron microscopy. Morphological changes induced by colchicine included dislocated organelles and other well-established reactions to such anti-microtubule drugs. Calcium pyroantimonate (Ca-PA) deposits in most ameloblast types were markedly reduced, with the greater reduction occurring in those cells more severely altered morphologically. However, the cell membranes of both control and experimental smooth-ended maturation ameloblasts were essentially devoid of Ca-PA. The normal distribution and intensity of Ca2+,Mg2(+)-ATPase was not affected by colchicine. Because the observed reduction of membrane-associated calcium is apparently not mediated by Ca2+,Mg2(+)-ATPase in this case, other aspects of the calcium regulating system of ameloblasts are apparently targeted by colchicine.     

Evaluation of a role for intracellular Na+, K+, Ca2+, and Mg2+ in hyperthermic cell killing

A dose of heat which renders 98% of a population of Chinese hamster ovary cells reproductively dead has no significant effect on their Na+, K+, or Mg2+ content by 28 h postheat. In contrast, the cellular Ca2+ content increases in a dose-dependent manner as observed at 22 h after heating for 15-35 min at 45 degrees C. However, the rates of both influx and efflux of Ca2+ were reduced by heating. Increasing the cellular Ca2+ content by incubating the cells in high extracellular Ca2+, either at the time of heating or for a period of 22 h following heat, does not potentiate the lethal effect of heat. Completely blocking the heat-induced increase in Ca2+ content by incubating the cells in medium containing a low Ca2+ concentration does not protect the cells. Therefore, we conclude that heat does not produce any significant changes in the Na+, K+, or Mg2+ content of cells and that the heat-induced increase in Ca2+ does not play an important role in hyperthermic cell killing.     

Inhibition of red cell Ca2+-ATPase by vanadate

1. The Mg2+- plus Ca2+-dependent ATPase (Ca2+-ATPase) in human red cell membranes is susceptible to inhibition by low concentrations of vanadate. 2. Several natural activators of Ca2+-ATPase (Mg2+, K+, Na+ and calmodulin) modify inhibition by increasing the apparent affinity of the enzyme for vanadate. 3. Among the ligands tests, K+, in combination with Mg2+, had the most pronounced effect on inhibition by vanadate. 4. Under conditions optimal for inhibition of Ca2+-ATPase, the K 1/2 for vanadate was 1.5 microM and inhibition was nearly complete at saturating vanadate concentrations. 5. There are similarities between the kinetics of inhibition of red cell Ca2+-ATPase and (Na+ + K+)-ATPase prepared from a variety of sources; however, (Na+ + K+)-ATPase is approx. 3 times more sensitive to inhibition by vanadate.     

Bacterial strain characterizing by EDTA requirement

A novel strain of bacteria (LPM-4) was isolated that is characterized by a unique EDTA requirement for cell growth. Suspensions of washed cells of strain LPM-4 degrated EDTA complexes with Ba2+, Mg 2+, Ca2+, and Mn2+ at constant rates (0.310-0.486 mmol EDTA/(g h)) and Zn-EDTA at an initial rate of 0.137 +/- 0.016 mmol EDTA/(g h). The temperature optima for cell growth and EDTA degradation were determined under pH-auxostat cultivation. As compared with the known EDTA-degrating bacteria, strain LPM-4 exhibited a higher specific growth rate (0.095 h(-1)) and lower mass cell yield (0.219 g cells/g EDTA) that is promising for its practical applications for EDTA removal in wastewater treatment plants.     

Role of cations in the flocculation of Saccharomyces cerevisiae and discrimination of the corresponding proteins

Asexual yeast flocculation was studied using strong flocculents of Saccharomyces cerevisiae. The inhibitory effect of cations on flocculation is considered to be caused by competition between those cations and Ca2+ at the binding site of the Ca(2+)-requiring protein that is involved in flocculation. Inhibition of flocculation by various cations occurred in the following order: La3+, Sr2+, Ba2+, Mn2+, Al3+, and Na+. Cations such as Mg2+, Co2+, and K+ promoted flocculation. This promoting effect may be based on the reduction of electrostatic repulsive force between cells caused by binding of these cations anionic groups present on the cell surface. In flocculation induced by these cations, trace amounts of Ca2+ excreted on the cell surface may activate the corresponding protein. The ratio of Sr2+/Ca2+ below which cells flocculated varied among strains: for strains having the FLO5 gene, it was 400 to 500; for strains having the FLO1 gene, about 150; and for two alcohol yeast strains, 40 to 50. This suggests that there are several different types of cell surface proteins involved in flocculation in different yeast strains.     

Cation-chromatin binding as shown by ion microscopy is essential for the structural integrity of chromosomes.

Mammalian interphase and mitotic cells were analyzed for their cation composition using a three-dimensional high resolution scanning ion microprobe. This instrument maps the distribution of bound and unbound cations by secondary ion mass spectrometry (SIMS). SIMS analysis of cryofractured interphase and mitotic cells revealed a cell cycle dynamics of Ca2+, Mg2+, Na+, and K+. Direct analytical images showed that all four, but no other cations, were detected on mitotic chromosomes. SIMS measurements of the total cation content for diploid chromosomes imply that one Ca2+ binds to every 12.5-20 nucleotides and one Mg2+ to every 20-30 nucleotides. Only Ca2+ was enriched at the chromosomal DNA axis and colocalized with topoisomerase IIalpha (Topo II) and scaffold protein II (ScII). Cells depleted of Ca2+ and Mg2+ showed partially decondensed chromosomes and a loss of Topo II and ScII, but not hCAP-C and histones. The Ca2+-induced inhibition of Topo II catalytic activity and direct binding of Ca2+ to Topo II by a fluorescent filter-binding assay supports a regulatory role of Ca2+ during mitosis in promoting solely the structural function of Topo II. Our study directly implicates Ca2+, Mg2+, Na+, and K+ in higher order chromosome structure through electrostatic neutralization and a functional interaction with nonhistone proteins.     

Characterization of a High-Affinity Mg2+-Independent Ca2+-ATPase from Rat Brain Synaptosomal Membranes

A high-affinity Mg2+-independent Ca2+-ATPase (Ca2+-ATPase) has been differentiated from the Mg2+-dependent, Ca2+-stimulated ATPase (Ca2+,Mg2+-ATPase) in rat brain synaptosomal membranes. Using ATP as a substrate, the K0.5 of Ca2+ for Ca2+-ATPase was found to be 1.33 microM with a Km for ATP of 19 microM and a Vmax of 33 nmol/mg/min. Using Ca-ATP as a substrate, the Km for Ca-ATP was found to be 0.22 microM. Unlike Ca2+,Mg2+-ATPase, Ca2+-ATPase was not inhibited by N-ethylmaleimide, trifluoperazine, lanthanum, zinc, or vanadate. La3+ and Zn2+, in contrast, stimulated the enzyme activity. Unlike Ca2+, Mg2+-ATPase activity, ATP-dependent Ca2+ uptake was negligible in the absence of added Mg2+, indicating that the Ca2+ transport into synaptosomal endoplasmic reticulum may not be a function of the Ca2+-ATPase described. Ca2+-ATPase activity was not stimulated by the monovalent cations Na+ or K+. Ca2+, Mg2+-ATPase demonstrated a substrate preference for ATP and ADP, but not GTP, whereas Ca2+-ATPase hydrolyzed ATP and GTP, and to a lesser extent ADP. The results presented here suggest the high-affinity Mg2+-independent Ca2+-ATPase may be a separate form from Ca2+,Mg2+-ATPase. The capacity of Mg2+-independent Ca2+-ATPase to hydrolyze GTP suggests this protein may be involved in GTP-dependent activities within the cell.     

Calcium-magnesium interactions in pancreatic acinar cells.

Although the role of calcium (Ca2+) in the signal transduction and pathobiology of the exocrine pancreas is firmly established, the role of magnesium (Mg2+) remains unclear. We have characterized the intracellular distribution of Mg2+ in response to hormone stimulation in isolated mouse pancreatic acinar cells and studied the role of Mg2+ in modulating Ca2+ signaling using microspectrofluorometry and digital imaging of Ca2+- or Mg2+-sensitive fluorescent dyes as well as Mg2+-sensitive intracellular microelectrodes. Our results indicate that an increase in intracellular Mg2+ concentrations reduced the cholecystokinin (CCK) -induced Ca2+ oscillations by inhibiting the capacitive Ca2+ influx. An intracellular Ca2+ mobilization, on the other hand, was paralleled by a decrease in [Mg2+]i, which was reversible upon hormone withdrawal independent of the electrochemical gradients for Mg2+, Ca2+, Na+, and K+, and not caused by Mg2+ efflux from acinar cells. In an attempt to characterize possible Mg2+ stores that would explain the reversible, hormone-induced intracellular Mg2+ movements, we ruled out mitochondria or ATP as potential Mg2+ buffers and found that the CCK-induced [Mg2+]i decrease was initiated at the basolateral part of the acinar cells, where most of the endoplasmic reticulum (ER) is located, and progressed from there toward the apical pole of the acinar cells in an antiparallel fashion to Ca2+ waves. These experiments represent the first characterization of intracellular Mg2+ movements in the exocrine pancreas, provide evidence for possible Mg2+ stores in the ER, and indicate that the spatial and temporal distribution of intracellular Mg concentrations profoundly affects acinar cell Ca2+ signaling.     

北京西山不同人工林枯落物层的水化学性质

通过采集降雨经枯落物后的水样,初步研究了北京西山地区油松林和栓皮栎林林下枯落物层的水化学性质.结果表明:大气降水经过林冠进入枯落物层后,油松林和栓皮栎林林下不同元素的浓度发生明显变化.枯落物水中K 、Na 、Ca2 、Mg2 、NH4 -N和NO3--N的浓度随时间的变化趋势基本一致.穿透雨经过枯落物层后,水中K 、Na 、Ca2 和Mg2 的平均浓度增加,而NH4 -N、NO3--N的平均浓度减小.其中,栓皮栎林和油松林中Ca2 浓度分别增加了7.54和5.27mg.L-1.栓皮栎林下枯落物层中K 、Na 、Ca2 、Mg2 的平均浓度高于油松林,而NH4 -N、NO3--N的平均浓度则低于油松林;经降水淋溶作用后,栓皮栎林和油松林林下枯落物归还林地的养分分别为41.59和58.12kg.hm-2,其中归还林地较多的是Ca2 ,其次是K .     

Effects of MgCl2 on the release and recycling of heparan sulfate proteoglycans in a rat parathyroid cell line.

Divalent cations, such as Mg2+, Ba2+, and Co2+, are known to mimic the effects of Ca2+ in parathyroid cells, but it is not clear whether the mechanism of their action is the same as that of Ca2+. We have shown that extracellular Ca2+ concentration ([Ca2+]e) regulates the distribution and recycling of cell-surface heparan sulfate (HS) proteoglycans in a rat parathyroid cell line; at normal to high [Ca2+]e (e.g., 2 mM) HS proteoglycans are primarily localized intracellularly, while at low [Ca2+]e (0.05 mM) they are translocated to the cell surface and rapidly recycle (Takeuchi, Y., Sakaguchi, K., Yanagishita, M., Aurbach, G. D., and Hascall, V. C., 1990, J. Biol. Chem. 265, 13661-13668). We now show that a high concentration of Mg2+ (8 mM) reduces the amount of recycling HS proteoglycans in low [Ca2+]e. However, the primary effects of high Ca2+ and high Mg2+ on the recycling HS proteoglycans are different. High [Ca2+]e causes translocation of HS proteoglycans to intracellular compartments, while high Mg2+ stimulates cleavage of their core proteins and subsequent shedding of HS proteoglycans into the medium, thereby depleting the recycling molecules. However, high Mg2+ does not induce shedding of HS proteoglycans in high [Ca2+]e. The effects of Ba2+ and Co2+ were similar to those of Mg2+, but Sr2+ showed no significant effects on HS proteoglycan translocation. Otherwise, 8 mM Mg2+ did not alter biosynthesis or intracellular catabolism of HS proteoglycans. These observations suggest that the recycling of HS proteoglycans in parathyroid cells is sensitive only to [Ca2+]e, whereas several other divalent cations can deplete the recycling HS proteoglycans by a distinctly different mechanism. Thus, the mechanism by which Ca2+ regulates the amounts of the recycling HS proteoglycans may be more physiological and play a functional role in parathyroid cells.     

四种生态型芦苇叶中离子分布对生境的生理适应

采用X射线微区分析技术,测定了4种生态型芦苇(Phragmites australis (CaV.) Trin. exSteud.)叶的表皮泡状细胞、叶肉细胞和叶脉维管束鞘细胞离子的含量.结果表明:沼泽芦苇的鞘细胞内,K+、Na+、Ca2+、Mg2+和Cl-分布均较叶肉细胞和泡状细胞高.沙丘芦苇的泡状细胞中Ca2+分布较叶肉细胞和鞘细胞高,而Mg2+在其叶肉细胞,以及K+、Na+和Cl-在其鞘细胞内分布均较高.在轻度盐化草甸芦苇的叶肉细胞内分布较多的Na+和Mg2+,而在鞘细胞内K+、Ca2+ 和Cl-的分布均较叶肉细胞和泡状细胞为高.重度盐化草甸芦苇的泡状细胞内Na+和Mg2+的分布较多;同样,在叶肉细胞中K+、Ca2+和Cl-的分布也较多.最后,讨论了上述各种离子在不同生态型芦苇叶内分布的状况, 以及与其环境适应的生理意义.     

A high affinity Ca2+-stimulated and Mg2+-dependent ATPase in rat corpus luteum plasma membrane fractions

Plasma membrane fractions from rat corpus luteum contain two kinds of Ca2+-stimulated ATPase, one having a high affinity for Ca2+, the other a low affinity for Ca2+. The high affinity ATPase had a specific Ca2+ requirement with a K 1/2 of 0.2 to 0.3 microM; it had a Vmax of 105 nmol min-1 mg-1 and distributed, upon subcellular fractionation, with recognized plasma membrane enzymes. The properties of this enzyme indicate that it is a CA2+ extrusion pump. The low affinity pump (K 1/2 for Ca2+, about 15 microM) was nonspecific, being stimulated equally well by Ca2+ of Mg2+; its function is unknown. Although the high affinity ATPase resembled the erythrocyte Ca2+-pumping ATPase in the properties mentioned above, it differed in that it failed to respond to Mg2+ or calmodulin. The lack of response to Mg2+ was due to the enzyme's retention of endogenous Mg2+; it did, after incubation with chelators, show a Mg2+ requirement. However, we were unable to show any effect of added calmodulin or trifluoperazine. This failure may be related to the high content of tightly bound calmodulin in these membranes. Much of this calmodulin could not be extracted even by washing with 1 mM EGTA and/or 0.1% (w/v) Triton X-100. This enzyme, the erythrocyte enzyme, and the adipocyte plasma membrane Ca2+ ATPase all belong to the class of Ca2+ ATPases with plasma membrane distribution and high affinity for Ca2+, indicating that they are Ca2+ extrusion pumps. However, the data indicate that tissue-specific differences exist within this class, with the enzyme from adipocytes and rat corpus luteum belonging to a subclass in which the requirement for Mg2+ and any response to calmodulin are difficult to demonstrate.     

Regulation of Na+-K+-ATPase: effect of Mg and Ca ions

The participation of Mg2+ and Ca2+ in complicated mechanisms of Na+, K(+)-ATPase regulation is discussed in the survey. The regulatory actions of Mg2+ on Na+, K(+)-ATPase such as its participation in phosphorylation and dephosphorylation of the enzyme, ADP/ATP-exchange inhibition, cardiac glycosides and vanadate binding with the enzyme, conformational changes induction during ATPase cycle are reviewed in detail. Some current views of mechanisms of above mentioned Mg2+ regulatory effects are discussed. The experimental evidence of Ca2+ immediate influence on the functional activity of Na+, K(+)-ATPase (catalytic, transport and glycoside-binding) are given. It's noted that these effects are based on the conformational changes in the enzyme and also on the phase transition in membrane induced by Ca2+. Unimmediate action of Ca2+ on Na+, K(+)-ATPase is also discussed, especially due to its effect on other membrane systems functionally linked with Na(+)-pump (for instance, due to Na+/Ca(+)-exchanger activation). It's concluded that Mg2+ and Ca2+ as "universal regulators" of the cell effectively influence the functional activity and conformational states of Na+, K(+)-ATPase.