Dependence on neutral salt concentration of the latency phase in the time course of hydrolysis of dimyristoylphosphatidylcholine liposomes by phospholipase A2.

The time course of the hydrolytic action of porcine pancreatic phospholipase A2 on sonicated dimyristoylphosphatidylcholine liposomes in the presence of variable NaCl concentrations has been studied at temperatures between 17 and 36 degrees C; at these temperatures liposomes are in the gel phase. At a NaCl concentration of 10 mM, the hydrolysis shows a small and constant lag period of 6-8 min at all temperatures within this range. As the temperature is raised into the liquid crystalline range, the latency phase lengthens monotonically so that at 36 degrees C it reaches 55 min. An increase in the NaCl concentration to 1 M makes the lag period longer at all temperatures studied, with the exception of the phase transition range (near 24 degrees C); within this temperature range, a small reduction in the lag time is observed. The increase in the length of the latency period at high salt concentrations may be due to screening of the negative surface charge generated by the nascent fatty acid which seems to be essential for the efficient interfacial binding of the enzyme. In the phase transition range of the lamellae, the unfavorable effect of high salt concentrations on the electrostatic binding of the enzyme appears to be overcome by another type of interaction. Recent findings raise the possibility that this interaction could be hydrophobic in nature.     

Role of extracellular calcium in the hyperthermic killing of CHL V79 cells

Two major questions are addressed by this study: Can an influx of calcium ion sensitize CHL V79 cells to hyperthermia, and, if so, does this occur during heating and does it play a crucial role in cell death? V79 cells are sensitized to hyperthermia by the calcium ionophore A23187 which also induces an influx of calcium at both 37 and 43 degrees C. Sensitization is at least partially dependent on the presence of extracellular calcium. In the absence of A23187, survival is independent of calcium concentration (from 0 to 25 mM) during heating, which differs from the behavior of hepatocytes which are sensitized to hyperthermia by 15 mM CaCl2. Calcium influx, as assayed by uptake of 45Ca measured after washing in LaCl3, is detectable in 3 mM CaCl2 only after 30 min at 45 degrees C, an exposure which reduces reproductive survival to less than 0.1%. Calcium uptake reaches 6 nmol/10(6) cells after 180 min at 45 degrees C. This is not due to a general loss of membrane permeability since there is no trypan blue staining during this time. In 15 mM CaCl2, influx occurs earlier (15 min) but still succeeds the loss of reproductive survival which is less than 1% at this time. Uptake is much higher in 15 mM CaCl2, reaching 10 nmol/10(6) cells by 30 min and 25 nmol/10(6) cells at 180 min, but the temporal pattern of uptake does not correlate with loss of reproductive survival. Thus, although A23187 sensitizes V79 cells to hyperthermia, probably by increased influx of calcium ion, and increased influx occurs during exposure to 45 degrees C, influx is not a crucial early event in the killing of V79 cells. This does not eliminate the possibility of intracellular calcium redistribution during hyperthermia.     

Role of gangliosides in adhesion and conductance changes in large spherical model membranes

The formation of two spherical model membranes at the tips of two syringes has allowed us to study the role of gangliosides in membrane adhesion and look for changes in conductance between two such membranes during the process of adhesion. Membranes were formed in aqueous 100 mM NaCl, 10 mM KCl, 1 mM CaCl2 from 1% (w/v) egg phosphatidylcholine in n-decane, with or without mixed bovine brain gangliosides. After thinning to the 'black' bilayer state, two membranes were moved into contact. With gangliosides, the contact area and conductance increased colinearly with time over a 5 to 20 min period of adhesion. The role of electrostatic bridging by calcium was investigated. In the absence of calcium or in the presence of 2 mM EDTA, adhesion proceeded after a longer lag time at about one-half the normal rate. As the ganglioside concentration was increased from 0 to 15 mol%, the electrical conductance of individual membranes decreased 3-fold from 48 +/- 30 nS/cm2 to 17 +/- 13 nS/cm2. The conductance was pH dependent with a minimum at neutral values. At neutral pH, when two membranes containing 4.1 mol% gangliosides adhered, the region of adhesion had a specific conductance three times that of the nonadhering regions of membranes. Without gangliosides, the specific conductance of the contact region was the same as that of non-adhering regions of the membrane. These data suggest that mixed gangliosides can mediate an adhesion-dependent increase in conductance.     

On the solubility of calcium deoxycholate: kinetics of precipitation and the effect of conjugated bile salts and lecithin

In view of the low solubility of calcium deoxycholate and the possible induction of cholesterol precipitation in the gallbladder by calcium insoluble salts, we find it of interest to study the precipitation of calcium deoxycholate and its dependence on other bile components. The findings of these studies were as follows: (i) Precipitation of calcium deoxycholate from mixtures of calcium chloride and monomeric deoxycholate (at concentrations below the critical micelle concentration (CMC] is very slow even at relatively high CaCl2 concentrations (more than 20 days at 50 mM CaCl2). (ii) At higher deoxycholic acid (DOC) concentrations, precipitation of micellar DOC is faster and requires much lower calcium chloride concentrations. For any given calcium concentration, the rate of precipitation is maximal at an optimal DOC concentration. In solutions containing 150 mM NaCl, the maximal rate of precipitation occurs at about 10 mM DOC, almost independent of Ca2+ concentration. At lower ionic strength (10 mM NaCl), the optimal DOC concentration is 30 mM. These observations suggest that the most important factors in determining the rate of Ca(DOC)2 precipitation are (a) the ratio between calcium ions bound to the surface of a DOC micelle, and the [DOC] (the Ca2+/DOC binding ratio) and (b) the concentration of DOC micelles. (iii) In the presence of conjugated deoxycholates, the crystallization of calcium deoxycholate is inhibited. Phosphatidylcholine has a similar, although smaller, inhibitory effect. Upon precipitation of calcium deoxycholate from a mixed micellar system containing sodium deoxycholate, phosphatidylcholine and cholesterol, the latter two components spontaneously form vesicles. The anti-nucleating effect of PC and conjugated bile salts is explained in terms of "poisoning" of the crystallization process. In view of the latter results we conclude that under normal conditions calcium deoxycholate is not likely to precipitate in the gallbladder.     

Cation-induced aggregation of acidic phospholipid vesicles: the role of fatty acid unsaturation and cholesterol

Cation-induced aggregation of acidic phospholipid vesicles consisting of dimyristoylphosphatidylglycerol (DMPG), dipalmitoylphosphatidylserine (DPPS), phosphatidylserine from bovine brain (brPS), and phosphatidylglycerol from egg yolk (eggPG) was studied. Significant differences were evident in the NaCl-induced aggregation of fully saturated and unsaturated acidic phospholipid vesicles. The threshold NaCl concentration of vesicle aggregation ([NaCl]Thr) for DPPS vesicles was 320 mM compared to 610 mM observed for brPS vesicles. For DMPG vesicles the [NaCl]Thr was 430 mM and no aggregation of eggPG vesicles could be observed upon addition of NaCl. The threshold CaCl2 concentrations of aggregation of DMPG and eggPG vesicles were 2.3 and 4.9 mM, respectively. The corresponding threshold CaCl2 concentrations for DPPS and brPS vesicles were 0.85 mM and 1.3 mM, respectively. The inclusion of cholesterol into vesicles attenuated NaCl- and CaCl2-induced aggregation of DMPG and DPPS vesicles. However, enhancement of aggregation by inclusion of cholesterol was observed in the case of NaCl-induced aggregation of brPS vesicles. It is concluded that cation mediated membrane-membrane interactions depend, in addition to polar headgroup structure, on the fatty acid composition of the phospholipids also.     

Na+-induced structural change of a soil bacterium, S34, and Ca2+ requirement for preserving its original structure.

A drastic change in the outer membrane structure of a salt-sensitive soil bacterium, S34, related to the genus Deinococcus was induced by 0.2 to 0.4% (wt/vol) NaCl. The change was relieved by 6 mM CaCl2 and induced by 1 mM EGTA. The results indicate the strong dependence of the organism on calcium.     

Change in the activity of calcium ions during illumination of a suspension of visual cell outer segment fragments]

Changes in the activity of calcium ions in the medium containing outer fragments suspension of bovine eye retina rods have been studied by the method of calcium-selective electrodes. Illumination of the suspension increases calcium ion activity in the incubation medium. Photoinduced yield of calcium ions depends on Ca+2 concentration: it equals 0.11+/-0.015 M Ca2+/1m rodopsin in the medium containing 0.1 mM CaCl2 and 0.046+/-0.002Ca2+/1M rodopsin in the medium containing 0.05 mM CaCl2. In the medium containing more than 10(-4) M CaCl2 both an increase and a decrease of Ca2+ ions have been observed.     

Pulse NMR study of the interaction of calcium ion with dipalmitoylphosphatidylcholine lamellae

Molecular motion of dipalmitoylphosphatidylcholine (DPPC)/CaCl2 lamellae in a gel phase was studied by pulse NMR. Proton 1/T1 for DPPC in a gel phase showed that the rate of reorientation about the long axis of the lipid molecule decreased gradually from 0 to 500 mM CaCl2. At 10-50 mM CaCl2 the correlation time reached the value of the inverse Larmor frequency (approx. 2.6 ns). A proton NMR absorption spectrum and a spin-pair-dipolar-echo (SPDE) decay showed that the second moment in the hydrocarbon chain region decreased below about 1 mM CaCl2 and increased from 1 to 500 mM CaCl2. The second moment in the polar head group increased gradually with an increase in the CaCl2 concentration. The increase in the second moment at the high CaCl2 concentrations was attributed to an increase in the order parameters of the segments both in the polar head group and in the hydrocarbon chain region. At the lower CaCl2 concentrations, however, calcium ion possibly induced disorder in the lamellae which led to a decrease in the order parameter in the hydrocarbon chain region.     

Role of calcium in stabilizing the structure of hyalin, a major protein component of the sea urchin extraembryonic hyaline layer

The interactions of NaCl and CaCl2 with the sea urchin embryo coat protein hyalin were investigated. Endogenous protein tryptophan fluorescence was enhanced by almost 45% in the presence of 200mM NaCl while 1mM CaCl2 reversed this effect and brought the intensity of fluorescence back close to that of the native protein. Half-maximal concentrations of 53 and 0.32mM were determined for NaCl and Ca+2, respectively. Hyalin conformation, as measured by circular dichroic spectroscopy, was altered by NaCl and CaCl2 in a fashion parallel to the effects of these salts on tryptophan fluorescence. Sodium chloride disrupted hyalin secondary structure while CaCl2 affected the return of hyalin to its native conformation. The interactions of NaCl and CaCl2 with hyalin were not modulated by MgCl2. These results suggest a role for CaCl2 in stabilizing hyalin against the disruptive effects of the high concentration of NaCl present in sea water.     

Calcium specificity of the antigen-induced channels in rat basophilic leukemia cells

Ion channels, activated upon IgE-Fc epsilon receptor aggregation by specific antigen, were studied in micropipet-supported lipid bilayers. These bilayers were reconstituted with purified IgE-Fc epsilon receptor complex and the intact 110-kDa channel-forming protein, both isolated from plasma membranes of rat basophilic leukemia cells (line RBL-2H3). In order to identify the current carrier through these ion channels and to determine their ion selectivity, we investigated the currents flowing through the IgE-Fc epsilon receptor gated channels in the presence of a gradient of Ca2+ ions. Thus, the solution in which the micropipet-supported bilayer was immersed contained 1.8 mM CaCl2, while the interior of the micropipet contained 0.1 microM Ca2+ (buffered with EGTA). Both solutions also contained 150 mM of a monovalent cation chloride salt (either K+ or Na+). The currents induced upon specific aggregation of the IgE (by either antigen or anti-IgE antibodies) were examined over a range of potentials imposed on the bilayer. The type of conductance event most frequently observed under the employed experimental conditions was a channel that has a slope conductance of 3 pS and a reversal potential practically identical with the calculated value for the reversal potential of calcium (134 +/- 11 mV in the presence of sodium, 125 +/- 13 mV in the presence of potassium). These results indicate that this channel is highly selective for calcium against the monovalent cations sodium and potassium. This same channel has a conductance of 4-5 pS in the presence of symmetrical solutions containing only 100 mM CaCl2 and 8 pS in the presence of 0.5 M NaCl with no calcium.(ABSTRACT TRUNCATED AT 250 WORDS)     

Modulation of intestinal absorption of calcium]

1. Absorption of ingested calcium (2 ml of a 10mM CaCl2 solution + 45Ca) by the adult rat was shown to be facilitated by the simultaneous ingestion of an active carbohydrate, L-arabinose. As the carbohydrate concentration is increased from 10 to 200 mM, the adsorption of calcium is maximized at a level corresponding to about twice the control adsorption level. 2. A similar doubling of calcium adsorption is obtained when a 100 mM concentration of any one of a number of other carbohydrates (gluconic acid, mannose, glucosamine, sorbitol, lactose, raffinose, stachyose) is ingested simultaneously with a 10 mM CaCl2 solution. 3. Conversely, the simultaneous ingestion of increasing doses (10 to 100 mM) of phosphate (NaH2PO4) with a 10 mM CaCl2 solution results in decreased 45Ca absorption and retention by the adult rat. 4. The maximum inhibition of calcium adsorption by phosphate is independent of the concentration of the ingested calcium solution (from 5 to 50 mM CaCl2). 5. The simultaneous ingestion of CaCl2 (10 mM) with lactose and sodium phosphate (50 and 10 mM, respectively) shows that the activating effect of lactose upon 45Ca adsorption may be partly dissimulated by the presence of phosphate. 6. These various observations indicate that, within a large concentration range (2 to 50 mM CaCl2), calcium adsorption appears to be a precisely modulated diffusion process. Calcium absorption varies (between minimum and maximum levels) as a function of the state of saturation by the activators (carbohydrates) and inhibitors (phosphate) of the calcium transport system.     

Two different effects of calcium on aquaporins in salinity-stressed pepper plants

Two different effects of calcium were studied, respectively, in plasma membrane vesicles and in protoplasts isolated from roots of control pepper plants (Capsicum annuum L cv. California) or of plants treated with 50 mM NaCl, 10 mM CaCl(2) or 10 mM CaCl(2) + 50 mM NaCl. Under saline conditions, osmotic water permeability (P ( f )) values decreased in protoplasts and plasma membrane vesicles, and the same reduction was observed in the PIP1 aquaporin abundance, indicating inhibitory effects of NaCl on aquaporin functionality and protein abundance. The cytosolic Ca(2+) concentration, [Ca(2+)](cyt), was reduced by salinity, as observed by confocal microscope analysis. Two different actions of Ca(2+) were observed. On the one hand, increase in free cytosolic calcium concentrations associated with stress perception may lead to aquaporin closure. On the other hand, when critical requirements of Ca(2+) were reduced (by salinity), and extra-calcium would lead to an upregulation of aquaporins, indicating that a positive role of calcium at whole plant level combined with an inhibitory mechanism at aquaporin level may work in the regulation of pepper root water transport under salt stress. However, a link between these observations and other cell signalling in relation to water channel gating remains to be established.     

Affinity-repulsion chromatography. Principle and application to lectins

The interactions of proteins with their immobilized ligands in an electrically charged microenvironment were studied. The binding of lectins to erythrocytes and to affinity matrices was used as a model system. Lectins bind and agglutinate erythrocytes in the presence of at least 10 mM NaCl or 1 mM CaCl2, but not in deionized water. The salt dependence of the agglutination process is due to the ability of salts to provide counterions neutralizing the forces of repulsion between the electrostatic charges of similar sign present on the erythrocyte cell surface and on the lectins. The same salt dependence is observed for the binding of lectins to affinity matrices. These observations are the basis of a protein separation process coined affinity-repulsion chromatography in which the electrostatic charges present, or purposely introduced, on affinity matrices are exploited and allow the elution, by electrostatic repulsion, of proteins carrying electrostatic charges of the same sign as that of the matrix. In this process, proteins are loaded on the affinity matrix in a salt solution and eluted with deionized water. Affinity-repulsion chromatography has been successfully applied here to the isolation of several lectins. Its physicochemical basis, merits, and potential applications are discussed.     

Mechanism of enhancement of the responses of the frog glossopharyngeal nerve to electrolytes by enhancers

In frogs, the responses of the glossopharyngeal nerve (GL) to NaCl are enhanced after treatment of the tongue with 8-anilino-1-naphthalene-sulfonic acid (ANS), a hydrophobic probe for biological membranes. The enhancement by ANS treatment has been explained by removal of Ca2+ from the receptor membrane treated with ANS. To explore the mechanism of enhancement by ANS treatment, we recorded neural responses from the frog GL. After ANS treatment, treatment with 10 mM CaCl2 prior to stimulation of NaCl did not affect the enhanced responses to 100 mM NaCl. The response to a relatively high concentration of CaCl2 (50 mM) was enhanced after ANS treatment. It is difficult to interpret these neural events in terms of modulation of the responses by membrane-bound calcium. The presence of NiCl2 in stimulating solution is known as an enhancer. Neural events after ANS treatment were similar to those caused by NiCl2. Our previous studies have demonstrated that enhancement of the responses to electrolytes by NiCl2 is due to modulation of the responses of water fibers in the GL. Water fibers are characterized by sensitivity to water or CaCl2, and they also respond to relatively high concentrations of electrolytes such as NaCl and choline Cl. Using a suction electrode method, we recorded unitary impulses from single water fibers. The ANS treatment led greatly enhanced responses to NaCl or choline Cl in water fibers, suggesting that enhancement by the ANS treatment is due to modulation of the responses of water fibers as well as enhancement by NiCl2. It appears that distinct receptors for each separate cation responsible for the neural responses in water fibers interact with a membrane element that is affected by ANS or Ni2+.     

Equilibrium and kinetic studies of oxygen binding to the haemocyanin from the freshwater snail Lymnaea stagnalis.

The binding of oxygen by the haemocyanin of the gastropod Lymnaea stagnalis was studied by equilibrium and kinetic methods. The studies were performed under conditions in which the haemocyanin molecule was in the native state. Over the pH range 6.8-7.6, in the presence of 10mM-CaCl2 the haemocyanin bound O2 cooperatively. Over this pH range the haemocyanin molecule displayed a normal Bohr effect whereby the O2 affinity of the molecule decreased with a fall in the pH of the solution. The maximum slope of the Hill plot (hmax.) was 3.5, obtained at pH 7.5. An increase in the CaCl2 concentration from 5 to 20 mM at pH 6.8 resulted in a slight increase in the oxygen affinity, with hmax. remaining virtually unchanged. At constant pH and CaCl2 concentration, an increase in NaCl concentration from 0 to 50 mM resulted in a small decrease in O2 affinity, but a significant increase in the value of hmax. from 3.5 to 8.6. Temperature-jump relaxation experiments over a range of O2 concentrations produced single relaxation times. The dependence of the relaxation time on the reactant concentrations indicated a simple bimolecular binding process. The calculated association and dissociation rate constants for this process at pH 7.5 are 29.5 X 10(6) M-1 X S-1 and 49 S-1 respectively. The association rate constant kon was found to be essentially independent of pH and CaCl2 concentration. The dissociation rate constant, koff, however, increased with a decrease in the pH, but was also independent of CaCl2 concentration. These results indicate that the stability of the haemocyanin-O2 complex is determined by the dissociation rate constant.     

The role of calcium ions during mitosis

Calcium-containing solutions were microinjected into dividing PtK1 cells to assess the effect of calcium ion concentration on the morphology and physiology of the mitotic spindle. Solutions containing 50 microM or more CaCl2 are immediately and irreversibly toxic to PtK1 cells. Those containing 5-10 microM CaCl2 cause reversible reduction in spindle birefringence followed by normal anaphase and cytokinesis. Microinjection of 5 microM or less CaCl2 into anaphase PtK1 cells has no detectable effect on the rate or extent of chromosome movement. Metaphase cells tend to enter anaphase 4-5 min after injection with 1-10 microM CaCl2, compared with an average of 16 min after injection with calcium-free buffer. Reducing the intracellular calcium concentration by injection of EGTA-CaCl2 buffers increases the lag between injection and anaphase to 20 min or more. Microinjection of calcium solutions does not promote precocious chromatid separation in nocodazole-arrested metaphase cells, indicating that the increase in calcium concentration does not induce centromere separation directly. An increase in the concentration of free calcium ions during metaphase appears to stimulate the onset of anaphase. Such an increase, regulated by the cell itself, may contribute to the initiation of chromosome separation in mammalian cells.     

Influence of the NaCl or CaCl2 concentration on the structure of heat-set bovine serum albumin gels at pH 7

The structure of heat-set systems of the globular protein bovine serum albumin (BSA) was investigated at pH 7 in different salt conditions (NaCl or CaCl(2)) using light scattering. Cross-correlation dynamic light scattering was used to correct for multiple scattering from turbid samples. After heat treatment, aggregates are formed whose size increases as the protein concentration increases. Beyond a critical concentration that decreases with increasing salt concentration, gels are formed. The heterogeneity and the reduced turbidity of the gels were found to increase with increasing salt concentration and to decrease with increasing protein concentration. The structure of the gels is determined by the strength of the repulsive electrostatic interactions between the aggregated proteins. The results obtained in NaCl are similar to those reported in previous studies for other globular proteins. CaCl(2) was found to be much more efficient in reducing electrostatic interactions than NaCl at the same ionic strength.     

Taste transduction mechanism: similar effects of various modifications of gustatory receptors on neural responses to chemical and electrical stimulation in the frog

Responses in the frog glossopharyngeal nerve induced by electrical stimulation of the tongue were compared with those induced by chemical stimuli under various conditions. (a) Anodal stimulation induced much larger responses than cathodal stimulation, and anodal stimulation of the tongue adapted to 5 mM MgCl2 produced much larger responses than stimulation with the tongue adapted to 10 mM NaCl at equal current intensities, as chemical stimulation with MgCl2 produced much larger responses than stimulation with NaCl at equal concentration. (b) The enhansive and suppressive effects of 8-anilino-1-naphthalenesulfonate, NiCl2, and uranyl acetate on the responses to anodal current were similar to those on the responses to chemical stimulation. (c) Anodal stimulation of the tongue adapted to 50 mM CaCl2 resulted in a large response, whereas application of 1 M CaCl2 to the tongue adapted to 50 mM CaCl2 produced only a small response. This, together with theoretical considerations, suggested that the accumulation of salts on the tongue surface is not the cause of the generation of the response to anodal current. (d) Cathodal current suppressed the responses induced by 1 mM CaCl2, 0.3 M ethanol, and distilled water. (e) The addition of EGTA or Ca-channel blockers (CdCl2 and verapamil) to the perfusing solution of the lingual artery reversibly suppressed both the responses to chemical stimulus (NaCl) and to anodal current with 10 mM NaCl. (f) We assume from the results obtained that electrical current from the microvillus membrane of a taste cell to the synaptic area supplied by anodal stimulation or induced by chemical stimulation activates the voltage-dependent Ca channel at the synaptic area.     

New evidence about the relationship between water channel activity and calcium in salinity-stressed pepper plants

This study, of how Ca2+ availability (intracellular, extracellular or linked to the membrane) influences the functionality of aquaporins of pepper (Capsicum annuum L.) plants grown under salinity stress, was carried out in plants treated with NaCl (50 mM), CaCl2 (10 mM), and CaCl2 (10 mM) + NaCl (50 mM). For this, water transport through the plasma membrane of isolated protoplasts, and the involvement of aquaporins and calcium (extracellular, intracellular and linked to the membrane) has been determined. After these treatments, it could be seen that the calcium concentration was reduced in the apoplast, in the cells and on the plasma membrane of roots of pepper plants grown under saline conditions; these concentrations were increased or restored when extra calcium was added to the nutrient solution. Protoplasts extracted from plants grown under Ca2+ starvation showed no aquaporin functionality. However, for the protoplasts to which calcium was added, an increase of aquaporin functionality of the plasma membrane was observed [osmotic water permeability (Pf) inhibition after Hg addition]. Interestingly, when verapamil (a Ca2+ channel blocker) was added, no functionality was observed, even when Ca2+ was added with verapamil. Therefore, calcium seems to be involved in plasma membrane aquaporin regulation via a chain of processes within the cell but not by alteration of the stability of the plasma membrane.     

A neutron diffraction study of the influence of ions on phospholipid membrane interactions.

Neutron diffraction is used to examine the effects of Ca2+ and ClO4- ions on interactions and some structural features of dipalmitoylphosphatidylcholine membranes in both solid and fluid lamellar phases. The results are described within the framework of Derjaguin-Landau-Verwey-Overbeek (DLVO) theory with reference to electrostatic, van der Waals, and hydration components of disjoining pressure. The Hamaker constants are evaluated under equilibrium conditions. Addition of 100 mM CaCl2 to the aqueous phase substantially increases the lamellar repeat spacing (d), which is interpreted in terms of adsorption of Ca2+ ions to bilayers followed by electrostatic repulsion between membranes. The rise of NaClO4 concentration in the presence of 100 mM CaCl2 leads to gradual decrease in d, evidently resulted from the diminution of Ca(2+)-induced positive surface potential by both electrostatic screening and binding of ClO4- ions. In the absence of CaCl2, elevation of NaClO4 concentration to 100-300 mM drastically enhances the repeat spacing and then dramatically decreases d at about 1 M NaClO4. Estimation of the hydration coefficients showed that the pronounced decrease of the repeat spacing at high NaClO4 concentrations was resulted mainly from the (partial) disruption of the structure of intermembrane bound water by chaotropic ClO4- ions and subsequent decrease in hydration repulsive pressure. Moreover, in the case of solid membranes (20 degrees C) high concentrations of ClO4- induced formation of interdigitated phase paralleled with marked reduction in bilayer thickness and corresponding increase in the effective cross-sectional area per lipid molecule.