Effects of prostaglandin E1 on contractility and Ca release in rabbit aortic smooth muscle

Concentrations of prostaglandin E₁ (PGE₁; 10⁻⁷ M) that do not elicit tension responses in aortic strips potentiate contractions induced by submaximal concentrations (10⁻⁸ − 10⁻⁷ M) of norepinephrine (NE) or angiotensin III (Ang III) but not those of high K⁺ depolarization or maximal NE or Ang III concentrations. Higher concentrations of PGE₁ (10⁻⁶ M and above) initiate contractions which are additive with submaximal responses to NE and Ang III but not to K⁺. These same concentrations of PGE₁ also decrease ⁴⁵Ca retention at high affinity La⁺⁺⁺-resistant sites in a manner similar to but not additive with NE and Ang III. Uptake of ⁴⁵Ca at low affinity La⁺⁺⁺-resistant sites (which is increased by high K⁺-depolarization) is not altered by 10⁻⁶ M PGE₁. The effects of PGE₁ are not altered by decreased extracellular Ca⁺⁺ (0.1 mM), decreased temperature, phentolamine or meclofenamate. Thus, PGE₁ does not appear to increase uptake of extracellular Ca⁺⁺ in this smooth muscle tissue. Instead, PGE₁ increases mobilization of Ca⁺⁺ from the same high affinity La⁺⁺⁺-resistant sites affected by Ang III and NE and, in this manner, may increase responses to these two stimulatory agents.     

Role of external calcium in sodium and chloride transport in the gills of seawater-adaptedMugil capito

Summary When the mulletMugil capito is transferred to medium lacking Ca⁺⁺ (either Ca⁺⁺-free seawater or distilled water) the passive permeability of the gill to Na⁺ and Cl^– is increased and the activating effect of external K⁺ on the Na⁺ and Cl^– effluxes in hyposaline media is inhibited. The permeability of the gill increases progressively in proportion to the time of Ca⁺⁺ deprivation; it declines when Ca⁺⁺ is added again to the external medium. The active mechanisms for ion excretion are not reversible. At external Ca⁺⁺ concentrations from 0.1 to 10 mM the Na⁺ permeability is constant but the activation of Na⁺ efflux by K⁺ shows a maximum at a Ca⁺⁺ concentration of about 1 mM. For activation of Cl^– efflux external bicarbonate must be present, in addition to Ca⁺⁺, suggesting the existence of a Cl^–/HCO ₃ ^– exchange. The mechanism by which Ca⁺⁺ controls the passive branchial permeability is thus probably different from that involved in K⁺ activation of ion excretion. The Ca⁺⁺ effect on the K⁺ sensitive ionic excretory mechanisms seems to be related to intracellular Ca⁺⁺ movements. Thus, on the one hand, substances such as Ruthenium Red and La⁺⁺⁺ which both inhibit Ca⁺⁺ exchange, in media containing Ca⁺⁺ and HCO ₃ ^– also inhibit K⁺ activation of Na⁺ and Cl^– effluxes; on the other hand, the ionophore A 23187, a stimulator of Ca⁺⁺ exchange, when added to these media, activates the Na⁺ and Cl^– effluxes; its maximal effect on the Na⁺ flux occurs at 2 mM Ca⁺⁺.Abbreviations ASW-Ca artificial seawater minus calcium - DW deionised water - DWCa deionised water with 1 mM Ca⁺⁺ added - DWCaHCO ₃ DW with calcium plus bicarbonate - DWHCO ₃ DW with 1 mM sodium bicarbonate added - FW freshwater (tap water) - FWK freshwater with K⁺ added - P. D. potential difference - SW seawater The experiments reported in this paper were done with Jean Maetz who tragically died in August 1977. It is the last report about several years of friendly collaboration     

Triple helix formation and disulphide bonding during the biosynthesis of glomerular basement membrane collagen.

White erythrocyte membranes, or ghosts, were monoconcave discocytes when incubated in 50mM N-tris (hydroxymethyl) methyl-2-aminoethane sulfonic acid titrated to pH 7.4 with triethanolamine. If 3mM MgCl₂ was included in the incubation medium, the ghosts were predominantly echinocytes. The echinocytic form could also be induced by Co⁺⁺, Ni⁺⁺, Li⁺, Na⁺, K⁺, $\text{NH}{}_4{}^{\mathrm{+}}$ and tetramethylammonium ion, all as chloride salts. The concentration of cation necessary for 50% of the ghosts to be echinocytes was correlated with the hydrated charge density of the cation with the most highly charged cations being the most effective. The cations Ca⁺⁺, Sr⁺⁺, Ba⁺⁺ and La⁺⁺⁺, (also as chloride salts) did not induce the normal echinocytic form, but at high levels induced a few misshapen forms with some resemblance to echinocytes. Instead Ca⁺⁺, Sr⁺⁺, Ba⁺⁺ and La⁺⁺⁺ suppressed the formation of echinocytes in the presence of Mg⁺⁺ and other ions. This suggests the presence of a specific Ca⁺⁺ binding site important to shape control in the erythrocyte membrane.     

Lanthanum-induced alterations of cellular electrolytes in Ehrlich ascites tumor cells: a new view

We have shown previously that Ehrlich ascites tumor cells maintained at room temperature under an oxygen atmosphere lose Na⁺, K⁺ and Cl^? isosmotically when exposed to La⁺⁺⁺ (0.1 to 1.0 mM). Concomitant with these changes there is an increase in the recorded membrane potential (increasing intracellular negativity). The present studies further characterize the effect of La⁺⁺⁺ on electrolyte distribution. Ehrlich ascites tumor cells were maintained at 0.5° C to permit Na⁺ gain and K⁺ loss. The addition of 1 mM La⁺⁺⁺ to low temperature cells induces rapid loss of Na⁺, K⁺ and Cl^?. This net loss of cellular electrolytes occurs even in cells depleted of ATP content using 2-deoxyglucose (5 mM) and rotenone (10^?6 M ). Analysis of the appearance of tracer ²²Na in the environment of cells preloaded with the radioisotope shows that La⁺⁺⁺-induced changes in membrane permeability or in active ion transport mechanisms are not responsible for the dramatic loss of electrolytes from experimental cells. The electrolyte loss occurs only when the cells are resuspended mechanically during the washing procedure used to prepare the cells for electrolyte determination. We conclude that the results of La⁺⁺⁺ interaction with Ehrlich ascites tumor cells are twofold. As we have previously reported, La⁺⁺⁺ stabilizes and causes a hyperpolarization of the membrane potential. Secondly, La⁺⁺⁺ predisposes the cell membrane to become highly permeable when subjected to mechanical stress.     

COMBINATION OF SALTS AND PROTEINS : III. THE COMBINATION OF CuCl2, MgCl2, CaCl2, AlCl3, LaCl3, KCl,AgNO3AND Na2SO4WITH GELATIN.

1. The combination of Cu⁺⁺, Ca⁺⁺, Mg⁺⁺, Al⁺⁺⁺, La⁺⁺⁺, K⁺, Ag⁺, and Cl^- with gelatin has been determined. 2. The equivalent combining value for copper is about 0.9 millimols per gm. of gelatin and is therefore the same as that of hydrogen. The value for copper with deaminized gelatin is about 0.4 to 0.5, again the same as that of hydrogen. The sum of the hydrogen and copper ions combined in the presence of an excess of either is 0.9 millimols showing that there is an equilibrium between the copper hydrogen and gelatin and that the copper and hydrogen are attached to the same group. 3. The equivalent combining value of La⁺⁺⁺ and Al⁺⁺⁺ is about 0.5 millimols per gm. of gelatin. This value is not significantly different with deaminized gelatin so that it is possible these salts combine only with groups not affected by deaminization. 4. No calcium is combined on the acid side of pH 3. The value rises rapidly from pH 3 to 4.7 and then remains constant. 5. No combination of K, Li, Na, NO₃ or SO₄ could be detected. 6. Cl combines less than the di- and trivalent metals so that the protein is positive in CaCl₂ but negative in KCl.     

Lanthanum in heart cell culture. Effect on calcium exchange correlated with its localization

Correlation of the localization of La⁺⁺⁺ with its effects on Ca⁺⁺ exchange in cultured rat heart cells is examined with the use of a recently developed technique. 75% of cellular Ca⁺⁺ is exchangeable and is completely accounted for by two kinetically defined phases. The rapidly exchangeable phase has a t ½ = 1.15 min and accounts for 1 1 mmoles Ca⁺⁺/kg wet cells or 43% of the exchangeable Ca⁺⁺ (cells perfused with [Ca⁺⁺]_o = 1 mM) Phase 2 has a t ½ = 19.2 min and accounts for 1.5 mmoles Ca⁺⁺/kg wet cells or 57% of the exchangeable Ca⁺⁺. 0.5 mM [La⁺⁺⁺]_o displaces 0 52 mmoles Ca⁺⁺/kg wet cells—all from phase 1—and almost completely abolishes subsequent Ca⁺⁺ influx and efflux The presence of La⁺⁺⁺ in the washout converts the washout pattern to a single phase system with a t ½ = 124 min. The effects upon Ca⁺⁺ exchange are coincident with abolition of contractile tension but regenerative depolarization of the tissue is maintained Electron microscope localization of the La⁺⁺⁺ places it exclusively in the external lamina or basement membrane of the cells. The study indicates that negatively charged sites in the basement membrane play a crucial role in the E-C coupling process in heart muscle     

Prostaglandin E2 modulates Na+,K+-ATPase activity in rat hippocampus: implications for neurological diseases

Prostaglandin E₂ (PGE₂) is quantitatively one of the major prostaglandins synthesized in mammalian brain, and there is evidence that it facilitates seizures and neuronal death. However, little is known about the molecular mechanisms involved in such excitatory effects. Na⁺,K⁺‐ATPase is a membrane protein which plays a key role in electrolyte homeostasis maintenance and, therefore, regulates neuronal excitability. In this study, we tested the hypothesis that PGE₂ decreases Na⁺,K⁺‐ATPase activity, in order to shed some light on the mechanisms underlying the excitatory action of PGE₂. Na⁺,K⁺‐ATPase activity was determined by assessing ouabain‐sensitive ATP hydrolysis. We found that incubation of adult rat hippocampal slices with PGE₂ (0.1–10 μM) for 30 min decreased Na⁺,K⁺‐ATPase activity in a concentration‐dependent manner. However, PGE₂ did not alter Na⁺,K⁺‐ATPase activity if added to hippocampal homogenates. The inhibitory effect of PGE₂ on Na⁺,K⁺‐ATPase activity was not related to a decrease in the total or plasma membrane immunocontent of the catalytic α subunit of Na⁺,K⁺‐ATPase. We found that the inhibitory effect of PGE₂ (1 μM) on Na⁺,K⁺‐ATPase activity was receptor‐mediated, as incubation with selective antagonists for EP1 (SC‐19220, 10 μM), EP3 (L‐826266, 1 μM) or EP4 (L‐161982, 1 μM) receptors prevented the PGE₂‐induced decrease of Na⁺,K⁺‐ATPase activity. On the other hand, incubation with the selective EP2 agonist (butaprost, 0.1–10 μM) increased enzyme activity per se in a concentration‐dependent manner, but did not prevent the inhibitory effect of PGE₂. Incubation with a protein kinase A (PKA) inhibitor (H‐89, 1 μM) and a protein kinase C (PKC) inhibitor (GF‐109203X, 300 nM) also prevented PGE₂‐induced decrease of Na⁺,K⁺‐ATPase activity. Accordingly, PGE₂ increased phosphorylation of Ser943 at the α subunit, a critical residue for regulation of enzyme activity. Importantly, we also found that PGE₂ decreases Na⁺,K⁺‐ATPase activity in vivo. The results presented here imply Na⁺,K⁺‐ATPase as a target for PGE₂‐mediated signaling, which may underlie PGE₂‐induced increase of brain excitability.     

3H-mepyramine binding to histamine H1-receptors in guinea pig lung: Characteristics and regulation by ions

The antogonist [³H]-mepyramine is used to label histamine H₁-receptors in guinea pig lung. Scatchard analysis reveals two classes of binding sites. Monovalent cations decrease steady-state binding (Na⁺ > Li⁺ > K⁺), while divalent cations (Mg⁺⁺, Ca⁺⁺, Mn⁺⁺, Ba⁺⁺) exhibit a biphasic curve, increasing binding at low concentrations and decreasing it at higher levels. Na⁺ decreases both affinity and number of binding sites. Dissociation curve shows two components, and Na⁺ accelerates the rate of dissociation of the slower component. GTP does not affect the binding of the antagonist ³H-Mepyramine.     

THE AFFINITY OF ONION CELL WALLS FOR CALCIUM IONS

Cell walls prepared from onion bulbs were found to exhibit an affinity for Ca⁺⁺. The adsorption of this ion was enhanced by the action of pectin methylesterase. It was confirmed that Ca⁺⁺ reacts with two COO“ groups and the corresponding affinity constant, K, was found to be: log K = 4.25. The action of pectin methylesterase had no effect upon K. The cell walls, as prepared, had 25 % of the total COO⁻ groups occupied by Ca⁺⁺, 14 % by Mg⁺⁺, and 39 % by H⁺. Treatment with acidified ethanol removed all of the metallic cations. K⁺ and Mg⁺⁺ could displace Ca⁺⁺ from the cell walls. At concentrations from 10⁻³ to 3 times 10⁻³ m it required from 4.9 to 13.2 moles of Mg⁺⁺ to displace one mole of Ca⁺⁺. For K⁺ it required 80 moles to displace 1 mole of Ca⁺⁺ at K⁺ concentrations from 0.65 × 10⁻² to 1.6 × 10⁻² M.     

Selectivity of interaction of univalent cations with mammalian ribosomes studied by equilibrium dialysis in the presence of the K+ analogue, 204Tl+

The interaction of K⁺ with mammalian ribosomes was studied by equilibrium dialysis and compared with that of other univalent cations. The heavy K⁺ analogue, Tl⁺, binds more firmly than K⁺ to ribosomes and, unlike K⁺, has a practically useful isotope. With ²⁰⁴Tl⁺ as a marker of K⁺-selective binding the ribosome-cation interaction could be followed down to levels below 0.1 average Tl⁺-occupied site per ribosome. The Tl⁺/ribosome ratio varied with the free Tl⁺ concentration in a multiple way. At high Tl⁺ saturation Tl⁺ was easily displaced by Mg²⁺. With decreasing Tl⁺ saturation the competitive activity of Mg⁺⁺ was strikingly reduced, indicating that Tl⁺ and Mg⁺⁺ compete with different efficiency for different classes of sites.The experiments on univalent cations were performed at 1.5 mM Mg²⁺ under two complementary conditions: (1) Ribosomes were pretreated with 5 × 10^?2, 5 × 10^?3, and 5 × 10^?4 M LiNO₃, NaNO₃, KNO₃, and CsNO₃, and then equilibrated with different concentrations of ²⁰⁴TlNO₃ in the same buffers. (2) Ribosomes were pretreated with 10^?2, 10^?4, and 10^?6 M ²⁰⁴TlNO₃, and then equilibrated with different concentrations of LiNO₃, NaNO₃, KNO₃, and CsNO₃ (displacement experiments). At high Tl⁺ saturation Na⁺ and Li⁺ were about as active as K⁺ and Cs⁺ in competing with ²⁰⁴Tl⁺. With decreasing Tl⁺ saturation a differentiation occurred in favor of K⁺ and Cs⁺, with some preference for K⁺. It is concluded that ribosomes contain a limited number of sites with pronounced ion specificity. Of physiological cations K⁺ is most firmly bound to these sites.     

MIGRATORY CELL LOCOMOTION VERSUS NERVE AXON ELONGATION : Differences Based on the Effects of Lanthanum Ion

The effects of lanthanum ions (La⁺⁺⁺) on the locomotion and adhesion of g lial cells and elongating nerve axons are reported. La⁺⁺⁺ increases adhesion of both glia and of nerve growth cones to a plastic substratum. La⁺⁺⁺ also markedly reduces glia locomotion, but it does not inhibit nerve elongation. Electron-opaque deposits are seen on the cell surface and within cytoplasmic vesicles of glia and nerves cultured in a La⁺⁺⁺-containing medium. Possible modes of action for La⁺⁺⁺ are discussed, particularly the possibilities that Ca⁺⁺ fluxes or Ca⁺⁺ involvement in adhesion are altered by La⁺⁺⁺. The results are consistent with the hypothesis that cell migration and nerve axon elongation differ in mechanism, with respect to both adhesive interactions and the activity of microfilament systems.     

Effects of Cations on (Ca2++ Mg2+)-Activated ATPase from Rat Brain

Abstract: With a partially purified, membrane-bound (Ca + Mg)-activated ATPase preparation from rat brain, the K_0.5 for activation by Ca²⁺ was 0.8 p μm in the presence of 3 mm -ATP, 6 mm -MgCl₂, 100 m_M-KCI, and a calcium EGTA buffer system. Optimal ATPase activity under these circumstances was with 6-100 μm -Ca²⁺, but marked inhibition occurred at higher concentrations. Free Mg²⁺ increased ATPase activity, with an estimated K_0.5, in the presence of 100 μm -CaCl₂, of 2.5 mm ; raising the MgCl₂ concentration diminished the inhibition due to millimolar concentrations of CaCl₂, but antagonized activation by submicromolar concentrations of Ca²⁺. Dimethylsulfoxide (10%, v/v) had no effect on the K_0.5 for activation by Ca²⁺, but decreased activation by free Mg²⁺ and increased the inhibition by millimolar CaCl₂. The monovalent cations K⁺, Na⁺, and TI⁺ stimulated ATPase activity; for K⁺ the K_0.5 was 8 mm , which was increased to 15 mm in the presence of dimethylsulfoxide. KCI did not affect the apparent affinity for Ca²⁺ as either activator or inhibitor. The preparation can be phosphorylated at 0°C by [γ-³²P]-ATP; on subsequent addition of a large excess of unlabeled ATP the calcium dependent level of phosphorylation declined, with a first-order rate constant of 0.12 s^?1. Adding 10 mm -KCI with the unlabeled ATP increased the rate constant to 0.20 s^?1, whereas adding 10 mm -NaCl did not affect it measurably. On the other hand, adding dimethyl-sulfoxide slowed the rate of loss, the constant decreasing to 0.06 s^?1. Orthovanadate was a potent inhibitor of this enzyme, and inhibition with 1 μm -vanadate was increased by both KCI and dimethylsulfoxide. Properties of the enzyme are thus reminiscent of the plasma membrane (Na + K)-ATPase and the sarcoplasmic reticulum (Ca + Mg)-ATPase, most notably in the K⁺ stimulation of both dephosphorylation and inhibition by vanadate.     

The effect of multivalent cations on the membrane potential of the Ehrlich ascites tumor cell

The effect of di- and trivalent cations on the membrane potential of the Ehrlich ascites tumor cell has been investigated using micro-electrode techniques. In solutions free of multivalent cations the average membrane potential for 46 cells was 8.3 ± 0.5 mv (SE). However, the potentials were not stable and decayed with a half-time of about six seconds. Addition of Ca⁺⁺ decreased the rate of decay and concomitantly increased the membrane potential. The magnitude of these effects was a function of the Ca⁺⁺ concentration. At the optimum concentration (2 mM ), the half-time of decay was increased to 12 seconds and the membrane potential was raised to 17.8 ± 1.7 mv (SE). The related alkaline-earth cations, Sr⁺⁺, Ba⁺⁺ and Mg⁺⁺ had similar effects on both the stability and magnitude of the membrane potential. The effect of La⁺⁺⁺, which was qualitatively similar to that of the divalent cations, was also concentration dependent. However, 100-fold lower concentrations were adequate to achieve comparable effects. Moreover, membrane potentials were stable for up to ten minutes in La⁺⁺⁺-containing solutions. Variations in intracellular Cl^? content induced by temperature changes were paralleled by changes in membrane potentials. However, the potentials were not those expected for a simple Cl^? electrode.     

Further Studies on the Roles of Sodium and Potassium in the Generation of the Electro-Olfactogram : Effects of mono-, di-, and trivalent cations

In the negative EOG-generating process a cation which can substitute for Na⁺ was sought among the monovalent ions, Li⁺, Rb⁺, Cs⁺, NH₄⁺, and TEA⁺, the divalent ions, Mg⁺⁺, Ca⁺⁺, Sr⁺⁺, Ba⁺⁺, Zn⁺⁺, Cd⁺⁺, Mn⁺⁺, Co⁺⁺, and Ni⁺⁺, and the trivalent ions, Al⁺⁺⁺ and Fe⁺⁺⁺. In Ringer solutions in which Na⁺ was replaced by one of these cations the negative EOG's decreased in amplitude and could not maintain the original amplitudes. In K⁺-Ringer solution in which Na⁺ was replaced by K⁺, the negative EOG's reversed their polarity. Recovery of these reversed potentials was examined in modified Ringer solutions in which Na⁺ was replaced by one of the above cations. Complete recovery was found only in the normal Ringer solution. Thus, it was clarified that Na⁺ plays an irreplaceable role in the generation of the negative EOG's. The sieve hypothesis which was valid for the positive EOG-generating membrane or IPSP was not found applicable in any form to the negative EOG-generating membrane. The reversal of the negative EOG's found in K⁺- , Rb⁺- , and Ba⁺⁺-Ringer solutions was attributed to the exit of the internal K⁺. It is, however, not known whether or not Cl^- permeability increases in these Na⁺-free solutions and contributes to the generation of the reversed EOG's.     

Prostaglandins have limited actions on abnormalities of beating induced in cultured heart cells

Prostaglandins are antiarrhythmic in a variety of situations including ischaemic arrhythmias, but the mechanisms involved are not known. In view of this, the protective actions of prostaglandins A₂, E₂, F_1α, F_2β, and I₂ against abnormalities of beating induced in cultured heart cells were investigated. Abnormalities of beating were induced in single cells by a variety of agents including ouabain, Ca⁺⁺, K⁺, dinitrophenol (DNP), and toxic material from the jellyfish . Abnormalities were assessed in terms of rate, rate range, subjective arrhythmic behaviour and percent cells beating. The prostaglandins (at 10⁻⁷-10⁻⁵ M) were added with the arrhythmogenic agent to test for their ability to modify agent-induced beating abnormalities and were compared with lidocaine and quinidine. Prostaglandins alone had minimal direct effects on the cells and only minimally reduced responses to arrhythmogenic agents. The most protective prostaglandins, PGE₂ and PGF_1α, tended to normalise beating behaviour most noticeably in DNP-treated cells, unlike lidocaine and quinidine which were effective against Ca⁺⁺-induced changes while worsening those of K⁺. Thus, a general ability to protect disturbed cardiac cells is not seen with high concentrations of prostaglandins.     

Distribution of prostaglandin E2-sensitive adenylate cyclase along the rat nephron

To define sites of prostaglandin action of renal tubules, the distribution of adenylate cyclase sensitive to prostaglandin E₂ (PGE₂) was examined in single nephron segments dissected from rat kidney. Further, the interaction between PGE₂ and vasopressin on adenylate cyclase activity in nephron segments sensitive to vasopressin was evaluated. Procedures involved in isolating nephron segments were without effects on adenylate cyclase stimulation by PGE₂. PGE₂ stimulated adenylate cyclase activity of the thin descending limb of Henle (tDL), cortical collecting tubules (CCT), and medullary collecting tubules (MCT) at concentrations of 1.4 × 10^?5 to 2.8 × 10^?5 M. PGE₂ was without effects in other nephron segments tested including proximal convoluated tubules, proximal pars recta, the thin and thick ascending limb of Henle's loop, and distal and connecting tubules. PGE₂, at both high (2.8 × 10^?5 M) and low (2.8 × 10^?8 M) concentrations, did not inhibit adenylate cyclase activity stimulated by submaximal doses of vasopressin in medullary thick ascending limb of Henle (MTAL), CCT, and MCT. These data define the distribution of PGE₂-sensitive adenylate cyclase in the rat nephron, i.e., tDL, CCT, and MCT, and show the lack of direct inhibitory actions of PGE₂ on vasopressin sensitive adenylate cyclase in MTAL, CCT, and MCT.     

Ca++ fluxes in resealed synaptic plasma membrane vesicles

The effect of the monovalent cations Na⁺, Li⁺, and K⁺ on Ca⁺⁺ fluxes has been determined in resealed synaptic plasma membrane vesicle preparations from rat brain. Freshly isolated synaptic membranes, as well as synaptic membranes which were frozen (?80°C), rapidly thawed, and passively loaded with K₂/succinate and ⁴⁵CaCl₂, rapidly released approximately 60% of the intravesicular Ca⁺⁺ when exposed to NaCl or to the Ca⁺⁺ ionophore A 23187. Incubation of these vesicles with LiCl caused a lesser release of Ca⁺⁺. The EC₅₀ for Na⁺ activation of Ca⁺⁺ efflux from the vesicles was approximately 6.6mM. exposure of the Ca⁺⁺-loaded vesicles to 150 mM KCl produced a very rapid (?1 sec) loss of Ca⁺⁺ from the vesicles, but the Na⁺-induced efflux could still be detected above this K⁺ - sensitive effect. Vesicles pre-loaded with NaCl (150 mM) exhibited rapid ⁴⁵Ca uptake with an estimated EC₅₀ for Ca⁺⁺ of 7–10 μM. This Ca⁺⁺ uptake was blocked by dissipation of the Na⁺ gradient. These observations are suggestive of the preservation in these purified frozen synaptic membrane preparations of the basic properties of the Na⁺Ca⁺⁺ exchange process and of a K⁺ - sensitive Ca⁺⁺ flux across the membranes.     

Mitogen-activated Ca++ channels in human B lymphocytes

Two complementary experimental methods have been used to examine mitogen-induced transmembrane conductances in human B cells using the Daudi cell line as a model for human B cell activation. Spectrofluorometry was used to investigate mitogen-induced changes in [Ca⁺⁺]_i and transmembrane potential. Activation of human B cells with anti-μ antibodies resulted in a biphasic rise in [Ca⁺⁺]_i, the second phase being mediated by the influx of extracellular Ca⁺⁺. Ca⁺⁺ influx was inhibited by high [K⁺]e, suggesting that this influx was transmembrane potential sensitive. Membrane currents of Daudi cells were investigated using voltage clamp techniques. Before mitogenic stimulation, the cells were electrically quiet. Within several minutes of the addition of anti-μ antibodies to the bath solution, inward currents were observed at negative voltages. Whole-cell currents changed instantly with voltage steps and were transmembrane potential sensitive in that at potentials more positive than ?40 mV no currents were detectable. A similar conductance was also activated by the introduction of IP₃ into the intracellular solution, suggesting that IP₃ generation after surface IgM crosslinking is involved in the activation of this conductance. Both anti-μ and IP³ induced currents were blocked by 1 mM La⁺⁺⁺, which is known to block Ca⁺⁺ channels. These results strongly support the presence of membrane Ca⁺⁺ channels in human B cells that function in the early stages of activation. Changes in transmembrane potential appear to be important in regulating Ca⁺⁺ influx. These mechanisms work in concert to regulate the level of [Ca⁺⁺]_i during the early phases of human B cell activation. © 1993 Wiley-Liss, Inc.     

A highly ion-sensitive ATP-phosphorylation system in lobster nerve

The transfer of -phosphate from ³²P labeled adenosine-triphosphate (ATP) at low concentrations (10^?10 to 10^?7M) into the peripheral nerve of the lobster was found to be highly sensitive to external ionic environments. The phosphorylation process is inhibited at conditions similar to extracellular environments (high Na⁺, Ca⁺⁺ and pH) and stimulated by those close to intracellular medium (high K⁺, Mg⁺⁺ and low pH). This system is not related to NaK ATPase (pump ATPase) which is highly sensitive to ouabain and is active only at higher ATP concentrations (>10^?6M). The system is membrane bound and sensitive to a variety of neuro-active agents which are known to interfere with ionic conductance changes in axons.