Replacement Of KCl Research Articles

Effect of partial substitution of NaCl by KCl , CaCl 2 , and MgCl 2 on properties of mixed gelation from myofibrillar protein and Flammulina velutipes protein

The effects of substitution of sodium chloride (NaCl) by chloride salt mixtures (KCl, CaCl2, and MgCl2) with different formulas on quality characteristics of mixed protein system containing 80% myofibrillar protein (MP) and 20% Flammulina velutipes protein (FVP) were investigated in this study. The addition of chloride salts (KCl, CaCl2, and MgCl2) into a reduced-sodium (0.3 M NaCl) mixed gel system, increased the gel properties, and network structure of the MP-FVP gel, which might be attributed to the increase in protein solubility and rheological properties. The examination of the turbidity, particle size, zeta potential, UV spectroscopy, and Fourier transform infrared spectroscopy indicated that the protein molecules were obviously aggregated, that the electrostatic interaction was weakened, that the hydrophobic interaction and β-sheet content was increased. The results showed that when KCl, CaCl2, and MgCl2 were used to replace NaCl at 35, 10, and 5%, respectively, the gel properties reached the maximum. Practical applications This study evaluated the effects of different types of sodium salt substitution on the structural properties of proteins and the structural properties of mixed myofibrillar protein and Flammulina velutipes protein gels. The results revealed that the mixed protein molecules apparently aggregated under different replacement of KCl, CaCl2, and MgCl2 (i.e., 35, 10, and 5%). The hydrophobicity and β-sheet content significantly increased, while the electrostatic interaction decreased, leading to enhanced mixed gel properties. The results indicated the different replacements of sodium salts could improve the gels characteristics, which may be a guide for the future production of low-salt meat products.

Replacement of KCl with organic fertilizer on cultivation of porang (Amorphophallus muelleri Blume) under sonokeling stands

KCl fertilization is generally done to increase the productivity of porang tubers, but the continuous use of KCl can degrade soil fertility. Based on this, KCl needs to be replaced with organic fertilizer. This research aimed to determine the effect and replacement of KCl with organic fertilizer which caused the highest porang yield. The research was held in the BKPH Pojok forest area, Grobogan, Central Java in March-July 2020. The research was using one-factor RCBD. This treatment was the replacement of KCl with organic fertilizer with percentage 100:0%, 75:25%, 50:50%, 25:75%, 0:100%, and control in 4 replications. Observation parameters included tuber fresh weight, tuber storage weight, tuber diameter, tuber thickness, and tuber chip weight. The data were processed by using ANOVA 5% and DMRT 5%. The results showed that KCl had a strong influence on the yield of porang which included tuber fresh weight, tuber storage weight, and tuber chip weight, and organic fertilizer was able to replace the role of KCl. The usage of KCl and organic fertilizer was selected based on the yield of the highest tuber diameter and thickness and appeared with the same percentage (50% KCl:50% organic fertilizer).

Basis of Protein Stabilization by K Glutamate: Unfavorable Interactions with Carbon, Oxygen Groups

Potassium glutamate (KGlu) is the primary Escherichia coli cytoplasmic salt. After sudden osmotic upshift, cytoplasmic KGlu concentration increases, initially because of water efflux and subsequently by K+ transport and Glu− synthesis, allowing water uptake and resumption of growth at high osmolality. In vitro, KGlu ranks with Hofmeister salts KF and K2SO4 in driving protein folding and assembly. Replacement of KCl by KGlu stabilizes protein-nucleic acid complexes. To interpret and predict KGlu effects on protein processes, preferential interactions of KGlu with 15 model compounds displaying six protein functional groups—sp3 (aliphatic) C; sp2 (aromatic, amide, carboxylate) C; amide and anionic (carboxylate) O; and amide and cationic N—were determined by osmometry or solubility assays. Analysis of these data yields interaction potentials (α-values) quantifying non-Coulombic chemical interactions of KGlu with unit area of these six groups. Interactions of KGlu with the 15 model compounds predicted from these six α-values agree well with experimental data. KGlu interactions with all carbon groups and with anionic (carboxylate) and amide oxygen are unfavorable, while KGlu interactions with cationic and amide nitrogen are favorable. These α-values, together with surface area information, provide quantitative predictions of why KGlu is an effective E. coli cytoplasmic osmolyte (because of the dominant effect of unfavorable interactions of KGlu with anionic and amide oxygens and hydrocarbon groups on the water-accessible surface of cytoplasmic biopolymers) and why KGlu is a strong stabilizer of folded proteins (because of the dominant effect of unfavorable interactions of KGlu with hydrocarbon groups and amide oxygens exposed in unfolding).

Role of chloride ions in the promotion of auxin-induced growth of maize coleoptile segments.

The mechanism of auxin action on ion transport in growing cells has not been determined in detail. In particular, little is known about the role of chloride in the auxin-induced growth of coleoptile cells. Moreover, the data that do exist in the literature are controversial. This study describes experiments that were carried out with maize (Zea mays) coleoptile segments, this being a classical model system for studies of plant cell elongation growth. Growth kinetics or growth and pH changes were recorded in maize coleoptiles using two independent measuring systems. The growth rate of the segments was measured simultaneously with medium pH changes. Membrane potential changes in parenchymal cells of the segments were also determined for chosen variants. The question of whether anion transport is involved in auxin-induced growth of maize coleoptile segments was primarily studied using anion channel blockers [anthracene-9-carboxylic acid (A-9-C) and 4,4'-diisothiocyanatostilbene-2,2'-disulphonic acid (DIDS)]. In addition, experiments in which KCl was replaced by KNO3 were also performed. Both anion channel blockers, added at 0·1 mm, diminished indole-3-acetic acid (IAA)-induced elongation growth by ~30 %. Medium pH changes measured simultaneously with growth indicated that while DIDS stopped IAA-induced proton extrusion, A-9-C diminished it by only 50 %. Addition of A-9-C to medium containing 1 mm KCl did not affect the characteristic kinetics of IAA-induced membrane potential changes, while in the presence of 10 mm KCl the channel blocker stopped IAA-induced membrane hyperpolarization. Replacement of KCl with KNO3 significantly decreased IAA-induced growth and inhibited proton extrusion. In contrast to the KCl concentration, the concentration of KNO3 did not affect the growth-stimulatory effect of IAA. For comparison, the effects of the cation channel blocker tetraethylammonium chloride (TEA-Cl) on IAA-induced growth and proton extrusion were also determined. TEA-Cl, added 1 h before IAA, caused reduction of growth by 49·9 % and inhibition of proton extrusion. These results suggest that Cl(-) plays a role in the IAA-induced growth of maize coleoptile segments. A possible mechanism for Cl(-) uptake during IAA-induced growth is proposed in which uptake of K(+) and Cl(-) ions in concert with IAA-induced plasma membrane H(+)-ATPase activity changes the membrane potential to a value needed for turgor adjustment during the growth of maize coleoptile cells.

Biosorption of Chromium, Cadmium, and Cobalt from Aqueous Solution by Immobilized Living Cells of Chryseomonas luteola TEM 05

In this work, the potential use of the immobilized cells of Chryseomonas luteola TEM 05 for the removal of Cr+6, Cd+2 and Co+2 ions from aqueous solutions was investigated. The living cells of C. luteola TEM 05 were firstly entrapped both in carrageenan and chitosan coated carrageenan gels and then used in biosoption of the metal ions in batch reactors at pH 6.0, 25°C, in 100 mg L−1 of each metal solution. Besides this, a process of competitive biosorption of these metal ions was also described and compared to single metal ion adsorption in solution. According to the immobilization results, the replacement of KCl by KCl-chitosan as gelling agent improved the mechanical strength and thermal stability of the gel. In addition, the C. luteola TEM 05 immobilized carrageenan-chitosan gel system was quite more efficient for the fast adsorption of metal ions from aqueous solution than the carrageenan gels without biomass.

Lithium Desensitizes Brain Mitochondria to Calcium, Antagonizes Permeability Transition, and Diminishes Cytochrome c Release

Among the numerous effects of lithium on intracellular targets, its possible action on mitochondria remains poorly explored. In the experiments with suspension of isolated brain mitochondria, replacement of KCl by LiCl suppressed mitochondrial swelling, depolarization, and a release of cytochrome c induced by a single Ca2+ bolus. Li+ robustly protected individual brain mitochondria loaded with rhodamine 123 against Ca2+-induced depolarization. In the experiments with slow calcium infusion, replacement of KCl by LiCl in the incubation medium increased resilience of synaptic and nonsynaptic brain mitochondria as well as resilience of liver and heart mitochondria to the deleterious effect of Ca2+. In LiCl medium, mitochondria accumulated larger amounts of Ca2+ before they lost the ability to sequester Ca2+. However, lithium appeared to be ineffective if mitochondria were challenged by Sr2+ instead of Ca2+. Cyclosporin A, sanglifehrin A, and Mg2+, inhibitors of the mitochondrial permeability transition (mPT), increased mitochondrial Ca2+ capacity in KCl medium but failed to do so in LiCl medium. This suggests that the mPT might be a common target for Li+ and mPT inhibitors. In addition, lithium protected mitochondria against high Ca2+ in the presence of ATP, where cyclosporin A was reported to be ineffective. SB216763 and SB415286, inhibitors of glycogen synthase kinase-3beta, which is implicated in regulating reactive oxygen species-induced mPT in cardiac mitochondria, did not increase Ca2+ capacity of brain mitochondria. Altogether, these findings suggest that Li+ desensitizes mitochondria to elevated Ca2+ and diminishes cytochrome c release from brain mitochondria by antagonizing the Ca2+-induced mPT.

Effect of potassium magnesium chloride in the fertigation solution as partial source of potassium on growth, yield and quality of greenhouse tomato

The effect of partial replacement of KCl in the fertigation by KCl·MgCl 2 on growth, yield and quality of greenhouse tomato (cv. Durinta) was studied in a soil-less system. Forty-seven days after planting (DAP), three treatment solutions were applied to the plants using different K sources: (1) KNO 3, (2) KCl, and (3) KCl·MgCl 2+KCl (25%:75% in terms of K supplied). In both treatments 2 and 3, NH 4NO 3, Ca(NO 3) 2 and HNO 3 were added as source of N. Plant height and total chlorophyll were the highest in the KCl+KCl·MgCl 2 treatment. Leaf Mg content was significantly lower in the KCl treatment, whereas highest in the KCl+KCl·MgCl 2 treatment. Both KCl and KCl+KCl·MgCl 2 led to a significantly higher leaf Cl content as compared with the KNO 3 treatment, but no Cl toxicity was observed in either treatment. Total yield was not different among treatments. Fruit firmness and freshness of the calyx were significantly improved by KCl and KCl+KCl·MgCl 2, and the number of rotten and blotchy fruits were significantly reduced by both these treatments. KCl+KCl·MgCl 2 also led to significantly higher levels of glucose, Mg and dry matter content in the fruit. Lower NO 3 and higher Fe contents were measured in both treatments 2 and 3. Although KCl as sole K source showed lower foliar Mg level as compared to KNO 3, the use of the KCl in tomato fertigation improved tomato fruit appearance and qualities. However, 25% replacement of KCl by KCl+KCl·MgCl 2 increased the foliar Mg level and improved fruit qualities even further.

Microbial degradation of 4-chlorophenol by microorganisms entrapped in carrageenan-chitosan gels

A new cell immobilization method based on the replacement of KCl by KCl +chitosan as gelling agent was developed. The results showed that through addition of chitosan into gelling agent, the mechanical strength and thermal stability of the carrageenan gel were greatly improved. The new immobilization method was used to entrap a chlorophenol-degrading microorganism. The immobilized microbial cells were applied for chlorophenol biodegradation. The experiments demonstrated that immobilized cells exhibit a higher bioactivity in the degradation of chlorophenol than free cells.

Chronic Extracellular Acidosis Induces Plasmalemmal Vacuolar Type H+ ATPase Activity in Osteoclasts

Proton extrusion into an extracellular resorption compartment is an essential component of bone degradation by osteoclasts. Chronic metabolic acidosis is known to induce negative calcium balance and bone loss by stimulating osteoclastic bone resorption, but the underlying mechanism is not known. The present studies were undertaken to evaluate whether chronic acidosis affects proton extrusion mechanisms in osteoclasts cultured on glass coverslips. Acidosis, mimicked experimentally by maintaining the cells at extracellular pH 6.5, rapidly lowered intracellular pH to 6.8. However, after 2 hours, a proportion of cells demonstrated the capacity to restore intracellular pH to near normal levels. To define the mechanism responsible for this recovery, the activity of individual H+ transport pathways was analyzed. We found that chronic acid treatment for up to 6 h did not significantly affect the cellular buffering power or Na+/H+ antiport activity. In contrast, chronic acidosis activated vacuolar H+ pumps in the osteoclasts. Although only approximately 5% of the control cells displayed proton pump activity, about 40% of cells kept at extracellular pH 6. 5 for 4-6 h were able to recover from the acute acid load by means of bafilomycin A1-sensitive proton extrusion. Conversely, the H+-selective conductance recently described in the plasma membrane of osteoclasts was clearly inhibited in the cells exposed to chronic acidosis. Following acid treatment, the activation threshold of the H+ conductance was shifted to more positive potentials, and the current density was significantly reduced. Considered together, these results suggest that induction of plasmalemmal vacuolar type ATPase activity by chronic acidosis, generated either systemically due to metabolic disease or locally at sites of inflammation, is likely to stimulate osteoclastic bone resorption and thus to promote bone loss.

Mechanism of the stimulatory effect of a potassium‐rich medium on the phosphorylation of glucose in isolated rat hepatocytes

We have investigated the mechanism by which the replacement of a Na(+)-rich medium by a K(+)-rich medium causes an increase in the apparent affinity of glucokinase (hexokinase IV or D) for glucose in isolated hepatocytes [Bontemps, F., Hue, L. & Hers, H. G. (1978) Biochem. J. 174, 603-611]. The stimulatory effect of a K(+)-rich medium on the rate of glucose phosphorylation, as assessed by the release of tritium from [2-3H]glucose, was only partially additive with the effect of fructose, suggesting that it was also due to a decrease in the inhibition exerted on glucokinase by its regulatory protein. Measurements of metabolites indicated that the effect of the K(+)-rich medium was neither due to the formation of fructose 1-phosphate, nor to changes in the concentrations of fructose 6-phosphate or Pi, two other effectors of the regulatory protein. Replacement of Na+ by K+ in the medium resulted in a time-dependent and dose-dependent increase in cell volume that paralleled the changes in the rate of detritiation observed at 5 mM glucose. The water and chloride contents, estimated using radiolabelled compounds, were threefold and tenfold higher, respectively, in K+ cells than in Na+ cells, and the intracellular Cl- concentration about threefold higher (94 versus 29 meq/l). The effects of the K(+)-rich medium on cell volume, Cl- concentration and rate of detritiation were greatly reduced by including 80 mM trehalose or sucrose in the medium at the start of the incubation. Addition of trehalose to cells incubated for 45-50 min in the K(+)-rich medium caused an immediate decrease in cell volume whereas the rate of detritiation and the Cl- concentration underwent a transient increase followed by a decrease. Replacement of KCl by KBr, potassium acetate or potassium trichloroacetate in the K(+)-rich medium resulted in different relationships between cell volume and the rate of detritiation, in agreement with the differential effect of these salts on the activity of purified glucokinase assayed in the presence of regulatory protein. From these results we conclude that the increase in the activity of glucokinase induced by a KCl-rich medium is at least partly due to an increase in the concentration of Cl-, which relieves the inhibition exerted by the regulatory protein on purified glucokinase.

Luteinizing hormone inhibits potassium outward currents in swine granulosa cells by intracellular calcium mobilization.

Potassium currents of swine granulosa cells were studied using the patch clamp technique in the whole cell configuration. Granulosa cells stepped to positive potentials (+60 mV) from -40 mV holding potential exhibit a slowly activating, noninactivating outward potassium current. Tail current reverse potential (between -90 and -100 mV) and the current inhibition brought about by the replacement of KCl with CsCl in the pipette solution indicate that this current is carried by K ions. LH was found to significantly reduce the amplitude of this current. The effect was dose and time dependent. Similar inhibition (20-30% of the initial current) was reached with doses of 1-50 micrograms/ml, but in times proportionally shorter as the dose increased (50% inhibition was reached in 170-180 and 30-40 sec with 1 and 50 micrograms LH/ml, respectively). Much longer and variable times (3-10 min) were required with lower doses (0.2 microgram/ml). The effect of LH was independent of extracellular Ca, while preexposure of cells to TMB-8, an inhibitor of intracellular Ca mobilization, completely prevented the effect of LH. Outward currents after LH treatment could be completely restored by perfusing the cells with ionomycin in Ca-free medium to facilitate calcium efflux from the cells. The present studies indicate that LH modifies the bioelectrical properties of swine granulosa cells. This effect is mediated by an elevation of intracellular calcium, probably mobilized from intracellular stores. The induced changes in K conductance may play a specific role in the transduction mechanisms for LH.

Inositol trisphosphate-induced calcium release in brain microsomes

The effects of different ionic media on Ca 2+ uptake and release in isolated brain microsomes were investigated. KCl (100 mM) provided the best medium for Ca 2+ uptake in the presence of ATP. The effect of myo-inositol 1,4,5-trisphosphate (IP 3) on Ca 2+ release was examined and was maximum at 0.2 μM. IP 3-induced Ca 2+ release was dependenton extramicrosomal free Ca 2+ concentration with maximal release at 5.0 μM free Ca 2+. Replacement of KCl by sucrose or NaCl did not show any response to IP 3. Electron microscopy showed that the microsomal fraction consisted of characteristic endoplasmic reticulum-derived vesicular profiles and were free of mitochondria or plasma membrane contamination. Our results support the concept that the endoplasmic reticulum is the target for IP 3-induced mobilization of Ca 2+ in the cell.

Replacement of potassium chloride by potassium glutamate dramatically enhances protein-DNA interactions in vitro.

Although protein-nucleic acid interactions exhibit dramatic dependences on both ion concentration and type in vitro, large variations in intracellular ion concentrations can occur in Escherichia coli and other organisms without apparent effects on gene expression in vivo. E. coli accumulates K+ and glutamate as cytoplasmic osmolytes. The cytoplasmic K+ concentration in E. coli varies from less than 0.2 to greater than 0.9 m as a function of external osmolarity; corresponding cytoplasmic glutamate concentrations range from less than 0.03 to greater than 0.25 m. Only low levels of chloride occur in the cytoplasm of E. coli at all osmotic conditions. Since most in vitro studies have been performed in chloride salts, whereas glutamate is the more relevant physiological anion, we have measured the effects of the substitution of potassium glutamate (KGlu) for KCl on the kinetics and equilibria of a variety of site-specific protein-DNA interactions in vitro. Both the interaction of E. coli RNA polymerase with two phage lambda promoters and the interactions of various restriction enzymes with their DNA cleavage sites are enhanced by this substitution. Using the abortive initiation assay, we find a greater than 30-fold increase in the second-order rate constant for open complex formation at the lambda PR promoter and a 10-fold increase at the lambda PR' promoter, when KGlu is substituted for KCl. Replacement of KCl by KGlu does not affect the strong salt dependences of these interactions; increasing either KCl or KGlu concentrations decreases both reaction rates and extents. Substitution of glutamate for chloride does, however, shift the range of salt concentrations over which these interactions are observable to higher K+ concentrations.(ABSTRACT TRUNCATED AT 250 WORDS)

The inhibitory effects of monovalent ions on force development in detergent-skinned ventricular muscle from guinea-pig.

Trabeculae from guinea-pig ventricles were bathed in a physiological saline, were stimulated electrically and were stretched until isometric force production was maximal. They were then 'skinned' by superfusion for 30 min with a solution containing the non-ionic detergent, Triton X-100. The steady isometric force generated by these skinned trabeculae was measured in solutions of different pCa (-log10 [Ca2+]) and salt concentration to investigate the influence of salt concentration on (i) the maximum Ca2+-regulated force (at pCa approximately 3.8), (ii) the resting force (at pCa greater than 8) and (iii) the pCa required for 50% Ca2+ activation (pCa50). The apparent affinity constants of EGTA (ethyleneglycol-bis-(beta-aminoethylether)-N,N,N',N'-tetraacetic acid) for Ca2+ were measured in the solutions of different salt composition by a pH-metric method and these constants were used in the calculation of solution pCa. Increases in ionic strength (gamma/2) (and osmolarity) over the range 0.1-0.4 mol/l produced by the addition of various monovalent Cl or K salts resulted in an approximately linear decrease in the maximum Ca2+-regulated force. The inhibitory actions of the different salts were similar despite the differences in size and structure of their constituent ions. Increases in solution osmolarity alone, produced by the addition of sucrose up to 0.6 mol/l, did not significantly alter the maximum Ca2+-regulated force. It was concluded that the influence of monovalent salts on the maximum Ca2+-regulated force was due entirely to the changes in ionic strength and was not related to changes in solution osmolarity or to specific ion effects. Resting force was little affected by changes in salt concentration at ionic strengths of 0.2 mol/l and above. At lower ionic strengths the muscles exhibited an elevated resting force and an inability to relax completely after a Ca2+ -regulated contracture. A decrease in ionic strength from 0.2 to 0.1 mol/l produced by a reduction in KCl concentration raised the pCa50 from 5.56 to 5.85, i.e. the Ca2+ sensitivity was increased. However, the same increase in pCa50 was observed when the [K+] was lowered at constant ionic strength, osmolarity and [Cl-] by the replacement of KCl with tetramethylammonium Cl or choline Cl. Therefore, the change in pCa50 induced by an alteration in KCl concentration was due entirely to a specific inhibitory action of K+ on the Ca2+ sensitivity of cardiac myofibrils and was unrelated to changes in ionic strength, osmolarity and [Cl-]. Substitution of KCl 0.1 mol/l in the solutions by NaCl did not significantly alter the pCa50.(ABSTRACT TRUNCATED AT 400 WORDS)

Failure of Dietary-Casein-Induced Acidosis to Explain the Hypercholesterolemia of Casein-Fed Rabbits

The partial replacement of casein by a mixture of gelatin, fish protein and soy protein in cholesterol-free semipurified diets of rabbits reduced the hypercholesterolemic response. The partial replacement of casein by the protein mixture also increased the feed intake and alleviated or reversed the weight loss observed from the casein diet. The data indicate that casein alone is not an ideal protein source for rabbits probably because of the imbalance of the amino acid composition. When KCl in the semipurified diets was replaced by KHCO3, a higher feed intake and a better growth were obtained, irrespective of the protein source in the diet. In addition, the feeding of semipurified diets containing KCl resulted in acidosis, which could be prevented by its replacement with KHCO3. A semipurified diet containing casein and KCl produced a more severe acidosis and higher serum cholesterol levels than the diet containing the protein mixture. Nevertheless, the correction of the acidosis by the replacement of KCl in the diet by KHCO3 did not lead to an abrogation of the casein-induced hypercholesterolemia.

Effects of potassium and sodium ions on the killing action of a Pichia kluyveri toxin in cells of Saccharomyces cerevisiae.

Loss of viability of toxin-treated cells of Saccharomyces cerevisiae SCF 1717 could be prevented in the period before they altered physiologically if cells were incubated in media with a suitable concentration of potassium (0.08 to 0.13 M) and hydrogen ions (pH 6.2 to 6.7). Incorporation of higher amounts of potassium chloride in the media had a pronounced negative effect on cell survival, particularly when the pH of the medium was lowered. Replacement of KCl by NaCl in the plate media was even more deleterious to toxin-treated cells and, in contrast with potassium, low concentrations of sodium ions could not sustain recovery of cells. Complete recovery of a toxin-treated cell suspension required an incubation of 3 h in a suitable medium. The recovery process was blocked by cycloheximide.

Influences of ionic environments on ACh-induced secretory responses in isolated perfused pancreas of rats.

The influence of extracellular Ca2+ concentration, [Ca2+]o, on the secretory response to acetylcholine (ACh) was analyzed in isolated perfused rat pancreas. The decrease of [Ca2+]o strongly diminished the amylase output and pancreatic juice flow in response to continuous stimulation with 5 X 10(-8) M ACh. A quantitative relation was found between the amount of amylase release by 5 X 10(-8) M ACh and the [Ca2+]o over a range of 0.1--2.5 mM. The partial replacement of NaCl with LiCl produced a diminution in both amylase output and pancreatic juice flow. A quantitative relation existed between the amount of ACh-induced amylase release and the [Na+]o over a range of 86--157 mM. The partial replacement of KCl with NaCl produced falls in both amylase output and pancreatic juice flow. Again, a quantitative relation existed between ACh-induced amylase release and [K+]o over a range of 1.0--5.6 mM. These results are compatible with the view that both the amylase output and the juice flow induced by 5 X 10(-8) M ACh are proportional to the amount of carrier-Ca complex and that the inward movement of the complex may be linked closely to the activation of Na pumps on the pancreatic acinar cell. A dose-response relation was found between the concentration of ACh and the amylase output. The relation was shifted to the left when 1 mU/ml cholecystokinin-pancreozymin (CCK-PZ) was added. A similar shift was observed when 1 mU/ml secretin was added. These results support the view that ACh, CCK-PZ, and secretin may activate the common cellular process in stimulus-secretion coupling, although these secretagogues may severally act on the different receptor sites.

Influence of external potassium concentration on secretory responses to cholecystokinin‐pancreozymin and ionophore A23187 in the pancreatic acinar cell.

1. The inhibitory effect of a diminution in [K+]o on the secretory processes of the pancreatic acinar cell was analysed in the isolated and perfused rat pancreas. 2. Partial replacement of KCl with NaCl produced falls in both amylase output and pancreatic juice flow which were induced by 5 m-u. cholecystokinin-pancreozymin (CCK-PZ)/ml., and the responses were slowly regained after reintroduction of a standard concentration of KCl. 3. A quantitative relation was found between the amount of amylase released by CCK-PZ and [K+]o over the range 1.0--5.6 mM. A linear relation was obtained between the ratio of E1K+]3(0)/amylase output and [K+]3(0). 4. A quantitative relation was also found between the pancreatic juice flow induced by CCK-PZ and [K+]o over the range 1.0--5.6 mM. A linear relation was obtained between the ratio of [K+]3(0)/juice flow and [K+]3(0). 5. The results obtained may be explained by a model as follows: (1) amylase output and juice flow induced by CCK-PZ are simply proportional to the amount of complex composed of a carrier molecule bearing one Ca and four Na atoms, (2) the inward movement of the complex is tightly linked to the CCK-PZ-induced activation of pumps which are postulated on both basal and apical sides of the membrane of the acinar cell, and (3) the Na:K coupling ratio of the pumps is close to 3:2. 6. A quantitative relation was also found between the amount of amylase released by Ca reintroduction after pre-treatment with Ca-ionophore A23187 and [K+]o. This may be explained by a model essentially similar to that mentioned above. 7. Total replacement of NaCl with NaBr slightly reduced the CCK-PZ-induced fluid and amylase secretion. Total replacement of NaCl with sodium isethionate or with sodium pyruvate, however, virtually abolished the CCK-PZ-induced fluid and amylase secretion.

Biochemical aspects of the visual process. XXXVI. Calcium accumulation in cattle rod outer segments: Evidence for a calcium-sodium exchange carrier in the rod sac membrane

Accumulation of calcium has been studied in bovine rod outer segments (rods), isolated by sucrose density gradient centrifugation. Calcium-depleted rods are obtained by having ethyleneglycol-bis-( β-aminoethylether)- N, N′-tetraacetic acid (EGTA) present during isolation. Rods thus isolated have a leaky plasma membrane, as shown by the effects of ionophore A23187 and by their light-induced phosphorylation behaviour. The accumulation of 45Ca, determined by incubation followed by a single fast washing-filtration procedure, thus represents translocation across the rod sac membrane. Accumulation in non-depleted rods is independent of the external calcium level and of ATP, suggesting exchange of 40Ca by 45Ca. In depleted rods in the presence of ATP there is net uptake, sigmoidally increasing with the external calcium concentration to the level attained in non-depleted rods. This net uptake is abolished by omission of ATP, its replacement by β,γ-methylene ATP and lowering the temperature to 0° C, suggesting involvement of enzymatic hydrolysis of ATP. Replacement of KCl by NaCl in the medium causes marked inhibition of 45Ca uptake, both net uptake and exchange. Oligomycin, ruthenium red, lanthanum and ouabain do not inhibit accumulation. Efflux of 45Ca from pre-loaded rods is slow in a KCl medium ( t 1 2 ∼30 min at 25° C), but is greatly accelerated by addition of NaCl or Ca 2+ ( t 1 2 10 s at 25°C). It is concluded that the rod sac membrane contains a carrier system, which is sensitive towards Ca 2+ and Na + and which requires ATP for net uptake of Ca 2+ but not for exchange transport of Ca 2+ with Ca 2+ or Na +.

The membrane structure studied with cationic dyes. 1. The binding of cationic dyes to submitochondrial particles and the question of the polarity of the ion-translocation mechanism.

Cationic dyes interact weakly with de‐energized submitochondrial particles incubated in a KCl medium. Energization of the particles results in a marked increase of binding. The Scatchard plots reveal, in the case of neutral red, and at pH 7.0, binding sites of 150–200 μmol per g protein with a dissociation constant of about 2.5 μM. In the case of acridine orange at the same pH, the value for binding sites is about 50 μmol per g protein with a dissociation constant of about 2 μM.The binding of the cationic dyes to the energized particles is accompanied by a large decrease of absorbance which is reversed on addition of uncouplers. The change of absorbance is stoichiometrically dependent on the utilization of ATP, but independent of whether energy is supplied from respiration or from ATP. Within a certain range of protein/dye concentration ratio the reaction kinetics appears of pseudo first‐order character.The change of absorbance is proportional to the amount of particles until a saturation is obtained. This permits titration the binding sites of the membrane for the dye, using the “end‐point” of the absorbance change or using the method of continuous variations. The number of binding sites found through these procedures is in agreement with those obtained from the binding experiments.The pH of the medium has a marked effect on the dye‐membrane interaction. Increasing the pH from 6 to 8 results in an increase of the rate of binding, in a decrease of the dissociation constant and in a decrease of the number of binding sites.The lipophilic cation, nupercaine, exerts a marked inhibition on the binding of the cationic dye and the concomitant decrease of absorbancy. The inhibition is of competitive type with a Ki of 20 μM.The presence of electrolytes in the medium is required for the high affinity binding and the change of absorbance. Replacement of KCl with sucrose results in abolition of both phenomena.The results are discussed in relation to the problem of the proposed inverted polarity of the ion translocation mechanism in the submitochondrial particles. The suggestion is made that sonication of mitochondria results in a “loss of polarity” rather than in “inversion of polarity”. The kinetics of the interaction indicates a limiting step which is probably that required for the interaction of the dye with the energized site.