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Biomedical subjects

N Kraus-Friedmann

Publications and source records attributed to N Kraus-Friedmann.

At least 37 records · Page 2Linked to original sources
PubMed1989

Demonstration of adenylate-kinase activity in hepatic microsomes. Relevance to Ca2+ uptake.

It is demonstrated that the hepatic microsomal fraction contains significant adenylate-kinase activity. This explains a paradoxical ADP-stimulated Ca2+ uptake into microsomal vesicles which is inhibited when adenylate kinase is inhibited. The presence of adenylate kinase in the microsomal fraction helps to prevent sudden drops in ATP level, and thus has a stabilizing effect on the many ATP-dependent reactions carried out in this subcellular compartment.

Adenosine Diphosphate↗
PubMed1988

Inhibitor of anion transport, DIDS, releases Ca2+ from hepatic microsomes.

Addition of 4,4'-diisothiocyanostilbene-2, 2'-disulfonic acid (DIDS) to Ca2+ loaded hepatic microsomal vesicles evoked a dose-dependent release of the accumulated Ca2+. Ca2+ uptake was also inhibited. The effects of DIDS do not seem to be due to the inhibitions of either Cl- or proton fluxes. The results indicate that DIDS inhibits Ca2+ uptake and releases Ca2+ by inhibiting the Ca2+-ATPase and the formation of the phosphorylated intermediate of the enzyme, and that it might interact with a specific site on the vesicle which is involved in the translocation of Ca2+ across the microsomal and mitochondrial membranes.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗
PubMed1988

Demonstration of the presence of G-proteins in hepatic microsomal fraction.

The presence of G-proteins in isolated hepatic microsomal vesicles is demonstrated. The G-proteins were identified by their capacity to be ADP-ribosylated by cholera and pertussis toxins. Cholera toxin identified 42 and 45 kDa proteins, corresponding to alpha s-1 and alpha s-2, respectively. Pertussis toxin identified a 40 kDa protein corresponding to alpha i. The microsomal G-proteins are identical to the corresponding G proteins of the plasma membrane, but are present in different proportions; the microsomes have considerably less alpha s proteins than the plasma membrane.

Adenosine Diphosphate Ribose↗
PubMed1987

Inhibition of rat liver microsomal Ca2+-ATPase by fluorescein-5'-isothiocyanate.

Rat liver microsomal fraction was incubated at pH 8.8 with fluorescein-5'-isothiocyanate in a Tris-buffered sucrose medium. This treatment completely inhibited ATP-dependent Ca2+ transport, Ca2+-ATPase activity, and Ca2+-ATPase phosphoenzyme intermediate formation. Inhibition of Ca2+ transport and phosphoenzyme intermediate formation by fluorescein-5'-isothiocyanate was partially prevented by including ATP in the treatment medium. These data taken together are consistent with the proposal that fluorescein-5'-isothiocyanate binds the Ca2+-ATPase ATP-binding site, suggesting the presence of a lysine residue in this domain. Fluorescein-5'-isothiocyanate labeling of microsomal proteins had no measurable effect on the basal, Mg2+-ATPase activity. Using fluorescein-5'-isothiocyanate-labeled microsomal fraction, we demonstrated that the Mg2+-ATPase activity was inhibited by Ca2+.

Adenosine Triphosphate↗
PubMed1987

A comparison between 45Ca2+ and atomic absorption in calcium flux determinations in perfused rat liver.

Calcium efflux from perfused rat liver following the administration of Ca2+ releasing agents was measured with two methods, 45Ca2+ labeling and atomic absorption. The values obtained with atomic absorption were usually higher than the values obtained with 45Ca2+. These indicate that the intracellular Ca2+ did not equilibrate with the perfusate ca2+ during the 90-minute labeling period. A similar conclusion was reached by measuring the liver 45Ca2+ and 40Ca2+ content. In addition, the types of albumin added to the perfusate influenced the amounts of Ca2+ released.

Animals↗
PubMed1986

Glucagon-stimulated respiration and intracellular Ca2+.

The effects of extra- and intracellular Ca2+ on glucagon-stimulated respiration were examined in perfused rat liver. Glucagon increased the uptake of O2 to a significantly greater extent in Ca2+-containing perfusate than in Ca2+-free perfusate. If, however, the livers were perfused first with Ca2+-containing perfusate for 60 min in order to load the hormone-sensitive Ca2+ pool(s) and subsequently with Ca2+-free perfusate, glucagon was able to stimulate O2 uptake to the same extent in Ca2+-free, as in Ca2+-containing perfusate. These experiments support previous observations of a connection between Ca2+ and the hormonal stimulation of respiration, but indicate a role for intracellular, rather than extracellular, Ca2+ in the process.

Animals↗
PubMed1986

The effect of Mg2+ on hepatic microsomal Ca2+ and Sr2+ transport.

The ATP-dependent uptake of Ca2+ by rat liver microsomal fraction is dependent upon Mg2+. Studies of the Mg2+ requirement of the underlying microsomal Ca2+-ATPase have been hampered by the presence of a large basal Mg2+-ATPase activity. We have examined the effect of various Mg2+ concentrations on Mg2+-ATPase activity, Ca2+ uptake, Ca2+-ATPase activity and microsomal phosphoprotein formation. Both Mg2+-ATPase activity and Ca2+ uptake were markedly stimulated by increasing Mg2+ concentration. However, the Ca2+-ATPase activity, measured concomitantly with Ca2+ uptake, was apparently unaffected by changes in the Mg2+ concentration. In order to examine the apparent paradox of Mg2+ stimulation of Ca2+ uptake but not of Ca2+-ATPase activity, we examined the formation of the Ca2+-ATPase phosphoenzyme intermediate and formation of a Mg2+-dependent phosphoprotein, which we have proposed to be an attribute of the Mg2+-ATPase activity. We found that Ca2+ apparently inhibited formation of the Mg2+-dependent phosphoprotein both in the absence and presence of exogenous Mg2+. This suggests that Ca2+ may inhibit (at least partially) the Mg2+-ATPase activity. However, inclusion of the Ca2+ inhibition of Mg2+-ATPase activity in the calculation of Ca2+-ATPase activity reveals that this effect is insufficient to totally account for the stimulation of Ca2+ uptake by Mg2+. This suggests that Mg2+, in addition to stimulation of Ca2+-ATPase activity, may have a direct stimulatory effect on Ca2+ uptake in an as yet undefined fashion. In an effort to further examine the effect of Mg2+ on the microsomal Ca2+ transport system of rat liver, the interaction of this system with Sr2+ was examined. Sr2+ was sequestered into an A23187-releasable space in an ATP-dependent manner by rat liver microsomal fraction. The uptake of Sr2+ was similar to that of Ca2+ in terms of both rate and extent. A Sr2+-dependent ATPase activity was associated with the Sr2+ uptake. Sr2+ promoted formation of a phosphoprotein which was hydroxylamine-labile and base-labile. This phosphoprotein was indistinguishable from the Ca2+-dependent ATPase phosphoenzyme intermediate. Sr2+ uptake was markedly stimulated by exogenous Mg2+, but the Sr2+-dependent ATPase activity was unaffected by increasing Mg2+ concentrations. Sr2+ uptake and Sr2+-dependent ATPase activity were concomitantly inhibited by sodium vanadate. In contrast to Ca2+, Sr2+ had no effect on Mg2+-dependent phosphoprotein formation. Taken together, these data indicate that Mg2+ stimulated Ca2+ and Sr2+ transport by increasing the Ca2+ (Sr2+)/ATP ratio.(ABSTRACT TRUNCATED AT 400 WORDS)

Adenosine Triphosphatases↗
PubMed1986

Effects of glucagon and vasopressin on hepatic Ca2+ release.

The effects of physiological levels of glucagon on Ca2+ efflux were examined in the perfused rat liver. Two methods were used to estimate Ca2+ efflux: prior labeling of the calcium pools with 45Ca2+ and measurement of perfusate Ca2+ with atomic absorption. According to both methods, glucagon administration at the physiological level evoked Ca2+ release. The released Ca2+ originated mostly from a carbonylcyanide p-trifluoromethoxyphenylhydrazone (FCCP)-depletable pool and also from an FCCP-insensitive pool from which Ca2+ could be released with A23187. Maximally effective doses of glucagon and vasopressin had no additive effect on Ca2+ release. Prior administration of vasopressin resulted in markedly reduced Ca2+ release by glucagon. These results indicate that glucagon releases Ca2+ from the same pool that vasopressin does.

Animals↗
PubMed1985

Phosphorylated intermediates of two hepatic microsomal ATPases.

The hepatic microsomal Ca2+- and Mg2+-dependent ATPase phosphoenzyme intermediates were distinguished by using the chelators EGTA and CDTA (trans-cyclohexane-1,2-diamine-NNN'N'-tetra-acetic acid). The Ca2+-ATPase intermediate is a hydroxylamine-labile base-labile 125 000-Mr phosphoprotein. The Mg2+-ATPase intermediate is a hydroxylamine-stable base-stable 30 000-Mr phosphoprotein. This enzyme intermediate probably reflects the large basal ATPase activity of hepatic microsomal fraction. It is dependent on Mg2+, since formation of the phosphoenzyme is abolished in the presence of CDTA. Under these conditions, the basal ATPase activity is dramatically decreased. These data demonstrate two separate and distinct enzymes which are responsible for the two ATPase activities of hepatic microsomal fraction. Furthermore, these data indicate that more meaningful data about the microsomal Ca2+-ATPase might be obtained if the free ion concentrations are controlled with CDTA.

Adenosine Triphosphatases↗
PubMed1984

Liposome encapsulated tetracaine lowers blood glucose.

Tetracaine, a local anesthetic, was previously shown to block hormonal stimulation of gluconeogenesis and glycogenolysis ( Friedmann , N. and Rasmussen, H. (1970) Biochim. Biophys. Acta 222, 41-52). In the present studies tetracaine incorporated into liposomes (phospholipid vesicles) was injected into intact rats and epinephrine was administered an hour later. Liposomal tetracaine blocked 50% of the hyperglycemic response. When tetracaine, incorporated into liposomes, was injected into diabetic rats it reduced transiently, but significantly, blood glucose levels. Equivalent doses of free tetracaine were toxic. These studies indicate that liposomal drug administration might be developed into a tool to influence hepatic metabolism and, consequently, blood glucose levels.

Animals↗
PubMed1984

Unique calcium-dependent hydrophobic binding proteins: possible independent mediators of intracellular calcium distinct from calmodulin.

Calcium-dependent regulation of cellular processes is mediated by specific intracellular proteins. A newly described set of proteins isolated from chicken gizzard with Mr of 67 X 10(3), 35 X 10(3), 33 X 10(3) and 30 X 10(3) also express a hydrophobic site in the presence of calcium. These proteins are isolated from several other cellular tissues and are termed calcimedins. These proteins differ from calmodulin in isoelectric point, DEAE-cellulose binding characteristics and heat stability. The calcimedins do not activate calmodulin-dependent cyclic nucleotide phosphodiesterase but do activate a hepatic microsomal Ca2+ -ATPase system. Hence, the possibility is opened that calcium regulation of cellular processes is mediated by calcium-binding proteins in addition to calmodulin.

Animals↗
PubMed1983

Hepatic microsomal Ca2+-dependent ATPase. Calmodulin-dependence and partial purification.

The hepatic microsomal fraction contains tightly bound calmodulin as demonstrated by affinity chromatography. When this calmodulin was partially removed by EGTA treatment (0.5 mM-EGTA), the uptake of 45Ca2+ by the microsomal vesicles was stimulated by added calmodulin and inhibited by trifluoperazine (TFP). The Ca2+-dependent ATPase was partially purified on a calmodulin column. This partial purification resulted in a 500-fold increase in the specific activity of the enzyme when measured in the presence of added calmodulin. Antibodies prepared against calmodulin prevented this stimulatory effect. The fraction eluted from the calmodulin column contained several protein bands indicating that the specific activity of the Ca2+-dependent ATPase is probably still underestimated. There are likely to be other calmodulin-sensitive processes present in the hepatic microsomal fraction.

Animals↗
PubMed1983

45Ca2+ uptake and phospholipid methylation in isolated rat liver microsomes.

The effects of glucagon, epinephrine and insulin on hepatic phospholipid methylation were studied. Glucagon, either injected into rats or added to perfused livers, stimulated methylation in subsequently isolated microsomes. Epinephrine also increased phospholipid methylation. Insulin by itself did not influence the rate of the reaction, but, when administered prior to glucagon, it blocked the effect of the latter. The possibility that the observed stimulation of phospholipid methylation might be causally linked to the reported stimulation by glucagon of 45Ca2+ uptake in subsequently isolated liver microsomes was examined. Both the substrate and the competitive inhibitor of the methylation reaction, S-adenosylmethionine and S-adenosylhomocysteine, had profound effect on the rate of phospholipid methylation, without having comparable effects on Ca2+ uptake. S-adenosylmethionine in increasing concentration stimulated methylation four-fold, while no significant changes in 45Ca2+ uptake were seen. S-adenosylhomocysteine did not inhibit 45Ca2+ uptake even at levels causing more than 95% decrease in methylation. In conclusion, while both phospholipid methylation and 45Ca2+ uptake seem to be hormonally controlled, the correlation between these two processes was not sufficient to support the notion that the changes in 45Ca2+ uptake are caused by the changes in phospholipid methylation.

Animals↗
PubMed1982

Glucagon stimulation of hepatic Na+, K+-ATPase.

In the perfused rat liver administration of glucagon was shown to result in a transiently increased uptake of K+, indicating the possible involvement of the Na+, K+-ATPase. Direct measurement of the activity of Na+, K+-ATPase revealed a two-fold stimulation of the enzyme by glucagon. The effect of glucagon on the activity of the enzyme was immediate. Simultaneously with the increase in the activity of the Na+, K+-ATPase, the activity of Mg2+-ATPase decreased. In order to evaluate whether the activation of the Na+, K+-ATPase by glucagon is related to the metabolic effects of the hormone, experimental conditions known to interfere with the activity of the enzyme were employed and glucagon stimulation of Ca2+-efflux, mitochondrial metabolism and gluconeogenesis were measured. K+-free perfusate, high K+ perfusate or ouabain interfered to varying degrees with the glucagon stimulation of these responses. The combination of K+-free perfusate and ouabain almost completely abolished the glucagon stimulation of all three parameters. These results demonstrate the glucagon stimulation of Na+, K+-ATPase and raise the possibility that the activation of the enzyme by glucagon might be a necessary link for the manifestation of its metabolic effects.

Animals↗
PubMed1982

Calcium uptake in isolated hepatic plasma-membrane vesicles.

A liver plasma-membrane fraction capable of Ca2+ uptake was isolated. The fraction exhibited high Na+, K+-ATPase, low glucose-6 phosphatase activity, and transported alanine in a Na+-dependent fashion. The uptake of Ca2+ was ATP-dependent; UTP, GTP, or CTP did not substitute for ATP. The presence of oxalate did not significantly alter the rate of uptake. The pH optimum of the reaction was basic (no uptake was visible at pH 6.8). These properties are at variance with those of the endoplasmic reticulum Ca2+ uptake system, which is oxalate-dependent, and has an acid pH optimum. The ATP-dependent Ca2+ uptake has a Km(Ca2+) of 1.4 X 10(-8) M and a Vmax of transport of 30 nmol X mg protein-1 X min-1. No conclusive results were obtained on the calmodulin-sensitivity of the process: addition of calmodulin to the vesicles did not stimulate uptake, and the anti-calmodulin drug trifluoperazine had no inhibitory effect. However, another anti-calmodulin drug (R24571) had a limited, but statistically significant, inhibitory action. A partial release of the accumulated Ca2+ from the vesicles could be induced by the addition of Na+, and incubation of the vesicles in a high Na+ medium (as compared to high K+ medium) resulted in lower (about 25%) calcium uptake. Partial release of the accumulated Ca2+ could be induced also by the addition of H+. The releasing effect of H+, taken together with the absence of Ca2+ uptake at acid pH, suggests the possibility of a H+/Ca2+ exchange.

Animals↗