Influence of ethanol on calcium,inositol phospholipids and intracellular signalling mechanisms

Summary Studies have implicated Ca⁺⁺ in the actions of ethanol at many biochemical levels. Calcium as a major intracellular messenger in the central nervous system is involved in many processes, including protein phosphorylation enzyme activation and secretion of hormones and neurotransmitters. The control of intracellular calcium, therefore, represents a major step by which neuronal cells regulate their activities. The present review focuses on three primary areas which influence intracellular calcium levels; voltage-dependent Ca⁺⁺ channels, receptor-mediated inositol phospholipid hydrolysis, and Ca⁺⁺/Mg⁺⁺-ATPase, the high affinity membrane Ca⁺⁺ pump.Current research suggests that a subtype of the voltage-dependent Ca⁺⁺ channel, the dihydropyridine-sensitive Ca⁺⁺ channel, is uniquely sensitive to acute and chronic ethanol treatment. Acute exposure inhibits, while chronic ethanol exposure increases⁴⁵Ca⁺⁺-influx and [³H]dihydropyridine receptor binding sites. In addition, acute and chronic exposure to ethanol inhibits, then increases Ca⁺⁺/Mg⁺⁺-ATPase activity in neuronal membranes. Changes in Ca⁺⁺ channel and Ca⁺⁺/Mg⁺⁺-ATPase activity following chronic ethanol may occur as an adaptation process to increase Ca⁺⁺ availability for intracellular processes. Since receptor-dependent inositol phospholipid hydrolysis is enhanced after chronic ethanol treatment, subsequent activation of protein kinase-C may also be involved in the adaptation process and may indicate increased coupling for receptor-dependent changes in Ca⁺⁺/Mg⁺⁺-ATPase activity.The increased sensitivity of three Ca⁺⁺-dependent processes suggest that adaptation to chronic ethanol exposure may involve coupling of one or more of these processes to receptor-mediated events.     

Serotonin-stimulated protein phosphorylation in aortic smooth muscle cells

Summary The effects of serotonin on the formation of inositol phosphates and protein phosphorylation were examined in cultured smooth muscle cells. Serotonin stimulated the formation of [³H]inositol monophosphate, [³H]inositol bisphosphate and [³H]inositol trisphosphate. This effect was prevented by 5-HT₂ specific antagonist, 6-methyl-1-(1-methylethyl)ergoline-8-carboxylic acid, 2-hydroxy-1-methylpropyl ester [Z]-2-butenedioate (LY53857). Serotonin stimulated the phosphorylation of many polypeptides, among which a 20 kDa polypeptide was the most prominent. The phosphorylation was also inhibited by LY53857. LY53857 alone produced no effects on protein phosphorylation. The 20 kDa polypeptides were also phosphorylated by the addition of 12-O-tetradecanoylphorbol-13-acetate. These results suggest that serotonin stimulates protein phosphorylation through 5-HT₂ receptors and possibly activates protein kinase C in intact vascular smooth muscle cells.Part of the data contained in this paper was presented at the 74th local meeting of the Japanese Society of Pharmacology at Kanagawa.     

Role of calcium in the histamine release induced by D-galactosamine from rat mast cells

Summary Rat peritoneal mast cells were isolated and purified by differential centrifugation in Ficoll. Cells pooled from three to four rats were suspended at approximately 10⁶ cells/ml in a buffered salt solution and incubated for 1 h at 37°C in 300 l volumes in the absence or presence (9×10^–4 M) of calcium chloride. Addition of D-galactosamine hydrochloride (DGM; 2.8×10^–4 M) caused (in addition to basal release) a mean ±SEM percent histamine release of 15.7±5.2 in the presence of Ca⁺⁺ and 19±4.9 in the absence of Ca⁺⁺ (p>0.05). It is suggested that D-galactosamine does not require extracellular Ca⁺⁺ for the release of histamine from the rat mast cell.A preliminary analysis of these results was presented at the International Symposium on calcium entry blockers and tissue protection, Rome, 15–16 March 1984.     

Effects of prostaglandin E1 on45Ca++-incorporation and spike activity in longitudinal smooth muscle of cat jejunum

Summary Prostaglandin E₁ (0.3 M) decreased both the⁴⁵Ca⁺⁺-incorporation and the spike activity in isolated longitudinal smooth muscle preparations of the cat jejunum probably by an inhibition on the Ca⁺⁺ influx.     

Possible role of intestinal alkaline phosphatase activity in thiamine transport

Summary Thiamine deficiency caused a marked decrease of intestinal alkaline phosphatase (al-Pase) activity, but had no effect on the Ca⁺⁺-ATPase activity and Ca⁺⁺-absorption in rats. The al-Pase activity was significantly decreased 1 h after oral administration of ethanol at 0.5 and 2.5 g/kg. In contrast, Mg⁺⁺-, Ca⁺⁺- and (Na⁺+K⁺)-ATPase activities did not change after the administration of ethanol. These findings show that the al-Pase activity, unlike the Ca⁺⁺-ATPase activity, is not related to Ca⁺⁺-absorption. A possible role of al-Pase activity in the active transport of thiamine in the intestine was discussed.     

Unterschiede zwischen Tyramin und Dimethylphenylpiperazin in der Ca++-Abhängigkeit und im zeitlichen Verlauf der Noradrenalin-Freisetzung am isolierten Kaninchenherzen

Summary On the perfused rabbit heart a constant infusion of tyramine released noradrenaline continuously and independently of the external Ca⁺⁺ concentration. In contrast, noradrenaline release by DMPP was only transient and required the presence of Ca⁺⁺.

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Mit Unterstützung der Deutschen Forschungsgemeinschaft.     

Ambiguous effect of caffeine upon the transmembrane Ca current in mammalian ventricular myocardium

Zusammenfassung Coffein (20 mM) wirkt auf den transmembranären Ca⁺⁺-Strom der Warmblütermyokardfaser biphasisch. Zunächst nimmt der Ca⁺⁺-Strom zu, was auf einer Verbesserung der Ca⁺⁺-Leifähigkeit des langsamen Membrankanals beruht. Nach längerer Einwirkungsdauer von Coffein kommt es dagegen zu einer Abnahme des Ca⁺⁺-Stroms. Als Ursache hierfür wird vor allem eine Verminderung des transmembranären Konzentrationsgradienten für Ca⁺⁺ — infolge eines Anstiegs der freien Ca⁺⁺-Konzentration im Zellinnern — postuliert.     

Effect of trifluoperazine and calcium ions on gregarine gliding

Summary The drug trifluoperazine (TFP) inhibits the gliding motility of gregarine protozoans. This suggests that the Ca⁺⁺ binding protein, calmodulin, plays a role in the motility process. However the presence of extracellular Ca⁺⁺ was not required for gliding to occur.Acknowledgments. We should like to thank L. Cooper for technical assistance, Smith, Kline and French for the gift of trifluoperazine, and Terry Preston for valuable discussion.     

Experiments on mitochondrial swelling in vivo

Riassunto E stato provato l'effetto sui mitocondri in vivo dei seguenti composti che producono rigonfiamento dei mitocondri isolati in vitro:l-cistina,l-cisteina, glutatione, L-ascorbato,l-tiroxina,l-3, 5, 3-triiodotironina, fosfato e Ca⁺⁺. Solo il Ca⁺⁺ ha provocato un aumento di volume dei mitocondri nelle cellule provate, i mioblasti coltivati in vitro.     

Die Abhängigkeit der Ca++-Aufnahme isolierter Mitochondrien des Herzmuskels von der Na+-und K+-Konzentration als mögliche Ursache der inotropen Digitaliswirkung

Summary In isolated mitochondria of heart muscle from rabbits and oxen there is, under suitable conditions, an accumulation of Ca⁺⁺, which is significantly enhanced by elevating the K⁺/Na⁺ quotient of the incubation medium. K-strophanthine (10^–5–10^–7) does not influence the accumulation of Ca⁺⁺ by the mitochondria of heart muscle. Therefore the intracellular increase in exchangeable Ca⁺⁺ observed after digitalis-glycosides could be explained by a decrease of the intracellular K⁺/Na⁺ quotient, which is caused by inhibition of the membrane ATPase and diminishes the capacity for Ca⁺⁺ accumulation in mitochondria.     

Calcium and magnesium in plant cytokinesis and their antagonism with caffeine

Summary The efficiency of caffeine at different concentration on the induction of binucleate cells in onion root-tip was studied. The drug effect is strongly depressed in the Ca⁺⁺ and/or Mg⁺⁺ presence at half-rate of maximum efficiency (0.04%), about 2 mM). We therefore conclude that both cations must play a role in plant cytokinesis.     

Influence of bistramide A on the twitch tension in rat atrial heart muscle

Bistramide A, a new toxin isolated from the UrochordateLissoclinum bistratum Sluiter, was applied to rat auricular heart muscle bundles. At a stimulation frequency of 0.2 Hz, the toxin induces a dose-dependent reduction of the stimulated twitch tension force; it decreases and shortens the duration of the plateau and the slow repolarizing phase of the action potential. In the control solution, switching from a stimulation frequency of 0.2 Hz to 1 Hz decreases the force with which a positive potentiation develops either at a maintained high frequency or after switching from 1 Hz to 0.2 Hz. Bistramide A reduces both the force evoked at 1 Hz and the potentiation. The data suggest that Bistramide A blocks Na⁺ conductance; inhibits Ca⁺⁺ channels in a time-and frequency-dependent manner; reduces Na⁺–Ca⁺⁺ exchange activity; but does not modify the ability of the sarcoplasmic reticulum to be refilled although the rate of Ca⁺⁺ accumulation is decreased.     

The reaction of Ca++ with the inner and outer membrane of mitochondria

Riassunto È stata studiata la distribuzione del transporto attivo del Ca⁺⁺ e dei siti ad alta e bassa affinità per il Ca⁺⁺ tra la membrana esterna e interna dei mitocondri di fegato. La membrana interna trasporta attivamente il Ca⁺⁺, la membrana esterna non è in grado di farlo. Ambedue le membrane posseggono i siti a bassa affinità, mentre i siti ad alta affinità si trovano solo sulla membrana interna.     

Magnesium-calcium antagonism in the contraction of arterioles

Zusammenfassung Erhöhung des Mg-Spiegels im Blut hemmt Tonus und Kontraktilität der Widerstandsgefässe im Muskel. Durch Noradrenalin hervorgerufene Verengerung wird gleich stark gehemmt wie die durch Reizung sympathischer Nerven verursachte Kontraktion. Ein Überschuss an Ca⁺⁺ verhindert die blockierende Wirkung des Mg⁺⁺ auf die Kontraktilität, der Tonusverlust der Gefässe wird durch Ca⁺⁺ jedoch nicht rückgängig gemacht. Die antagonistischen Wirkungen von Mg⁺⁺ und Ca⁺⁺ beruhen wahrscheinlich auf deren gegensätzlicher Wirkung auf die elektromechanische Kopplung im Gefässmuskel.

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U.S. Public Health Service International Fellow.     

The affinity of mitochondria for Ca++

Riassunto E' stata studiata l'affinità dei mitocondri di fegato di ratto e di cuore di coniglio per il Ca⁺⁺, nel sistema dipendente da energia. Si sono usati tamponi EGTA-Ca⁺⁺ per mantenere la concentrazione del Ca⁺⁺ costante durante l'esperimento. Si è dimostrato che laK _m apparente per il Ca⁺⁺ non è lontana da 10^–6 M.

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We are indebted to Dr.B. Chance for many stimulating discussions and for facilitating the progress of the work with the generous hospitality in his Institute.     

ATPase activity of guinea pig heart muscle

Riassunto L'Autore ha studiato l'influenza della concentrazione degli ioni Ca⁺⁺ e Mg⁺⁺ sull'ATPasi degli atri e dei ventricoli di cuore di cavia. Tale attività è maggiore a pH 9,2 che a pH 6,8. Mentre il Ca⁺⁺ influisce differenziando decisamente le attività degli atri da quelle dei ventricoli, il Mg⁺⁺ accentua piuttosto una differenza tra la parte destra e la sinistra. Il calore distrugge quasi completamente l'attività enzimatica dopo una permanenza a 60° per 20 min.     

A comparative study of the intracellular Ca++ movements in white and red muscle

Riassunto Dopo iniezione di⁴⁵Ca⁺⁺ in conigli, radioattività appare nei mitocondri e nel reticolo sarcoplasmico del massetere dell'adduttore magno. La quantità di radioattività, e l'attività specifica del Ca⁺⁺, sono molto più elevate nei mitocondri che nel reticolo sarcoplasmico del massetere. Sono dello stesso ordine di grandezza nei due organelli dell'adduttore magno. I risultati suggeriscono un ruolo prominente dei mitocondri nei movimenti intracellulari del Ca⁺⁺ nel muscolo rosso, ed un analogo prominente ruolo del reticolo sarcoplasmico nel muscolo bianco.     

Studies of total and protein-bound plasma Mg++ in wild and hatchery-reared coho salmon smolts in freshwater and in seawater

Summary Total plasma Mg⁺⁺ and Ca⁺⁺, Mg⁺⁺ in erythrocytes as well as protein-bound plasma Mg⁺⁺ were investigated in wild and hatchery-reared smolts. The proportion of plasma Mg⁺⁺ which was bound to plasma protein did not change significantly during entry into seawater, even though the in vitro addition of exogenous Mg⁺⁺ to the plasma showed that additional binding was possible.     

The effects of the ‘calcium-antagonist’, prenylamine,on the action potential of crayfish muscle (Oronectes virilis)

Zusammenfassung Der organische Ca⁺⁺-Antagonist Prenylamin blockiert die leicht reversiblen Ca⁺⁺-Aktions-potentiale in der Muskelfaser des FlusskrebsesOronectes virilis. Etwas niedrigere Konzentrationen erhöhen die Schwelle und verlängern die Dauer, während schwache Konzentrationen von Prenylamin nur wirksam waren, wenn dieses verwendet wurde, bevor der Muskel dem Aktionspotentialerreger Procain oder Sr⁺⁺ ausgesetzt war.     

A specific endogenous inhibitor of two forms of Ca++ activated neutral proteases in platelets

Summary An endogenous, specific inhibitor of high molecular weight has been isolated from bovine blood platelets, which inhibits the activity of the 2 forms of platelet Ca⁺⁺ activated neutral proteases reported previously by us. The inhibition is not due to chelation of Ca⁺⁺ but results from a stoichiometric complex formation.