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Biomedical subjects

R H Michell

Publications and source records attributed to R H Michell.

At least 145 records · Page 8Linked to original sources
PubMed1978

A calcium-activated polyphosphoinositide phosphodiesterase in the plasma membrane of human and rabbit erythrocytes.

Haemoglobin-free human erythrocyte ghosts that were prepared in the presence of EDTA and were then exposed to Ca2+ showed a substantial loss of phosphatidylinositol phosphate and phosphatidylinositol diphosphate, measured either chemically or by loss of 32P from the lipids of prelabelled membranes. At the same time there was, as reported previously (Allan, D. and Michell, R.H., (1976) Biochim. Biophys. Acta 455, 824--830), and approximately equivalent rise in the diacylglycerol content of the membranes. Analysis of the 32P-labelled water-soluble material released during this process showed that the major products were inositol diphosphate and inositol triphosphate. No change was seen in the phosphatidylinositol or phosphatidate content of the membranes, and there was no Ca2+-activated loss of 32P from the phosphatidate of prelabelled membranes: this suggests that Ca2+ did not activate phosphoinositide phosphomonoesterases or phosphatidate phosphomonoesterase in human erythrocyte membranes. It is concluded that human erythrocyte membranes contain at their cytoplasmic surface a Ca2+-activated phosphodiesterase that is active against both phosphatidylinositol phosphate and phosphatidylinositol diphosphate. Rabbit erythrocytes also contained this enzyme, but in these cells there was also evidence for the presence of a Ca2+-activated phosphatidate phosphomonoesterase.

Animals↗
PubMed1977

Calcium ion-dependent diacylglycerol accumulation in erythrocytes is associated with microvesiculation but not with efflux of potassium ions.

Erythrocytes from several different species were exposed to Ca2+ and the bivalent-cation ionophore A23187. The lipid composition, morphology and K+ permeability of the treated cells were investigated. Erythrocytes from human, rat, guinea pig and rabbit (a) showed an increased concentration of 1,2-diacyl-sn-glycerol and enhanced labelling of phosphatidate with 32P, (b) underwent echinocytosis and outward vesiculation, and (c) rapidly released much of their intracellular K+. Pig cells showed only the K+ loss, and ox and sheep (high-K+) cells showed none of these Ca2+-evoked effects. All of the cells underwent stomatocytosis and inward vesiculation when treated externally with Clostridium perfringens phospholipase C. These results support the idea that there is a correlation between the asymmetric insertion of diacylglycerol (or ceramide) into the membrane and the shape-changes leading to microvesiculation, but they indicate that Ca2+-triggered K+ efflux and diacylglycerol production are unrelated events. Erythrocytes of chicken and turkey showed no Ca2+-stimulated K+ efflux. They showed slight ionophore A23187-stimulated vesiculation, but this appeared to be associated with the appearance in the membrane of ceramide rather than of diacylglycerol. Phospholipase C treatment caused very similar changes in morphology and phosphatidate labelling to those seen in mammalian erythrocytes.

Animals↗
PubMed1977

A comparison of the effects of phytohaemagglutinin and of calcium ionophore A23187 on the metabolism of glycerolipids in small lymphocytes.

1. The effects of phytohaemagglutinin and of a Ca2+ ionophore (A23187) on glycerolipid metabolism in lymphocytes from pig lymph nodes were compared (a) by studying the incorporation of [32P]Pi and [3H]glycerol, and (b) by following the redistribution of [3H]glycerol among the lipids caused by these agents in pulse-chase experiments. 2. Phytohaemagglutinin only stimulated 32P incorporation into phosphatidylinositol and, to a slight extent, phosphatidate. Removal of most of the extracellular Ca2+ somewhat decreased this response. 3. Ionophore A23187 stimulated the labelling of phosphatidate and phosphatidylinositol with 32P to a much greater extent than did phytohaemagglutinin: the increase in phosphatidate labelling, but not that of phosphatidylinositol, was almost abolished by the removal of extracellular Ca2+. 4. The combined effects of phytohaemagglutinin and ionophore appeared to be additive, rather than synergistic. 5. Treatment with ionophore A23187 somewhat decreased the total incorporation of [3H]glycerol into glycerolipids, possibly because it lowered cell ATP content. In these experiments di- and tri-acylglycerol behaved anomalously, triacylglycerol labelling being suppressed completely, whereas that of diacylglycerol was enhanced. The pulse-chase results revealed that triacylglycerol was converted into diacylglycerol in the ionophore-treated cells, and the availability of this diacylglycerol probably led to the enhanced labelling of phosphatidate and phosphatidylinositol in the these cells. 6. Thus an increase in intracellular Ca2+ concentration appeared to have three effects on glycerolipid metabolism: (a) slight inhibition of some metabolic step preceding phosphatidate synthesis, (b) inhibition of diacylglycerol acyltransferase and (c) activation of a triacylglycerol lipase. 7. In contrast, it seems likely that the only effect of phytohaemagglutinin is to stimulate phosphatidylinositol breakdown. 8. Pig polymorphonuclear leucocytes treated with ionophore A23187 showed metabolic changes that were similar to those demonstrated with lymphocytes. 9. A possible similarity is suggested between Ca2+-stimulated triacylglycerol lipase in lymphocytes and polymorphonuclear leucocytes and previous observations of enhanced triacylglycerol metabolism in stimulated cells whose metabolic functions involve membrane fusion.

Animals↗
PubMed1977

Permeability characteristics of erythrocyte ghosts prepared under isoionic conditions by a glycol-induced osmotic lysis.

A detailed study has been made of the permeability characteristics of human erythrocyte ghosts prepared under isoionic conditions by a glycol-induced lysis (Billah, M.M., Finean, J.B., Coleman, R. and Michell, R.H. (1976) Biochim. Biophys. Acta 433, 45-54). Impermeability to large molecules such as dextran (average molecular weight 70 000) was restored immediately and spontaneously after each of the 5-7 lyses that were required to remove all of the haemoglobin. Permeabilities to smaller molecules such as MgATP2-, [3H]inositol and [14C]choline were initially high but could be greatly reduced by incubation at 37 degrees C for an hour. The extent of such resealing decreased as the number of lyses to which the ghosts had been subjected increased. Both removal of haemoglobin and permeabilities to small molecules were affected significantly by pH, CA3+ concentrations and divalent cation chelators. Maximum resealing was achieved in ghosts prepared in the basic ionic medium (130 mM KCl, 10 nM NaCl, 2 mM MgCl2, 10 mM N-2-hydroxyethylpiperazine-N'-2-ethanesulphonic acid (HEPES)) at pH 7.0 (0 degrees C) and with a calcium level around 10(-5) M. Acidic pH facilitated the removal of haemoglobin whilst the presence of divalent cation chelators showed down its release. Retention of K+ by ghosts leaded with K+ during the first lysis and subsequently incubated at 37 degrees C was substantial but lation chelators slowed down its released. Retention of K+ by ghosts loaded with K+ during the first lysis and subsequently incubated at 37 degrees C was substantial but little K+ could be retained within the haemoglobin-free ghosts. Permeability of the ghosts to K+ after one lysis was affected by temperature, pH, Ca2+ concentrations and by the presence of divalent cation chelators.

Adenosine Triphosphate↗
PubMed1976

Production of 1,2-diacylglycerol in human erythrocyte membranes exposed to low concentrations of calcium ions.

A specific increase in the membrane content of 1,2-diacylglycerol occurred when erythrocytes were lysed at 20 degrees C in media which did not inclued a chelator of Ca2+ and also when Ca2+ was added to haemoglobin-free erythrocyte ghosts which had been prepared in the presence of ethyleneglycol-bis-(beta-aminoethylether)-N,N'-tetraacetic acid (EGTA). The maximum increase was about 20-fold. The production of 1,2-diacylglycerol appeared to be caused by an endogenous membrane-bound phospholipase C which was half-maximally activated at less than 1 muM Ca2+ and which had access to only about 0.6-0.8% of the cells' glycerolipids. This activity was optimal at pH 7.0-7.2 in the presence of 0.1 mM Ca2+; under these conditions diacylglycerol production was complete within 5-10 min. Enzyme activity was markedly decreased at low temperatures, and was abolished by heating at 100 degrees C for 1 min.

Calcium↗
PubMed1976

Effects of calcium-antagonistic drugs on the stimulation by carbamoylcholine and histamine of phosphatidylinositol turnover in longitudinal smooth muscle of guinea-pig ileum.

A number of drugs classed as calcium antagonists, spasmolytics, non-specific receptor antagonists or receptor antagonists with multiple sites of action were tested to determine whether they prevent the stimulation of phosphatidylinositol turnover caused in various tissues by the activation of receptors which increase cell-surface Ca2+ permeability. The experiments were done with fragments of longitudinal smooth muscle from guinea-pig ileum; these were incubated in vitro with 32Pi and either 100 muM-carbamoylcholine or 100 muM-histamine, in the presence of antagonistic drugs at concentrations at least sufficient to cause complete blockade of smooth-muscle contraction. The phosphatidylinositol response to carbamoylcholine was not changed by cinchocaine, papaverine, nifedipine, dibenamine, amethocaine, cinnarizine, lidoflazine, methoxyverapamil, prenylamine or two antimuscarinic alkane-bis-ammonium compounds, and the response to histamine was unaffected by the first four drugs. In contrast, phenoxybenzamine prevented the increase in phosphatidylinositol labelling caused by either carbamoylcholine or histamine. The insensitivity of the phosphatidylinositol response to most of the drugs provides further experimental support for the conclusion that the receptor-stimulated phosphatidylinositol breakdown which initiates the increase in phosphatidylinositol turnover is not caused by an increase in intracellular Ca2+. The simplest interpretation of the available information appears to be that phosphatidylinositol breakdown plays a role in the coupling between the receptor-agonist interaction and the opening of cell-surface Ca2+ gates [Michell, R. H. (1975) Biochim. Biophys. Acta 415, 81-147]. If this is correct, then phenoxybenzamine must exert its inhibitory effects on phosphatidylinositol breakdown early in this sequence of events, but the drugs must act at a stage later than phosphatidylinositol breakdown. The unexpected difference in the effects of dibenamine and phenoxybenzamine, which are chemically very similar, may provide a useful experimental tool with which to explore the way in which activated receptors provoke the opening of cell-surface Ca2+ gates.

Anesthetics, Local↗