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Biomedical subjects

H Rasmussen

Publications and source records attributed to H Rasmussen.

At least 199 records · Page 11Linked to original sources
PubMed1982

Studies on the Ca2+ transport mechanism of human erythrocyte inside-out plasma membrane vesicles. V. Chlortetracycline fluorescence.

The measurement of chlortetracycline fluorescence was employed as a probe for measuring the process to calcium transport by human erythrocyte inside-out vesicles. Chlortetracycline is a divalent metal chelator which increases its fluorescence when bound to calcium in the presence of a membrane. Addition of calcium and ATP to inside out vesicles in the presence of chlortetracycline increased the chlortetracycline fluorescence as a function of time following an initial delay. Only after a threshold level of calcium had been accumulated did the fluorescence increase. The presence of both ATP and calcium were required. The addition of calmodulin increased the rate and absolute magnitude of the chlortetracycline fluorescence change. Similarly, calmodulin stimulated the rate and extent of 45Ca transport by inside-out vesicles. Moreover, the presence of saponin abolished both chlortetracycline fluorescence change and 45Ca uptake; a non-hydrolyzable ATP analog would not substitute for ATP in either 45Ca transport or chlortetracycline fluorescence experiments. Comparison between the slopes of the linear portions of chlortetracycline fluorescence change and calcium transport time courses at varied free calcium concentrations showed a consistent ratio between the slopes. This suggests that calcium transport change can be calibrated by employing chlortetracycline fluorescence. Based on this data, it is concluded that chlortetracycline fluorescence is a rapid and accurate method for monitoring calcium transport by human erythrocyte inside-out vesicles.

Biological Transport, Active↗
PubMed1982

Role of changes in membrane lipid structure in the action of 1,25-dihydroxyvitamin D3.

Recent studies have shown that 1,25-dihydroxyvitamin D3 (1,25-(OH)2-D3) stimulates the entry of calcium into the duodenal mucosal cell of the chick by a mechanism that does not require the synthesis of new protein. Using isolated brush border membrane vesicles (BBMV) from these cells, we have explored the mechanism by which 1,25-(OH)2-D3 acts. Administration of the hormone leads to an increase in calcium uptake into BBMV. This calcium uptake is a saturable process. Addition of the methyl ester of cis-vaccenic acid to BBMV in vitro leads to a specific increase in calcium uptake into vesicles from vitamin D-deficient chicks but not in those from 1,25-(OH)2-D3-treated chicks. Administration of 1,25-(OH)2-D3 leads to an increase in the de novo synthesis of phosphatidylcholine (PC) and an increase in the total PC content of the brush border membrane. It also increases the turnover of fatty acids into PC, which results in an increase in the content of polyunsaturated fatty acids in the PC fraction. These changes in lipid structure and turnover either precede in time or occur simultaneously with the change in calcium transport rate, and neither is blocked by the administration of cycloheximide. It is proposed that the primary mechanism by which 1,25-(OH)2-D3 regulates calcium transport across the luminal membrane of the enterocyte is by inducing a specific alteration in membrane PC content and structure, which leads to an increase in membrane fluidity and thereby to an increase in calcium transport rate.

Animals↗
PubMed1981

Phenothiazines and related compounds disrupt mitochondrial energy production by a calmodulin-independent reaction.

Phenothiazines and related compounds bind to mitochondrial membranes in approximate proportion to their affinities for calmodulin. Penfluridol (16 microM), pimozide (20 microM), or trifluoperazine (66 microM) completely inhibit ADP-stimulated respiration in isolated rat liver mitochondria, but exert no effect on either uncoupler- or Ca2+-stimulated respiration. The inhibition of ADP-stimulated respiration results from inhibition of the oligomycin-sensitive ATPase. Inhibition of the ATPase does not involve interaction of phenothiazine with calmodulin. The addition of calmodulin with or without calcium to mitochondrial inner membrane preparations has no effect on ATPase activity. The addition of EGTA and the ionophore A23187 prior to the addition of phenothiazine does not prevent the phenothiazine-induced inhibiton of the ATPase. Measurements of inner membrane calmodulin content by gel electrophoresis or cyclic nucleotide phosphodiesterase activation are negative. Despite the absence of calmodulin in the inner membrane preparations, 12.5 nmol trifluoperazine bind per 100 microgram of membrane protein with an association constant, K, of 6.5 . 10(4) M-1. We conclude that calmodulin-binding neuroleptic agents, when added to whole cells, have the potential to disrupt mitochondrial energy production by a reaction which apparently does not involve a phenothiazine-calmodulin interaction.

Adenosine Triphosphatases↗
PubMed1981

Effect of 1,25-dihydroxyvitamin D3 on phospholipid metabolism in chick duodenal mucosal cell. Relationship to its mechanism of action.

Pretreatment of the D-deficient chick with 1,25-dihydroxyvitamin D3 increases de novo synthesis of phosphatidylcholine by a stimulation of CDP-choline: sn-1,2-diacylglycerol choline-phosphotransferase reaction. The time course of change in the incorporation of [3H]choline and [14C]ethanolamine into the brush border lipid fraction after 1,25-dihydroxyvitamin D3 treatment correlates closely with the time course of change in calcium uptake into the brush border membrane vesicles. Prior treatment with cycloheximide does not block this increase in phosphatidylcholine synthesis. In addition, 1,25-dihydroxyvitamin D3 administration increases the incorporation of [3H]arachidonic acid into the phosphatidylcholine fraction of the brush border to a great extent but does not increase the incorporation of [3H]palmitic acid into the phosphatidylcholine fraction. The incorporation of these 3H labeled fatty acids into diacylglycerol is not changed by 1,25-dihydroxyvitamin D3. These data indicate that 1,25-dihydroxyvitamin D3 enhances the synthesis of phosphatidylcholine independent of new protein synthesis, and also increases the incorporation of unsaturated fatty acids into phosphatidylcholine. From these results we suggest that changes in phospholipid metabolism in the enterocyte are the mechanisms by which 1,25-dihydroxyvitamin D3 acts to enhance calcium entry across the brush border membrane.

Animals↗
PubMed1981

Regulation of cyclic AMP metabolism in the rat erythrocyte during chronic beta-adrenergic stimulation. Evidence for calmodulin-mediated alteration of membrane-bound phosphodiesterase activity.

The regulation of cyclic AMP metabolism in the rat erythrocyte has been investigated during chronic exposure to the beta agonist isoproterenol. A triphasic response is observed: 1) an acute increase in cyclic AMP to levels four- to fivefold greater than basal, maximal by 1 minute (Phase I); 2) a gradual decline in cAMP content to levels near basal during the next 15-20 minutes (Phase II) and a second sustained rise in cAMP, maximal by 60 minutes, to a concentration greater than that observed during the first minute (Phase III). Extensively washed Phase II and Phase III cells are refractory to a second challenge by isoproterenol. In phosphodiesterase-inhibited intact Phase II and III cells adenylate cyclase activity is maximally activated. Isoproterenol has no effect on soluble phosphodiesterase activity but increases membrane-bound phosphodiesterase activity 3- and 2.2-fold in Phase II and Phase III cells, respectively. The activation of this membrane-bound enzyme activity appears to be mediated by the calcium-dependent regulatory protein, calmodulin, because 1) the amount of exogenous calmodulin required to achieve half-maximal activation of membrane-bound phosphodiesterase is 3.7, 2.0, and 1.2 micrograms in control, Phase III and Phase II membranes, respectively; and 2) there is less calmodulin in membrane-free lysates prepared from Phase II cells than control cells. These data support the idea that the major mechanism regulating cAMP content in the rat erythrocyte during chronic isoproterenol stimulation is the membrane-bound phosphodiesterase and that there is a translocation of calmodulin from the cytoplasm to the membrane during hormone stimulation.

3',5'-Cyclic-AMP Phosphodiesterases↗
PubMed1981

Studies of the Ca2+ transport mechanism of human erythrocyte inside-out plasma membrane vesicles. I. Regulation of the Ca2+ pump by calmodulin.

Calcium accumulation by human erythrocyte inside-out vesicles was linear for at least 30 min in the presence of ATP. In untreated inside-out vesicles, 3.76 +/- 1.44 nmol of calcium/min/unit of acetylcholinesterase were transported, compared with 10.57 +/- 2.05 (+/- S.D.; n = 11) in those treated with calmodulin. The amount of calmodulin necessary for 50% activation of Ca2+ accumulation was 60 +/- 22 ng/ml (+/- S.D.; n = 4). The Km (Ca2+) for calmodulin-stimulated accumulation was 0.8 +/- 0.05 microM (+/- S.D.; n = 5) using Ca2+ /ethylene glycol bis(beta-aminoethyl ether)N,N,N',N'-tetraacetic acid (EGTA) buffers, or 25 microM with direct addition of unbuffered calcium. In the absence of calmodulin, these values were 0.4 and 60 microM, respectively, Km (ATP) values of 90 and 60 microM in the presence and absence of calmodulin, respectively, were measured at constant magnesium concentration (3 mM). In the presence of calmodulin, a broad pH profile is exhibited from pH 6.6 to 8.2. Maximal calcium accumulation occurs at pH 7.8. In the absence of calmodulin, the pH profile exhibits a linear upward increase from pH 7.0 to 8.2. The (Ca2+-Mg2+)-ATPase activity, measured under identical conditions, was 2.40 +/- 0.72 nmol of Pi/min/unit of acetylcholinesterase in the untreated vesicles and 11.29 +/- 2.87 nmol of Pi/min/unit of acetylcholinesterase (+/- S.D.; n = 4) in calmodulin-treated vesicles. A stoichiometry of 1.6 Ca2+/ATP hydrolyzed was determined in the absence of calmodulin; in the presence of calmodulin, this ratio was decreased to 0.94 Ca2+/ATP hydrolyzed.

Adenosine Triphosphatases↗
PubMed1981

Long-term treatment of familial hypophosphatemic rickets with oral phosphate and 1 alpha-hydroxyvitamin D3.

Combined treatment with oral phosphate and 1 alpha (OH)D3 was carried out in nine children with familial hypophosphatemic rickets. All nine had positive responses over a four- to six-year period as judged by healing of rickets, change in growth rate, decrease in alkaline phosphatase activity, and symptomatic improvement. In two patients therapy was stopped for a short time because of hypercalcemia. In one patient in whom therapy was effective there was a significant reduction in creatinine clearance which necessitated cessation of treatment. The results of this study suggest that combined treatment with 1 alpha(OH)D3 and oral phosphate is an effective form of therapy for this condition, but that the balancing of these two modalities of therapy in each patient is essential if hypercalcemia and hypercalciuria, on the one hand, and secondary hyperparathyroidism, on the other, are to be avoided. A simple means of balancing these therapeutic modalities is suggested.

Administration, Oral↗
PubMed1981

Liponomic control of Ca2+ transport: relationship to mechanism of action of 1,25-dihydroxyvitamin D3.

Isolated vesicles prepared from the brush border membranes of chicken duodenal mucosal cells (enterocytes) take up calcium by a passive but saturable process. The rate of uptake (Vmax) is increased 2.5- to 3-fold, with no change in Km, in vesicles prepared from 1,25-dihydroxyvitamin D3 [1,25(OH)2D3]-treated chickens compared to vesicles from vitamin D-deficient controls. Preincubation of vesicles with either cis- or trans-vaccinic acid (cVA or tVA, respectively) or their methyl esters in vitro also alters the rates of calcium transport. Methyl cVA causes an increase in rate of calcium uptake into vesicles from vitamin D-deficient chickens but not in those from 1,25(OH)2D3-treated chickens. This increase is 80-90% of that seen after 1,25(OH)2D3 treatment. Higher concentrations of methyl cVA produce no further increases. Conversely, methyl tVA causes a decrease in rate of calcium uptake in vesicles from 1,25(OH)2D3-treated chickens but no change in vesicles from vitamin D-deficient controls. This decrease reduces the rate of calcium uptake to nearly the same value as seen in vesicles from vitamin D-deficient controls. Higher concentration of methyl tVA produce no further suppression of uptake rate. The changes seen were in the Vmax and not in the Km of the transport process. The fatty acids did not alter the process of Na+-dependent glucose uptake in the same membrane. These data demonstrate that a small alteration in the lipid structure of this membrane can specifically shift the activity of the calcium transport process.

Animals↗