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Biomedical subjects

K E Akerman

Publications and source records attributed to K E Akerman.

At least 109 records · Page 6Linked to original sources
PubMed1985

Interaction of Cd2+ with the calmodulin-activated (Ca2+ + Mg2+)-ATPase activity of human erythrocyte ghosts.

Treatment of erythrocyte ghosts with micromolar concentrations of Cd2+ results in a noncompetitive inhibition of the calmodulin-dependent (Ca2+ + Mg2+)-ATPase activity. Higher concentrations of Cd2+ are required for inhibition of the (Ca2+ + Mg2+)-ATPase activity of calmodulin-depleted ghosts. The interaction of Cd2+ is time-dependent with an apparent rate constant around 0.12/min. The inhibition is relieved by addition of EGTA with a rate constant around 0.15/min. If Cd2+ is allowed to interact with calmodulin prior to the association of the protein with the ghosts, the inhibition is mainly competitive. The results suggest that the inhibitory effect caused by Cd2+ is due to an interaction with calmodulin. The slow interaction of Cd2+ suggests that calmodulin bound to the (Ca2+ + Mg2+)-ATPase is inaccessible to Cd2+.

Ca(2+) Mg(2+)-ATPase↗
PubMed1985

Rapid activation of ornithine decarboxylase by mitogenic (but not by nonmitogenic) ligands in human T lymphocytes.

The T cell mitogens, concanavalin A and the monoclonal antibody OKT3, cause a rapid activation of ornithine decarboxylase (ODC) activity in human T lymphocytes, maximal within 10 min of mitogen addition. Nonmitogenic ligands to T cell surface structures do not induce ODC. The enzyme induction is dependent upon an intact mobility of ligand-receptor complex, requires a functioning energy metabolism, but is independent of de novo protein synthesis. The early induction of pre-existing ODC molecules appears to be specifically linked to the initiation of T lymphocyte proliferation.

Cycloheximide↗
PubMed1985

Kinetics of long-term (72 hr) calcium content during mitogen activation of cultured human T lymphocytes.

In vitro stimulation of human blood lymphocytes with mitogen resulted in an increased intracellular content of Ca2+ per unit cell volume. This increase in Ca2+ content of lectin-activated cells reached a maximum after 24 hr of culture and thereafter slowly declined. Brief treatment of cells at 24 hr of culture with the Ca2+ ionophore A23187 in combination with EGTA resulted in a larger release of Ca2+ from cells in mitogen-stimulated cultures than from cells in control cultures. This indicates that the Ca2+ is accumulated intracellularly but is readily exchangeable. At 24 hr of culture the increase in cellular Ca2+ correlated well with the proliferative response as measured by 3H-thymidine incorporation. Ca2+ influx at 24 and 48 hr of culture was markedly enhanced in the mitogenically stimulated cells as compared either to cells cultured for 1 and 72 hr or cells cultured without mitogen.

Biological Transport↗
PubMed1985

Effects of Cd2+ upon Ca2+ fluxes and proliferation in concanavalin A-stimulated lymphocytes.

The mitogenic response of human peripheral blood lymphocytes to the lectin concanavalin A (conA) is inhibited by micromolar concentrations of CdCl2. This inhibition is partially relieved by an increase in the external Ca2+ concentration (from 0.6 to 2.2 mM). The initial rate of conA-induced 45Ca2+ influx is unaltered by CdCl2, although the level of 45Ca2+ accumulation increases. The basal rate of 45Ca2+ entry is not measurably disturbed by CdCl2 (100 microM). The steady-state efflux of 45Ca2+ and the calmodulin-activated (Ca2+ + Mg2+)-ATPase activity of erythrocyte ghosts are inhibited by CdCl2 (10 microM). Thus, the mechanism behind the Cd2+-induced suppression of the mitogenic response to conA is not due to alteration of mitogen-stimulated Ca2+ influx. We suggest that Cd2+ competes with Ca2+ for intracellular Ca2+-binding molecules, such as calmodulin, essential for the induction of cell proliferation.

Biological Transport, Active↗
PubMed1984

Depolarization of the mitochondrial membrane potential increases free cytosolic calcium in synaptosomes.

Intracellular calcium transients in synaptosomes, isolated from the guinea pig brain, were measured using entrapped metallochromic indicator arsenazo III. Addition of 1 microM carbonyl cyanide p-trifluoromethoxyphenyl hydrazone (FCCP) increased rapidly the absorbance of the entrapped arsenazo III, indicating an increase in the cytosolic free calcium. The FCCP-induced increase in cytoplasmic free Ca2+ was not blocked by 200 microM verapamil, while the increment in calcium caused by 40 microM veratridine was verapamil-sensitive. The absorbance changes induced by FCCP were not significantly increased when the extracellular potassium concentration was elevated from 5.4 to 50 mM. These data indicate that in nerve endings of mammalian brain, cytoplasmic free calcium, which is essential for the release of transmitter, is increased on depolarization of major intracellular calcium buffers, mitochondria.

Animals↗
PubMed1984

Direct mitogenic effect of ionophore A23187 on isolated human T helper lymphocytes.

The Ca2+ ionophore A23187 induces only a weak mitogenic response in cultures of unfractionated mononuclear leukocytes from human blood. When a comitogen, 12-O-tetradecanoyl-phorbol-13-acetate, which by itself is nonmitogenic, is added, a greatly increased cell proliferation is obtained. Highly purified T lymphocytes respond by proliferation to A23187 alone. Studies on functional subsets of T cells, fractionated by using the monoclonal antibodies OKT4/OKT8, revealed that A23187 is a strong mitogen for pure T helper cells (T4+). This suggests that increased cytosolic Ca2+ directly triggers a proliferative response in T helper cells with no apparent need for accessory cells.

Calcimycin↗
PubMed1984

Electron-dense endoplasmic reticulum-like profiles closely associated with mitochondria in glomus cells of the carotid body after fixation with oxalate.

Fixation in the presence of oxalate was used to demonstrate the electron-dense Ca2+ precipitates in the endoplasmic reticulum in glomus cells of the carotid body. Glomus cells in intact carotid bodies or cells dissociated from the organ by treatment with collagenase were studied electron microscopically. In the intact organ as well as in dissociated glomus cells, electron-dense endoplasmic reticulum-like profiles were seen closely associated with mitochondria, while these lacked reaction product. The interspace between mitochondria was occupied by electron-dense, slightly distended ER, which appeared to contact the outer membrane of the mitochondria. Occasionally, a mitochondrion was in contact with several ER profiles or the ER formed an electron-dense 'cap' on the mitochondrion. The electron-dense precipitates could be removed from ultrathin sections with the calcium chelator ethyleneglycol-2(2-aminoethyl tetra-acetic acid) (EGTA). It is tentatively suggested that the endoplasmic reticulum could be involved in intracellular buffering of Ca2+ in the glomus cell, as has been previously suggested for neurons.

Animals↗
PubMed1984

Membrane potential generation and calcium transport in plasma membrane enriched fractions from fetal human brain.

Plasma membrane fractions isolated from fetal human brain of 14-19 weeks of gestation are capable of generating a membrane potential of 30-50 mV as a response to a gradient of K+ ions. Valinomycin, a K+ conducting ionophore, does not affect the membrane potential whereas it is markedly reduced by veratridine which opens Na+ channels in excitable membranes. The membrane fractions concentrate Ca2+ by an ATP-dependent mechanism. The uptake has a high affinity for Ca2+, it is enhanced by oxalate and abolished by the 2H+/Ca2+ exchanger A 23187. Trifluoroperazine (40 microM), a calmodulin antagonist, inhibits Ca2+ uptake by 80%. Addition of Na+ causes efflux of part of the Ca2+ taken up in the presence of ATP, suggesting that a Na+-linked Ca2+ transport is also present in the membranes. The results show that the neuronal membranes of the fetal human brain already in the early second trimester of gestation have properties similar to those of the adult animal brain.

Acetylcholinesterase↗
PubMed1984

Facilitation of transmission at the crayfish neuromuscular synapse by a convulsant phenol.

The effects of the convulsant drug 4-Cl phenol on synaptic transmission were studied in the opener muscle of the crayfish walking leg. 4-Cl phenol was found to increase the amplitude of the excitatory postsynaptic potential without affecting the resting potential or input resistance of the muscle fiber. The drug did not change the frequency of spontaneous miniature postsynaptic potentials in K+-depolarized fibers. The postsynaptic voltage response to bath-applied glutamate (the excitatory transmitter compound) was decreased while the Cl(-) -conductance increase related to the action of bath-applied gamma-aminobutyric acid (the inhibitory transmitter) was not affected. In the light of previous results obtained on crayfish axons it is concluded that convulsant phenols induce an increase in the evoked release of transmitter by increasing the duration of the presynaptic depolarization through a block of voltage-dependent potassium channels.

Action Potentials↗
PubMed1984

Free extracellular [Ca2+] at photoreceptor level equals that in vitreous in frog and carp eyes.

The extracellular Ca2+ concentration has a strong influence on the function of retinal photoreceptors. In the present study both single- and double-barrelled Ca2+-sensitive microelectrodes were used to measure the free calcium ion concentration in the extracellular fluid bathing the photoreceptors in opened excised eyes of frogs and carps. The electrode tip was localized electrophysiologically by voltage drops related to current pulses passed radially through the preparation, using the high-resistance pigment epithelium as a landmark. The measurements showed that the extracellular Ca2+ concentration is very close or identical to that prevailing in the vitreous humour, 1.0-1.1 mM in the frog retina and 1.2-1.4 mM in the carp.

Animals↗
PubMed1983

Phosphate dependent, ruthenium red insensitive CA2+ uptake in mung bean mitochondria.

Energy linked Ca2+ uptake into mung bean mitochondria has been studied. Using arsenazo III as a monitor of extramitochondrial Ca2+, we observe a respiration-linked uptake of Ca2+ which requires phosphate and is insensitive to ruthenium red. The rate of uptake is of the order of 5 nmol/mg protein/min. Acetate, sulphate and thiosulphate are unable to support Ca2+ uptake. The results suggest that although plant mitochondria accumulate Ca2+ in an energy dependent fashion, it is not via a simple electrophoretic uniport mechanism.

Biological Transport, Active↗
PubMed1983

Qualitative measurements of cytosolic calcium ion concentration within isolated guinea pig nerve endings using entrapped arsenazo III.

If arsenazo III is present during homogenization of brain this metallochromic indicator is entrapped within subsequently isolated synaptosomes. A large proportion of the entrapped indicator is released upon addition of digitonin to disrupt the synaptosomal plasma membrane. A similar proportion of [3H]sucrose is also trapped within synaptosomes if present in the homogenization medium, suggesting that homogenization causes a transient opening of the nerve ending as it is chopped off from the axon. Addition of the ionophore A23187 or depolarization of the plasma membrane by adding veratridine, gramicidin or increasing external K+ changes the absorbance of the entrapped dye, with peaks of absorbance around 600 and 650 nm, typical of the arsenazo III-Ca2+ complex. The response to veratridine is inhibited by the Ca2+-channel antagonist, verapamil, while that of A23187 is unaffected. The present method provides a sensitive technique for measurements of changes in cytosolic calcium ion concentrations within nerve endings.

Animals↗
PubMed1983

Evidence for a Ca2+ gradient across the plasma membrane of wheat protoplasts.

Fluxes of Ca2+ across the plasma membrane of isolated wheat protoplasts have been measured both as net accumulation and as uptake under steady-state conditions. The ATPase inhibitors, orthovanadate and diethylstibesterol, and the divalent cation ionophore, A23187, were all found to enhance net Ca2+ accumulation by protoplasts. The uptake of Ca2+ under steady-state conditions was also stimulated by A23187 but relatively unaffected by a range of plant hormones or by red or far red light. Light treatments were compared to dark controls with protoplasts isolated from etiolated wheat. The results suggest that plant cells maintain a Ca2+ gradient across their plasma membrane but it appears not to be under phytochrome control.

Calcimycin↗
PubMed1983

Endocytosis of cationized ferritin to vesicles in the Golgi region of the glomus cells dissociated from the adult rat carotid body.

Glomus cells were dissociated from the carotid bodies of adult rats by enzymatic digestion with collagenase. The cells were then incubated at 37 degrees C for 30 minutes to 3 hours in the continuous presence of cationized ferritin (CF) as a membrane marker and extracellular tracer to study the intracellular route of endocytosis in this cell type. After 30 minutes of incubation with CF, occasional solitary CF-containing vesicles were observed at the cell periphery and also in the Golgi region. After 2-3 hours of incubation with CF, cell viability was still preserved and CF-labeled vesicles were abundant in the Golgi region. CF particles were also seen in some vesicles having a dense core. The core of these labeled vesicles appeared to be less electron-dense than that of typical secretory granules. It is suggested that the Golgi apparatus is involved in membrane recycling in glomus cells and that the membrane is then possibly further transported to an immature type of storage vesicle for reusage.

Animals↗
PubMed1982

Concanavalin A binding and Ca2+ fluxes in rat spleen cells.

Addition of the mitogenic lectin concanavalin A to rat spleen cells results in a small increase in the steady-state Ca2+ content of the cells. 45Ca2+ fluxes were measured under conditions where artifacts due to Ca2+ binding to concanavalin A could be excluded. Both 45Ca2+ influx into and efflux from these cells are significantly activated by the lectin. If 45Ca2+ is added 30 min after concanavalin A the rate of influx is further enhanced. The increase in 45Ca2+ influx correlates well with binding of concanavalin A to the cells. At low concentrations (optimal mitogenic) of the lectin (1 and 3 micrograms/ml) no significant increase in 45Ca2+ influx occurs but an increase in 45Ca2+ efflux is still observed. The results suggest that concanavalin A binding to the cell surface causes an increase in Ca2+ influx into the cells and that activation of Ca2+ efflux occurs as a response to an increase in the cytosolic Ca2+ activity. Thus, Ca2+ may well play a role in triggering lymphocyte activation.

Animals↗
PubMed1982

Endocytotic uptake of cationized ferritin tracer into glomus cells dissociated from the adult rat carotid body.

Glomus cells from carotid bodies of adult rats dissociated by means of collagenase or collagenase + trypsin were used to study by electron microscopy the endocytotic uptake of cationized ferritin (CF) tracer into subcellular compartments. The glomus cells were incubated with the tracer (1) in a basic salt medium (BM), or (2) in the BM into which calcium ionophore A23187 had been added, or (3) in a potassium-rich medium. Incubation of the cells in BM containing CF for 30 min resulted in attachment of the tracer to the cell membrane and uptake of a few solitary tracer particles into small vesicles and multivesicular bodies. No uptake into the cisternae of the Golgi apparatus was observed. Further incubation in BM containing CF for another 30 min resulted in increased uptake of the tracer into small vesicles and multivesicular bodies. A similar pattern of uptake was observed when the dissociated glomus cells were first preincubated in BM with CF for 30 min and then incubated for 1 min or 30 min in the BM solution containing both the ionophore and CF. Upon such incubation, CF particles were seen to penetrate into coated pits and sites of exocytosis at the cell surface. When the 30-min preincubation in BM was followed by incubation in a CF-containing potassium-rich medium for 15-30 min, uptake into vesicles, small lysosomes and occasionally also into profiles of the smooth endoplasmic reticulum was seen. Endocytotic mechanisms of the glomus cells are outlined.

Animals↗