Biomedical subjects
H J Schatzmann
Publications and source records attributed to H J Schatzmann.
Haemolytic anaemia in analpha-lipoproteinaemia (Tangier disease): morphological, biochemical, and biophysical properties of the red blood cell.
A patient with familial analpha-lipoproteinaemia (Tangier disease) was found to have stomatocytosis and haemolytic anaemia. The analysis of the red cell membrane constituents revealed a low cholesterol content (90 nmol/ml red cells, control 130 nmol/ml red cells), a decreased cholesterol/phospholipid ratio (0.54, control 0.78), high phosphatidylcholine (41.5%, control 30.6%) and low sphingomyelin (18.8%, control 27.6%). The electrophoretic membrane protein pattern was normal. Osmotic gradient ektacytometry and osmotic resistance showed a decreased surface/volume ratio, which caused an increased filtration resistance in 3 microns pores. The elasticity of the membrane was unchanged. Functional membrane properties were altered: the anion exchange rate was increased, whereas alkali cation fluxes were normal. The capacity to release vesicles was reduced. This case represented a new type of stomatocytosis. It contributes to the understanding of the role of cholesterol and phospholipids in the red cell membrane and biomembranes in general.
ATP requirement of the sodium-dependent magnesium extrusion from human red blood cells.
1. Competitive behaviour detectable in the stimulatory action of external sodium (Nao+) and internal magnesium (Mgi2+) corroborates the idea that Nao+-dependent Mg2+ extrusion is a Mgi2+-Nao+ exchange. 2. Mg2+-loaded resealed cells made from metabolically starved cells (with less than 5 mumols/l cells of ATP), show hardly any Nao+-dependent Mg2+ outflow. Incorporation of ATP during lysis-resealing restores this Mg2+ transport. Half-saturation for the effect is reached at an initial ATP concentration of about 150 mumols/l cells. 3. Adenylyl(beta, gamma-methylene) diphosphonate (AMP-PCP) and AMP had no restituting effect, indicating that in order to act ATP must be hydrolysed. 4. Nao+-dependent Mg2+ outflow is not inhibited by vanadate concentrations that completely block the Ca2+ or Na+ pump. Therefore, the Nao+-Mgi2+ exchange does not fall into the class of cation pumps of the E1E2 type. 5. Yet the fact that reversal of the Na+ gradient fails to reverse the direction of the Na+-dependent Mg2+ transport in human red cells (Lüdi & Schatzmann, 1987) and that at equal Na+ concentration inside and outside the rate of Mg2+ transport is still 50% of that at a Na+ concentration difference of approximately 100 mM across the membrane suggests that the Na+ gradient, or the cation gradients in general, are not the only driving forces for Mg2+ movement. The assumption that there is energy input from ATP hydrolysis is compatible with these observations, whereas proposing the action of a protein kinase fails to explain them. 6. It is concluded that the Nao+-Mgi2+ exchange system has an absolute requirement for ATP and that it is more probable that ATP is supplying energy for transport rather than activating transport by protein phosphorylation or simply by binding.
The calcium pump of the surface membrane and of the sarcoplasmic reticulum.
The molecular architectures of SMP and SRP differ, yet the functional analogies between CAM-associated SMP and SRP are numerous. Most of the divergences observed may have their root in different behavior during dephosphorylation (see Table 1).
Dependence of the red blood cell calcium pump on the membrane potential.
(1) It is shown that the rate of calcium extrusion from intact human red cells is faster at a membrane potential of approximately +50 mV (inside) than at approximately -50 mV. (2) The positive potential applied was the chloride potential of KCl cells in a K-gluconate medium when the Ca2+ sensitive K+ channel was blocked by 0.3mM quinidine. The negative potential resulted from the high K+ permeability in Ca2+ loaded cells (the cells were loaded to a Ca2+ activity in the cell water of about 50 microM). (3) It is further demonstrated that the Ca2+ affinity of the pump ATPase is decreased both at the internal (high affinity) and external (low affinity) site by increasing the proton concentration. Acidification thus inhibits internally and stimulates externally. (4) An indirect effect of the membrane potential on the pump activity via the accompanying pH shifts on either side of the membrane could be ruled out by choosing Ca2+ concentrations which are fully activating at the internal Ca2+ binding site at pH 6.5 and not yet inhibitory at the external Ca2+ binding site at pH 8. (5) The result is compatible with the assumption that the human red cell Ca-pump is exchanging Ca2+ for protons, yet is electrogenic by virtue of a stoichiometry of 1H+:1Ca2+ for this exchange.
Some properties of a system for sodium-dependent outward movement of magnesium from metabolizing human red blood cells.
1. In agreement with the report by Féray & Garay (1986) it is shown that Mg2+ leaves human red cells mainly by a saturable pathway at a maximal rate of some 200 mumol 1(-1) cells h(-1) only if the medium contains Na+.Mg2+0.5 (i.e. the free Mg2+ concentration for half-maximal rate) at the internal membrane surface is 1.3 mM and the dissociation constant for Na+, KNa, at the external surface is 16-17 mM. Mg2+ shows co-operative behaviour. The Na+o-stimulated Mg2+ outflow is sensitive to millimolar amiloride concentrations. Implication of the Na+ or Ca+ pump can be ruled out. 2. Na+i is inhibitory by simple competition with Na+o. The affinity for Na+ inside is the same as outside. There is no detectable competition between Na+i and Mg2+i. 3. At approximately 1 mM-[Mg2+]i the outward Mg2+ movement stimulated by Na+o still proceeds when [Mg2+]o is increased up to 20 mM. Thus the Mg2+ movement is uphill and the apparent Mg2+0.5 at the external surface is larger than 20 mM. 4. Reversing the Na+ gradient (making [Na+]i greater than [Na+]o) does not elicit an inward Mg2+ movement, even if [Mg2+]o is simultaneously made larger than [Mg2+]i. 5. The Na+o-dependent Mg2+ outflow ceases nearly completely (falling to 5% of the control) in metabolically depleted cells. 6. The behaviour observed is compatible with the assumptions that (1) the system possesses distinct binding sites for Na+ and for Mg2+, (2) the ionophoric moiety is passively mobile when loaded with Na+, (3) the movement of the Na+ form is rate limiting, and (4) the Mg2+ form preferentially moves in the outward direction owing to an input of metabolic energy (ATP hydrolysis) and is immobile in starved cells. 7. Mg2+ may be required at a further site(s) not involved in the actual Mg2+ translocation but in the energy input. The simple kinetics suggesting translocation of one Na+ ion in exchange for one Mg2+ ion were found in selected cells of average maximum transport rate (Vmax) and may not hold for all cell specimens. 8. The conclusion is that the system is a Mg2+ extrusion pump driven by metabolic energy directly and not by the inward Na+ gradient, although net inward Na+ movement is necessary to bring the ionophoric part of the system back to the in-position. It appears that in intact cells the system operates far below saturation by Mg2+i and, by compensating for an inward leak of less than 5 mumol,1(-1) cells h(-1), sets the internal free Mg2+ concentration at about 0.5 of the equilibrium value.
Red blood cell calcium pump and its inhibition by vanadate and lanthanum.
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Hereditary leaky red cell syndrome in a Swiss family.
A Swiss family was found to have a hereditary hemolytic disorder associated with excessively leaky red cell membranes. Hemolysis was mild and fully compensated. Membrane lipid analysis revealed an increased phosphatidylcholine:phosphatidylethanolamine ratio. Membrane leaks included an increased permeability to sodium, potassium, calcium and, possibly, creatine. It is suggested that in hemolytic states, the combined finding of reticulocytosis and normal red cell creatine might be an easily obtainable clue to the presence of a leaky red cell syndrome.
Calcium transport by red blood cell membranes from young and adult cattle.
It is shown using inside-out membrane vesicles that cattle red cells extrude calcium by means of a calmodulin sensitive Ca-pump whose activity is high in calves and extremely low in adult cows. The decline is not due to loss of calmodulin susceptibility nor to a drop in Ca-affinity.
Some effects of removal of external calcium on pig striated muscle.
Bundles of about 800 cells from the m. thyreopharyngicus of pigs were used to measure activation and inactivation during contracture by K+ depolarization. When [Ca2+] in the medium was lowered to less than 5 X 10(-10) M for 3 min (replacing Ca2+ by Mg2+) the activation occurred at the same [K+] in the medium as in normal solution (3 mM-Ca2+) but inactivation was shifted to lower external [K+]. The absolute value of this shift in terms of membrane potential is uncertain, because [K+] at the cell surface is unknown. Exposure for 4 min to Ca2+-free medium (Ca2+ being replaced by Mg2+) had no effect on contractility tested after a subsequent rest of 22-25 min in normal solution ( [Ca2+] = 2 mM). However, if the muscle underwent one maximal K+ contracture in Ca2+-free medium the response (tetanus or K+ contracture) after the same interval in normal solution was strongly reduced, although the membrane potential recovered fully. K+ contractures in normal solution could be repeated without loss of contractile force. A K+ contracture in Ca2+-free medium had very little effect on the response to caffeine, tested after 25 min in normal solution. It seems that Ca2+ is lost into Ca2+-free medium only during depolarization, from a site which is not accessible to Ca2+ from outside at the resting membrane potential, or from inside at any membrane potential. This site might be located inside the transverse-tubular membrane and, when loaded with Ca2+, might represent the positive group of the model of Chandler, Rakowski & Schneider ( 1976b ), the movement of which during depolarization activates and inactivates the Ca2+ release from the sarcoplasmic reticulum.
The response to potassium of the Na-K pump ATPase in low-K red blood cells from cattle at birth and in later life.
It is shown that in low-K red blood cells of cattle the apparent affinity for K(1/Kapp K) at an inhibitory site of the Na-K ATPase increases markedly during the first 3 months of life. This site probably is the Na accepting site at the internal membrane surface and the change in Kapp K reflects an increase in KNa/KK, the ratio of the true dissociation constants. This effect may explain the concomitant fall in cellular K concentration.
The site of action of La3+ in the reaction cycle of the human red cell membrane Ca2+-pump ATPase.
Lanthanum (La3+) inhibits the Ca-pump of the red cell by arresting the protein in a phosphorylated form (PI). Similar La3+ concentrations are required to increase the amount of PI and to stop PI-decay. In the presence of La3+ phosphorylation becomes insensitive to Mg2+. PI made in the presence of Mg2+ is not prevented from decaying by subsequent addition of La3+, whereas that made in the absence of Mg2+ is. Taken together, these findings seem to indicate that La3+ blocks the transition between a 1st and a 2nd form of PI.
Trypsin activation of the red cell Ca2+-pump ATPase is calcium-sensitive.
Stimulation of the calmodulin-independent activity of the red cell Ca2+-pump ATPase by trypsin treatment (of calmodulin free red cell membranes) is sensitive to Ca2+ in a concentration range near the KCa of the transport site. The Ca2+ requirement for this effect is absolute, whereas the calmodulin sensitivity of the ATPase can be abolished by sufficient trypsin attack in the absence of Ca2+, although Ca2+ accelerates inactivation. This indicates that the two effects of trypsin are due to at least two distinct cleavage sites in the pump protein.
Is the red cell calcium pump electrogenic?
1. In inside-out vesicles of high potassium permeability, prepared from human red cell membranes, volume changes accompanying the action of the Ca2+ pump were measured by recording the intensity of light scattered by a suspension of these vesicles. Replacing Cl- by the impermeant gluconate anion changed swelling into shrinking. 2. Assuming that in Cl- media two Cl- ions accompany one Ca+ ion moved by the pump and in gluconate media two K+ ions are exchanged for one Ca2+ ion resulted in a good agreement between relative Ca2+ transport rate obtained from the volume change and from direct measurement of 45Ca uptake in the two media. 3. The fact that it is possible to change co-transport of Ca with Cl- into counter-transport of Ca2+ for K+ rules out that within the pump there is an obligatory coupling of Ca2+ movement with movement of another ion species (including the proton). The conclusion, therefore, is that the Ca2+ pump must be electrogenic. 4. The combination of measurement of volume change with direct measurement of 45Ca movement yielded 5-6 microliter/mg protein for the volume of the vesicles.
'Lactate type' response of ruminal fermentation to chloral hydrate, chloroform and trichloroethanol.
In ruminal fluid, taken from a cow 2 h after cereal feeding and supplemented with powdered grass, chloral hydrate diminished the rate of acetate, butyrate and propionate production. Instead, a considerable increase in lactate production was elicited by the drug. Chloroform and trichloroethanol had the same effect. Chloral hydrate and chloroform rendered the redox potential more negative which indicates inhibition of methane production. A concentration of chloral hydrate (2 mM), giving complete methane inhibition, reduced volatile fatty acid production by 30%. The findings are explained by assuming that chloral hydrate blocks methane production but in addition is inhibitory at a step below lactate in the reductive pathway to propionate and that the conditions chosen resulted in substrate saturation of one of the enzymes in the succinate pathway to propionate.
[Importance of pharmacokinetics for dosage in chemotherapy].
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Net ATP synthesis by running the red cell calcium pump backwards.
Ca2+ loaded inside-out vesicles from human red blood cells, yielding C2+ into a Ca2+ free medium with 4 mM EGTA, 2 mM ADP and 10 mM phosphate, produced an excess of 14.9 pmoles . min-1 . (mg protein)-1 of ATP compared to controls in which the transmembrane Ca2+ gradient was abolished by the ionophore A 23 187.