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Erythrocyte membrane ATPase and calcium pumping activities in porcine malignant hyperthermia
H S Thatte1, J R Mickelson, P B Addis
1Department of Medicine, Stanford University School of Medicine, California 94305.
Abstract:
To investigate possible abnormalities in erythrocyte membrane enzyme activities in the pharmacogenetic disorder MH, membrane ATPase activities have been examined in erythrocyte ghosts prepared from red blood cells of MHS and normal swine. While no differences were noted in Mg2+-ATPase activities, the (Na+, K+)-ATPase activity of MHS erythrocyte ghosts was less than that of normal ghosts. Ca2+-ATPase activity exhibited low- and high-affinity Ca2+-binding sites in both types of erythrocyte ghost. While the Km for Ca2+ was greater for normal than for MHS erythrocyte ghosts at the high-affinity Ca2+-binding site, the reverse was true at the low-affinity Ca2+-binding site. Irrespective of the type of calcium binding site occupied, the Vmax for normal erythrocyte ghost Ca2+-ATPase activity was greater than that for MHS ghosts. In the presence of calmodulin, there was now no difference between MHS and normal erythrocyte ghosts in either the Km for Ca2+ or the Vmax of the Ca2+-ATPase activity. To determine if the calcium pumping activity of intact MHS and normal pig erythrocytes differed, calcium efflux from the 45Ca-loaded erythrocytes was determined; this activity was significantly greater for MHS than for normal erythrocytes. Thus, the present study confirms that there are abnormalities in the membranes of MHS pig red blood cells. However, we conclude that these abnormalities are unlikely to result in an impaired ability of MHS erythrocytes to regulate their cytosolic Ca2+ concentration.
Insights
Malignant hyperthermia (MH) swine red blood cells show altered (Na+, K+)-ATPase and Ca2+-ATPase activities. Despite membrane abnormalities, MH erythrocytes likely maintain normal cytosolic calcium regulation.
Area of Science:
- Biochemistry
- Pharmacogenetics
- Cell Biology
Background:
- Malignant hyperthermia (MH) is a pharmacogenetic disorder affecting muscle contraction.
- Erythrocyte membrane enzyme abnormalities are investigated as potential indicators in MH.
- Swine models (MHS) are used to study MH-related physiological changes.
Purpose of the Study:
- To investigate erythrocyte membrane ATPase enzyme activities in MH susceptible (MHS) swine.
- To compare (Na+, K+)-ATPase and Ca2+-ATPase activities between MHS and normal swine erythrocytes.
- To assess the impact of these enzyme differences on cytosolic calcium regulation in MHS erythrocytes.
Main Methods:
- Erythrocyte ghosts were prepared from MHS and normal swine red blood cells.
- Activities of Mg2+-ATPase, (Na+, K+)-ATPase, and Ca2+-ATPase were measured.
- Calcium efflux from intact erythrocytes was determined using 45Ca loading.
Main Results:
- No difference in Mg2+-ATPase activity was observed.
- (Na+, K+)-ATPase activity was lower in MHS erythrocyte ghosts.
- Ca2+-ATPase kinetics (Km, Vmax) differed between MHS and normal ghosts, particularly at high-affinity sites; differences normalized with calmodulin.
- Calcium efflux was significantly higher in MHS erythrocytes compared to normal erythrocytes.
Conclusions:
- Confirmed abnormalities exist in MHS pig erythrocyte membranes, specifically in (Na+, K+)-ATPase and Ca2+-ATPase activities.
- These membrane alterations do not appear to impair the ability of MHS erythrocytes to regulate cytosolic calcium concentration.
- Calmodulin plays a role in normalizing Ca2+-ATPase function in MHS erythrocytes.