Jim S. Larsen

Diabetes Decreases Na + -K + Pump Concentration in Skeletal Muscles, Heart Ventricular Muscle, and Peripheral Nerves of Rat

Na+-K+-ATPase or the Na+-K+ pump is essential for some specific properties of muscle and nerve tissue such as contractility and excitability. Previous studies have shown conflicting variations in Na+-K+-ATPase activity or Na+-K+ pump concentration of muscle cells in experimental diabetes. Our study demonstrates that early untreated diabetes in rats induced by injection of streptozocin is associated with decreases in [3H]ouabain binding-site concentration of 24-48% in various skeletal muscles and 16% in peripheral nerves as well as a decrease in K+-dependent 3-O-methylfluorescein phosphatase activity of 21% in the heart ventricle. These effects could be prevented by insulin treatment. They probably represent a decrease in the concentration of Na+-K+ pumps. There was no evidence for more than one population of Na+-K+ pumps in intact samples of skeletal muscle and nerves from normal, diabetic, and insulin-treated animals. The decrease in Na+-K+ pump concentration in nerve cells may be due to atrophy of the axons. In skeletal muscles, myocardium, and peripheral nerves, the observed decrease in Na+-K+ pump concentration may be important for the pathophysiology of diabetes. We emphasize that quantification of Na+-K+-ATPase or the Na+-K+ pump in muscle and nerve tissue from diabetic animals should preferably be performed with either intact samples or crude homogenates of whole tissue.

Heart Na, K-ATPase activity in cardiomyopathic hamsters as estimated from K-dependent 3-O-MFPase activity in crude homogenates

The hamster hereditary cardiomyopathy provides a unique model for the study of membrane abnormalities during chronic congestive heart failure. It is associated with intracellular calcium accumulation, mitochondrial calcification and cell necrosis. Previous studies have shown a decrease in Na,K-ATPase activity purified from ventricle sarcolemma. The present study demonstrates a decrease in K-dependent 3-O-methylfluorescein phosphatase (3-O-MFPase) activity from 1.93 to 1.30 mumol/g wet wt. or 33% in crude homogenates from the left ventricle of 7-months-old cardiomyopathic hamsters as compared to control animals. This represents an equivalent decrease in Na, K-ATPase activity. The values are several times higher than previously published for membrane fractions of myocardium from the hamster. Concomitantly, there was an increase in intracellular Na-concentration of the myocardium of 42% whereas the K-concentration was unchanged. The decrease in Na,K-pump concentration may be of importance for the increase in intracellular sodium and ensuing calcifying necrosis observed in the myocardium of cardiomyopathic hamsters. It is emphasized that quantification of the Na,K-ATPase or Na,K-pump should preferably be performed using crude homogenates.

Effects of exercise on urinary excretion of albumin and β2-microglobulin in young patients with mild essential hypertension without treatment and during long-term propranolol treatment

The effect of exercise on the urinary excretion of albumin, beta 2-microglobulin, renal plasma flow (RPF) and glomerular filtration rate (GFR) was studied in two groups of young patients with mild essential hypertension, one comprising nine untreated patients (1) and one comprising eight patients treated with propranolol (2), and in a group comprising ten normotensive healthy control subjects (3). The urinary excretion of albumin and beta 2-microglobulin, RPF and GFR were measured during four consecutive periods: a control period of 40 min; two exercise periods of 20 min each, with a load of 75 W and 100 W, respectively; and a control period. During exercise group 1 showed an increase in albumin excretion and a decrease in beta 2-microglobulin excretion, while group 2 showed an increase in albumin excretion during heavy exercise only and unchanged beta 2-microglobulin excretion, and group 3 showed unchanged albumin and beta 2-microglobulin excretion. RPF and GFR were reduced during exercise in all groups, most markedly in the hypertensive groups. During both exercise loads albumin excretion was higher and RPF was lower in group 1 than in 3. In group 2 albumin excretion was lower than in 1 during the light exercise load, whereas RPF and GFR were lower during both exercise loads. It is concluded that young patients with mild essential hypertension have both an abnormally high albumin excretion and an abnormally low RPF during exercise. During propranolol therapy urinary albumin excretion was partly normal. The changes in albumin excretion suggest glomerular leakage.

Estimation of stability of [3H]-ouabain binding site concentration in rat and human skeletal muscle post mortem

The post mortem stability of the [3H]-ouabain binding site concentration and 3-O-methylfluorescein phosphatase (MFPase) activity was evaluated in rat skeletal muscle. As compared with the values measured in fresh tissue, the [3H]-ouabain binding site concentration in rat soleus muscle only dropped by around 1% per hour post mortem and a significant decrease was only seen after 12 h (15%, p less than 0.02). The 3-O-MFPase activity in rat gastrocnemius muscle showed a similar decrease. After 4 days, both parameters had dropped by 65% (p less than 0.001). In contrast, when intact fresh rat soleus muscles were incubated in Krebs-Ringer bicarbonate buffer at 20 degrees C for 4 days no significant decrease was seen in the [3H]-ouabain binding site concentration. In 10 human subjects the concentration of [3H]-ouabain binding sites was measured in specimens of the vastus lateralis muscle obtained within half an hour and at 6 and 12 h post mortem. The relative decrease after 6 h was insignificant (8%, p less than 0.30), whereas it was significant after 12 h (29%, p less than 0.005). These results have shown that the [3H]-ouabain binding sites in human skeletal muscle are resistant to post mortem degradation during the first 6 h after death. This makes it possible to perform measurements post mortem of the [3H]-ouabain binding site concentration in human skeletal muscle.

Reduction of cerebral cortical [3H]ouabain binding site (Na+,K+-ATPase) density in dementia as evaluated in fresh human cerebral cortical biopsies

Na+,K(+)-ATPase density in human cerebral cortex was for the first time studied by vanadate facilitated [3H]ouabain binding to intact samples. Fresh human cerebral cortical biopsies were obtained as a result of diagnostic frontal lobe biopsy from patients with normal pressure hydrocephalus (NPH) syndrome and associated dementia. For control measurements post-mortem samples were obtained from patients without clinically observed dementia. [3H]ouabain binding kinetics were evaluated: when incubating samples in 1 microM [3H]ouabain binding equilibrium was obtained after 6 h of incubation, non-specific uptake and retention amounted to only 2.3% of total uptake and retention of [3H]ouabain and release of specifically bound [3H]ouabain during washout in the cold occurred only slowly (T1/2 = 37 h). Evaluation of receptor affinity for ouabain was in agreement with a heterogeneous population of [3H]ouabain binding sites. [3H]Ouabain binding was significantly reduced after frozen storage of samples before measurements. Post-mortem degradation of cerebral [3H]ouabain binding sites occurred only slowly (T1/2 = 75 h). No significant variation in [3H]ouabain binding site density was observed between the cerebral lobes with occipital, parietal and temporal values (means +/- S.E.M., n = 5) amounting to 10281 +/- 649, 11267 +/- 1011 and 9263 +/- 615 pmol/g wet wt., respectively. [3H]Ouabain binding measured in frontal cortical samples gave values of (means +/- S.E.M., n = 5) 4274 +/- 1020 and 11397 +/- 976 pmol/g wet wt. delta % = 62; P < 0.05) in patients with dementia and controls, respectively. Human cerebral cortical capacity for active K+ uptake was around 37- and 16-fold greater than in skeletal muscular and myocardial tissue, respectively.

Myocardial Na,K-ATPase Concentration and Heart Failure

The membrane-bound Na, K-ATPase, which by deriving energy from ATP performs the active transport of Na and K across the cell membrane, is of major importance to excitable tissues. By adjusting the translocation of Na and K across the cell membrane, it plays an important role in generating and maintaining the membrane potential. Furthermore the Na,K-pump is of major importance for interstitial K homeostasis following depolarization [1–3]. Additionally, there is a general consensus that the inotropic effect of cardiac glycosides is achieved through binding to left ventricular Na,K-ATPase. Thus, occupancy of Na,K-ATPase with cardiac glycoside inhibits its active cation transport and therefore, indirectly increases intracellular Ca availability by the Na-Ca exchanger, which in turn activates contractile proteins [4].