A century of exercise physiology: effects of muscle contraction and exercise on skeletal muscle Na+,K+-ATPase, Na+ and K+ ions, and on plasma K+ concentration-historical developments.

This historical review traces key discoveries regarding K+ and Na+ ions in skeletal muscle at rest and with exercise, including contents and concentrations, Na+,K+-ATPase (NKA) and exercise effects on plasma [K+] in humans. Following initial measures in 1896 of muscle contents in various species, including humans, electrical stimulation of animal muscle showed K+ loss and gains in Na+, Cl- and H20, then subsequently bidirectional muscle K+ and Na+ fluxes. After NKA discovery in 1957, methods were developed to quantify muscle NKA activity via rates of ATP hydrolysis, Na+/K+ radioisotope fluxes, [3H]-ouabain binding and phosphatase activity. Since then, it became clear that NKA plays a central role in Na+/K+ homeostasis and that NKA content and activity are regulated by muscle contractions and numerous hormones. During intense exercise in humans, muscle intracellular [K+] falls by 21 mM (range - 13 to - 39 mM), interstitial [K+] increases to 12-13 mM, and plasma [K+] rises to 6-8 mM, whilst post-exercise plasma [K+] falls rapidly, reflecting increased muscle NKA activity. Contractions were shown to increase NKA activity in proportion to activation frequency in animal intact muscle preparations. In human muscle, [3H]-ouabain-binding content fully quantifies NKA content, whilst the method mainly detects α2 isoforms in rats. Acute or chronic exercise affects human muscle K+, NKA content, activity, isoforms and phospholemman (FXYD1). Numerous hormones, pharmacological and dietary interventions, altered acid-base or redox states, exercise training and physical inactivity modulate plasma [K+] during exercise. Finally, historical research approaches largely excluded female participants and typically used very small sample sizes.

Na+,K+-ATPase with Disrupted Na+ Binding Sites I and III Binds Na+ with Increased Affinity at Site II and Undergoes Na+-Activated Phosphorylation with ATP.

Na+,K+-ATPase actively extrudes three cytoplasmic Na+ ions in exchange for two extracellular K+ ions for each ATP hydrolyzed. The atomic structure with bound Na+ identifies three Na+ sites, named I, II, and III. It has been proposed that site III is the first to be occupied and site II last, when Na+ binds from the cytoplasmic side. It is usually assumed that the occupation of all three Na+ sites is obligatory for the activation of phosphoryl transfer from ATP. To obtain more insight into the individual roles of the ion-binding sites, we have analyzed a series of seven mutants with substitution of the critical ion-binding residue Ser777, which is a shared ligand between Na+ sites I and III. Surprisingly, mutants with large and bulky substituents expected to prevent or profoundly disturb Na+ access to sites I and III retain the ability to form a phosphoenzyme from ATP, even with increased apparent Na+ affinity. This indicates that Na+ binding solely at site II is sufficient to promote phosphorylation. These mutations appear to lock the membrane sector into an E1-like configuration, allowing Na+ but not K+ to bind at site II, while the cytoplasmic sector undergoes conformational changes uncoupled from the membrane sector.

Screening of Peptides that Specifically Binds to M3-M4 Extracellular Domain of Sodium Pump α1 Subunit and Analysis of Their Bioactivity In Vitro and In Vivo.

Interaction between ouabain (OUA) and Na+/K+-pump remains in the current focus of hypertension research. This study aimed to find an oligopeptide that would antagonize the inhibitory effect of endogenous OUA on Na+/K+-pump and examine its activity at the cellular and organism levels. To this end, Phage Random 12 Peptide Library was employed to screen for specific polypeptide ligands that interact with M3-M4 extracellular domain of Na+/K+-pump α1 subunit known as OUA-binding site. Synthetic sequence ILEYTWLEAGGGS of extracellular domain M3-M4 of Na+/K+-pump α1 subunit was used as the target. The phage positive clones were screened and identified using the phage library and double sandwich ELISA. DNA was extracted and sequenced to synthesize 3 peptide ligands to Na+/K+-pump: P-A, P-B, and P-C. We also studied the effects of the short peptide with the highest potency for countering OUA on proliferation and apoptosis of EA.hy926 vascular endothelial cells and on systolic BP in spontaneously hypertensive rats (SHR). The effect of peptide P-A on proliferation (stimulation with physiological concentrations of OUA) and on apoptosis (stimulation with OUA in high concentrations) of EA.hy926 vascular endothelial cells was assessed by the MTT test and flow cytometry, respectively. In SHR rats, intravenous injection of P-A decreased systolic BP. Oligopeptide P-A competitively antagonized the inhibitory action of OUA on Na+/K+-pump, OUA-induced proliferation, and OUA-provoked apoptosis of cultured EA.hy926 cells. Our findings open vista for the emergence of novel hypertensive drugs.

Altered expression of sodium transporters in the excretory duct-ligated submandibular gland in rat

Na(+)-Li(+) Countertransport and Na(+)-K(+) Pump of Red Blood Cells in Patients with Essential Hypertension

BACKGROUND: This study was performed in order to investigate the changes of Na+ transport system in the red blood cells of patients with essential hypertension. METHODS: Na(+)-Li(+) countertransport and Na(+)-K(+) pump activity were measured in 30 cases of essential hypertension and 20 healthy normal controls. And these measurements were analyzed in terms of some important clinical parameters in the patients, i.e., body mass index(BMI), status of hypertension and plasma lipids. RESULTS: Na(+) and K(+) contents of red cells in hypertensive patients were 16.9+/-1.4 and 77.8+/-2.1mmol/L cells, respectively, and no significant difference was found compared with respective value of normal control(14.2+/-0.9 and 82.2+/-2.8mmol/L cells). Na(+)-Li(+) countertransport in the patients group was significantly ancreased compared with control group(62.5+/-4.5 and 46.8+/-3.0umol/L cells.hr), and Na(+)-K(+) pump activity was also showed a significant depression(8.72+/-0.80 and 12.79+/-0.52umol Pi/mg.hr). In the analysis regarding the relationship between Na(+) transport system and some important clinical parameters of the patients with essential hypertension, Na(+)-Li(+) countertransport was related to BMI, and the level of triglyceride. On the other hand, Na(+)-K(+) pump activity was related to the WHO stage and the levels of total cholesterol and triglyceride. But level of blood pressure did not show a significant correlation with either are of the two Na(+) transport system. CONCLUSION: These resluts suggest that Na(+)-Li(+) countertransport and Na(+)-K(+) pump activity in patients with essential hypertension were significantly altered compared with heathy normal controls, and these Na(+) transport system were also influenced by BMI, WHO stage, and the levels of cholesterol and triglyceride. And the individual variation in Na(+) transport system were also suggested by the findings being overlapped between hypertensive patients and controls.

Effects of Zhiyu Granules on sciatic nerve function of diabetic peripheral neuritis / 中成药

AIM: To discuss the mechanism of action of Zhiyu Granules on treating diabetic peripheral neuritis. METHODS: The model of diabetic peripheral neuritis in mice was induced by streptozotocin.The effect of Zhiyu Granules on the contents of sciatic nerve inositol in model mice through ELSD(evaporative light scattering detector) was examined,and the activity of Na~+-K~+ pump in sciatic nerve was measured,and the activity of SOD and the contents of MDA in erythrocytes were detected,and the ultrastructures of sciatic nerve,pancreas and kidney in model mice were observed by electric microscope. RESULTS: Zhiyu Granules could increase the contents of sciatic nerve inositol in model rats,and enhance the activity of Na~+-K~+ pump,and improve the ultrastructures of sciatic nerve and pancreas cell.Zhiyu Granules couldn't improve the ultrastructure of renal corpuscles,or affect the activity of SOD and the contents of MDA in erythrocytes of model mice yet. CONCLUSION: Zhiyu Granules is effective in clinical treatment of diabetic peripheral neuritis.