Effect Of Potassium Depletion On Myocardial Potassium Content And Myocardial Na~+ -K~+ -ATPase In Rabbits

Objective:To investigate changes of plasma potassium(K) concentration and myocardial K content and response of myocardial Na⁺-K⁺-ATPase activity andα₁,α₂ isoform protein expression in K-depleted rabbits; to investigate effect of myocardial Na⁺-K⁺-ATPase on potassium distribution during potassium depletion and repletion. K-depleted rabbit models were prepared in this study with a low-K cotent diet.Methods:Sixty-four healthy male rabbits weighing 2.0².7kg were randomly divided into 8 groups (n=8/group):control group (group T₀) ,K-depleted 3 days (group T₁) , K-depleted 6 days (group T₂) , K-depleted 9 days (group T₃) , K-depleted 12 days (group T₄) , K-depleted 15 days (group T₅) ,repletion control group (group RC) and repletion experimental group (group RE).All rabbits were fed a normal diet for a week before model preparation.Rabbits in group T₀ and group RC were given a normal diet only(100g standard chow/d each rabbit,free access to normal water),rabbits in groupT₁,T₂,T₃,T₄,T₅ and RE were fed a low-K content diet(100g low-K content chow/d each rabbit,free access to distilled water) for 3days,6 days, 9 days, 12 days, 15 days and 15 days,respectively.After models were prepared,the hearts of group T₀,T₁,T₂,T₃,T₄,T₅ were excised after blood sampling, intravenous potassium repletion experiment was done in group RC and RE.Rabbits in group RC and RE were anesthetized .And then a femoral vein infusion catheter and a central auricular artery blood sampling catheter were inserted.Blood samples were taken from the central auricular artery intermittently during KCl infusion to monitor plasma K.When plasma K in the two groups reached ⁵.5mmol/L and maintained at this level for at least 1 hour,KCl infusions were stopped and the heart was immediately excised for K content measurement. Sampling blood to measure Plasma K,Na, hemoglobin concentration through central auricular artery and excising heart were performed on the day that models were prepared in group T₀,T₁,T₂,T₃,T₄,T₅.Rabbits of T₀,T₁,T₂,T₃,T₄,T₅ were anesthetized by 4% pentobarbital (1ml/kg) given through auricular vein,their thoracic cavity were opend and hearts were excised.Hearts of rabbits in group RC and RE were cut off after K repletion.About 35mg of anterior left ventricular wall from group T₀,T₁,T₂,T₃,T₄,T₅,RC and RE were washed with cold iso-osmia LiCl,dryed,weighed exactly and then preserved in -20℃for myocardial K and Na content measurement.Two samples of about 0.5g of anterior left ventricular wall from group T₀, T₃, T₅ were washed with physiology saline,marked,frozen in liquid nitrogen and preserved in -70℃for measuring activity of Na⁺-K⁺-ATPase and protein expression of Na⁺-K⁺-ATPaseα₁,α₂ isoform.One sample was preserved in group T₁,T₂,T₄ for measuring activity of Na⁺-K⁺-ATPase.Artial blood samples were analyzed for plasma K and Na concentrations by ion-sensitive electrodes, hemoglobin by spectrophotometric method.Myocardial K and Na contents were measured by flame atomic absorption photometry.Myocardial Na⁺-K⁺-ATPase activity was measured with an ultramicro-ATPase test box after total protein quantity with a coomssie brilliant blue protein measurement test box.Myocardial Na⁺-K⁺-ATPaseα₁ andα₂ isoform relative content were measured by western blot.Myocardial K uptake of group RC(or group RE) during KCl infusions were calculated as the difference between myocardial K content in each heart after KCl infusion and mean myocardial K content in group T₀(or group T₅);mean myocardial K uptake rate of each rabbit was calculated by respective myocardial K uptake divided by duration of K infusion .Results:1 Animals were clinically unaffected and the low-K content dietary regimens were well tolerated. 2 There was no significant difference of the rabbits'weight and ages in all the groups.3 The changes of plasma K,Na and hemoglobin concentrationCompared with group T₀,plasma K in group T₁,T₂,T₃,T₄,T₅ decreased gradually,the statistical differcence was significant(P <0.05); no significant difference in plasma Na and hemoglobin were observed in group T₀,T₁,T₂,T₃,T₄,T₅(P >0.05).4 The changes of myocardial K,Na contentCompared with group T₀, there were no significant decrease in myocardial K in group T₁ (P >0.05), myocardial K in group T₂ T₃,T₄,T₅ decreased,the statistical differcence was significant(P <0.05);there were no significant increse in myocardial Na in group T₁,T₂,T₃,T₄,T₅ compared with that in group T₀ (P >0.05).5 The changes of myocardial Na⁺-K⁺-ATPase activityCompared with group T₀, myocardial Na⁺-K⁺-ATPase activity in group T₁,T₂ increased,but the statistical differcence was not significant(P >0.05), myocardial Na⁺-K⁺-ATPase activity in group T₃ ,T₄,T₅ increased,the statistical differcence was significant(P <0.05)6 Relationship between myocardial K and Na⁺-K⁺-ATPase activityLinear regression analysis of data of myocardial K content and Na⁺-K⁺-ATPase activity in group T₀, T₁,T₂ ,T₃ ,T₄,T₅ showed a negative linear relationship between mean Na⁺-K⁺-ATPaseb activity and mean myocardial K content, linear regression equation was Y=20.16-0.11X,R2=0.894, P < 0.05.7 Protein expression of myocardial Na⁺-K⁺-ATPaseα₁ andα₂ isoformCompared with group T₀, myocardial Na⁺-K⁺-ATPaseα₁ isoform protein expression in group T₃,T₅ increased, the statistical differcence was significant(P <0.05), myocardial Na⁺-K⁺-ATPaseα₂ isoform protein expression in group T₃,T₅ decreased, the statistical differcence was significant(P <0.05).8 The comparison of myocardial K uptake and mean myocardial K uptake rate after potassium repletion Compared with group RC, myocardial K uptake in group RE increased, the statistical differcence was significant(P <0.05), mean myocardial K uptake rate in group RE increased, the statistical differcence was significant(P <0.05),infused KCl dosage in group RE was higher than that in group RC, the statistical differcence was significant(P<0.05).Conclusions:1 The present study prepared K-depleted rabbit models successfully used a low-K content diet,which induced a plasma K decrease in experimental rabbits.2 Myocardial K content gradually decreased during potassium depletion.3 Potassium depletion induced an upregulation in myocardial total Na⁺-K⁺-ATPaseb activity;linear regression analysis of data from K-depleted and contral rabbits showed a negative linear relationship between mean Na⁺-K⁺-ATPaseb activity and mean myocardial K content.4 The responsivity of myocardial Na⁺-K⁺-ATPaseα₁ andα₂ isoform to potassium is different,protein expression ofα₁ isoform was up-regulated andα₂ isoform down-regulated during K depletion.5 Potassium clearance capacity in cardial muscle of K-depleted rabbits increased signifiantly and tolerance to K content increase was upregulated.