RESUMEN
The activities of Na-K-ATPase and Na-K-2Cl cotransporter (NKCC1) were studied in the aorta, heart, and skeletal muscle of streptozotocin (STZ)-induced diabetic rats and control rats. In the aortic rings of STZ rats, the Na-K-ATPase-dependent (86)Rb/K uptake was reduced to 60.0 +/- 5.5% of the control value (P < 0.01). However, Na-K-ATPase activity in soleus skeletal muscle fibers of STZ rats and paired control rats was similar, showing that the reduction of Na-K-ATPase activity in aortas of STZ rats is tissue specific. To functionally distinguish the contributions of ouabain-resistant (alpha(1)) and ouabain-sensitive (alpha(2) and alpha(3)) isoforms to the Na-K-ATPase activity in aortic rings, we used either a high (10(-3) M) or a low (10(-5) M) ouabain concentration during (86)Rb/K uptake. We found that the reduction in total Na-K-ATPase activity resulted from a dramatic decrement in ouabain-sensitive mediated (86)Rb/K uptake (26.0 +/- 3.9% of control, P < 0.01). Western blot analysis of membrane fractions from aortas of STZ rats demonstrated a significant reduction in protein levels of alpha(1)- and alpha(2)-catalytic isoforms (alpha(1) = 71.3 +/- 9.8% of control values, P < 0.05; alpha(2) = 44.5 +/- 11.3% of control, P < 0.01). In contrast, aortic rings from the STZ rats demonstrated an increase in NKCC1 activity (172.5 +/- 9.5%, P < 0.01); however, in heart tissue no difference in NKCC1 activity was seen between control and diabetic animals. Transport studies of endothelium-denuded or intact aortic rings demonstrated that the endothelium stimulates both Na-K-ATPase and Na-K-2Cl dependent (86)Rb/K uptake. The endothelium-dependent stimulation of Na-K-ATPase and Na-K-2Cl was not hampered by diabetes. We conclude that abnormal vascular vessel tone and function, reported in STZ-induced diabetic rats, may be related to ion transport abnormalities caused by changes in Na-K-ATPase and Na-K-2Cl activities.
Asunto(s)
Aorta/enzimología , Proteínas Portadoras/metabolismo , Diabetes Mellitus Experimental/metabolismo , Angiopatías Diabéticas/metabolismo , Isoenzimas/metabolismo , ATPasa Intercambiadora de Sodio-Potasio/metabolismo , Animales , Transporte Biológico/efectos de los fármacos , Transporte Biológico/fisiología , Western Blotting , Bumetanida/farmacología , Diuréticos/farmacología , Endotelio Vascular/enzimología , Inhibidores Enzimáticos/farmacología , Hipertensión/metabolismo , Técnicas In Vitro , Isoenzimas/análisis , Isoenzimas/antagonistas & inhibidores , Masculino , Tono Muscular/fisiología , Músculo Liso Vascular/fisiología , Ouabaína/farmacología , Potasio/farmacocinética , Ratas , Ratas Sprague-Dawley , Radioisótopos de Rubidio , Simportadores de Cloruro de Sodio-Potasio , ATPasa Intercambiadora de Sodio-Potasio/análisis , ATPasa Intercambiadora de Sodio-Potasio/antagonistas & inhibidoresRESUMEN
Chronic renal failure in the rat is associated with an impaired extrarenal potassium handling, whereas a renal adaptive mechanism of the remaining nephrons has been described. To understand the molecular basis of potassium homeostasis during renal failure we investigated the in vitro pump activity and the catalytic mRNA transcription in three different tissues: skeletal muscle, isolated adipocytes and kidney. The activity of the sodium pump, as measured by ouabain-sensitive 86Rb/K uptake in isolated adipocytes and skeletal muscle fibers, revealed a significant reduction of the pump activity in uremic rats. The reduction of the Na, K-ATPase activity in adipose tissue was associated with a similar decrement of both catalytic subunits (alpha 1 and alpha 2), whereas in the skeletal muscle tissue was only related to a decrease in the activity of the alpha 1 isoform. The expression of rat Na, K-ATPase catalytic isoforms mRNAs in kidney, muscle and adipose tissue from control and chronic renal failure rats was investigated at the molecular level with cDNA probes specific for the catalytic isoforms (alpha 1 and alpha 2). Northern blot analysis revealed that the respective catalytic mRNAs of uremic rats are regulated in a tissue-specific manner that are in agreement with the sodium-potassium pump activity. Muscle and adipose tissue showed a decrement in the levels of expression for the alpha 1 isoform mRNA. In contrast to these tissues, an increment in alpha 1 mRNA expression was observed in the kidney of rats with chronic renal failure.(ABSTRACT TRUNCATED AT 250 WORDS)
Asunto(s)
Tejido Adiposo/enzimología , Regulación Enzimológica de la Expresión Génica , Riñón/enzimología , Músculos/enzimología , ATPasa Intercambiadora de Sodio-Potasio/metabolismo , Uremia/enzimología , Animales , Northern Blotting , Isoenzimas/metabolismo , Fallo Renal Crónico/enzimología , Masculino , Ouabaína/farmacología , Potasio/metabolismo , ARN Mensajero/metabolismo , Ratas , Ratas Sprague-Dawley , Radioisótopos de Rubidio/metabolismo , ATPasa Intercambiadora de Sodio-Potasio/genéticaRESUMEN
Previous studies have suggested that an alteration in the expression of the Na,K-ATPase of muscle may be an important determinant of enhanced insulin sensitivity in chronic renal failure. Therefore, in the present studies we have examined the effect of uremia on the Na,K-ATPase alpha isoforms in skeletal muscle, at the level of mRNA expression and enzymatic activity. The activity of the sodium pump, as measured ouabain-sensitive 86Rb/K uptake in soleus muscle, revealed a reduction in the activity in uremia, related to the increment in plasma creatinine values. The decrement in 86Rb uptake by the rat soleus muscle of experimental animals was associated with changes on Na,K-ATPase gene product. Northern analysis of mRNA revealed isoform-specific regulation of Na,K-ATPase by uremia in skeletal muscle: a decrease of approximately 50% in alpha 1 subunit Na,K-ATPase mRNA, as compared to controls. The decrement in alpha 1 mRNA correlates with the decreased activity of the Na,K-ATPase in uremia, under basal conditions and with the almost complete inhibition of the Na,K-ATPase, of uremic tissue by a concentration of 10(-5) M ouabain. Although the activity of the alpha 2 isoform pump was not modified by uremia, the 3.4-kb message for this enzyme was increased 2.2-fold; this discrepancy is discussed. Altogether these findings demonstrate that the defective extrarenal potassium handling in uremia is at least dependent in the expression of alpha 1 subunit of the Na,K-ATPase.
Asunto(s)
Isoenzimas/genética , Fallo Renal Crónico/enzimología , Músculos/enzimología , ARN Mensajero/metabolismo , ATPasa Intercambiadora de Sodio-Potasio/genética , Transcripción Genética , Animales , Masculino , Ouabaína/farmacología , Ratas , Ratas EndogámicasRESUMEN
Although bovine and ovine descending colon absorbed solute and sodium at approximately the same rate in vitro, water absorption by bovine colon was faster (15.0 +/- 2.0 microliters.h-1.cm-2, n = 8) than by ovine colon (8.6 +/- 1.3 microliters.h-1.cm-2, n = 9; P less than 0.01). Consequently, the observed osmolality of cattle absorbate was lower (364 +/- 13 mosmol/kg, n = 8) than with sheep (807 +/- 135 mosmol/kg, n = 9; P less than 0.01). Paracellular permselectivity was examined to elucidate this difference; the permeability of bovine descending colon to [3H]polyethylene glycol 400 and 4000 was higher than in sheep (P less than 0.001). A paracellular solvent drag was observed in bovine but not in ovine colon, and the electrical resistance of bovine colon was lower (16.0 +/- 1.4 omega.cm2) than ovine colon (28.0 +/- 2.4 omega.cm2; P less than 0.001). Pore radii of 2.5 nm for ovine and 5 nm for bovine colonic paracellular route were estimated from these data. It is concluded that an increased hydraulic conductance of the "active" route for solute absorption combined with raised solute reflux via the wider paracellular pathway may account for the failure of cattle to form a hypertonic absorbate and, consequently, hard feces.
Asunto(s)
Colon/fisiología , Absorción Intestinal , Mucosa Intestinal/fisiología , Animales , Transporte Biológico , Bovinos , Colon/citología , Colon/efectos de los fármacos , Conductividad Eléctrica , Epitelio/fisiología , Técnicas In Vitro , Absorción Intestinal/efectos de los fármacos , Mucosa Intestinal/citología , Cinética , Matemática , Modelos Biológicos , Músculo Liso/citología , Músculo Liso/efectos de los fármacos , Músculo Liso/fisiología , Ouabaína/farmacología , Perfusión/métodos , Permeabilidad , Potasio/metabolismo , Rubidio/metabolismo , Ovinos , Teofilina/farmacología , AguaRESUMEN
Translocation of potassium to the intracellular compartment is impaired in advanced chronic renal failure. The purpose of this study was to evaluate the role of endogenous insulin in the disposal of an oral potassium load in uremia. Experiments were done on male Sprague-Dawley rats. Chronic renal failure (CRF) was induced by 3/4 nephrectomy. The results show that the addition of oral glucose to a potassium load was more effective in the translocation of potassium to the intracellular compartment in uremic animals. Further, suppression of endogenous insulin secretion with somatostatin caused a much higher increase in plasma potassium (K) of uremic rats (1.09 +/- 0.15 mEq/liter in CRF vs. 0.28 +/- 0.03 mEq/liter in control). Experiments to assess the activity of the Na pump were done in soleus muscles derived from these animals. Although a 50% reduction of the basal Na pump activity was found in the uremic muscles, the addition of insulin 100 mU/ml caused a relatively greater stimulation of ouabain-sensitive 86Rb uptake in the uremic muscle as compared to the control tissue (203% vs. 77% increment). These data suggest a greater sensitivity to insulin action on extrarenal potassium disposal in uremia.