Vasopressin, insulin and peroxide(s) of vanadate (pervanadate) influence Na+ transport mediated by (Na+, K+)ATPase or Na+/H+ exchanger of rat liver plasma membrane vesicles

Open Access
Publisher Website
Google Scholar
Abstract
Uptake of 22Na+ by liver plasma membrane vesicles, reflecting Na+ transport by (Na+, K+)ATPase or Na+/ H+ exchange was studied. Membrane vesicles were isolated from rat liver homogenates or from freshly prepared rat hepatocytes incubated in the presence of [Arg8]vasopressin or pervanadate and insulin. The ATP dependence of (Na+, K+)ATPase‐mediated transport was determined from initial velocities of vanadate‐sensitive uptake of 22Na+, the Na+‐dependence of Na+/H+ exchange from initial velocities of amiloride‐sensitive uptake.By studying vanadate‐sensitive Na+ transport, high‐affinity binding sites for ATP with an apparent Km(ATP) of 15 ± 1 μM were observed at low concentrations of Na+ (1 mM) and K+ (1 mM). At 90 mM Na+ and 60 mM K+ the apparent Km(ATP) was 103 ± 25 μM. Vesiculation of membranes and loading of the vesicles prepared from liver homogenates in the presence of vasopressin increased the maximal velocities of vanadate‐sensitive transport by 3.8‐fold and 1.9‐fold in the presence of low and high concentrations of Na+ and K+, respectively. The apparent Km(ATP) was shifted to 62 ± 7 μM and 76 ± 10 μM by vasopressin at low and high ion concentrations, respectively, indicating that the hormone reduced the influence of Na+ and K+ on ATP binding. In vesicles isolated from hepatocytes preincubated with 10 nM vasopression the hormone effect was conserved. Initial velocities of Na+ uptake (at high ion concentrations and 1 mM ATP) were increased 1.6 – 1.7‐fold above control, after incubation of the cells with vasopressin or by affinity labelling of the cells with a photoreactive analogue of the hormone.The velocity of amiloride‐sensitive Na+ transport was enhanced by incubating hepatocytes in the presence of 10 nM insulin (1.6‐fold) or 0.3 mM pervanadate generated by mixing vanadate plus H2O2 (13‐fold). The apparent Km(Na+) of Na+/H+ exchange was increased by pervanadate from 5.9 mM to 17.2mM. Vesiculation and incubation of isolated membranes in the presence of pervanadate had no effect on the velocity of amiloride‐sensitive Na+ transport.The results show that hormone receptor‐mediated effects on (Na+,K+)ATPase and Na+/H+ exchange are conserved during the isolation of liver plasma membrane vesicles. Stable modifications of the transport systems or their membrane environment rather than ionic or metabolic responses requiring cell integrity appear to be involved in this regulation.