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The effects of ATP on the interactions between monovalent cations and the sodium pump in dialysed squid axons
Author(s) -
Beaugé Luis,
Di Polo Reinaldo
Publication year - 1981
Publication title -
the journal of physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.802
H-Index - 240
eISSN - 1469-7793
pISSN - 0022-3751
DOI - 10.1113/jphysiol.1981.sp013719
Subject(s) - efflux , sodium , chemistry , ouabain , atp hydrolysis , squid , biophysics , membrane potential , atpase , potassium , ion , ion transporter , membrane , biochemistry , enzyme , biology , ecology , organic chemistry
1. The efflux of Na in dialysed axons of the squid has been used to monitor the sidedness of the interactions of the Na pump with Na + ions, K + ions and ATP. The axons were under conditions such that most of the Na efflux went through the Na pump by means of a complete cycle of ATP hydrolysis. 2. With 310 m m ‐K i + , 70 m m ‐Na i + and 10 m m ‐K + artificial sea water (ASW) more than 97% of the Na efflux was abolished by removal of ATP. The efflux of Na was stimulated by ATP with a K ½ of about 200 μ m . This is similar to the K ½ of 150 μ m found for the ATP dependence of a ouabain‐sensitive Na,K‐ATPase activity in membrane fragments isolated from squid optical nerves. 3. A 100‐fold reduction in the ATP concentration (from 3‐5 m m to 30‐50 μ m ) increased the apparent affinity of the Na pump for K o + about 8‐fold. In addition, the maximal rate of K o + ‐stimulated Na efflux was reduced by a similar factor. Analogous results were seen in axons dialysed with 310 m m ‐K i + or without K i + . 4. The relative effectiveness of external monovalent cations as activators of the Na efflux was a function of the ATP concentration inside the axon. With 3‐5 m m ‐ATP the order of effectiveness was K + > NH 4 + > Rb + . With 30‐50 μ m ‐ATP the sequence was NH 4 + » K + » Rb + . These results were not affected by the removal of K i + . 5. When the ATP concentration was 3 m m and the Na i + concentration 70 m m , the removal of K i + produced a slight and reversible increase in the total efflux of Na (15%) and no change in the ATP‐dependent Na efflux. When the ATP concentration was reduced to 30‐50 μ m , or the Na i + concentration lowered to 5‐10 m m , the removal of K i + reversibly increased the total and the ATP‐dependent efflux of Na. The largest increase in Na efflux was seen when both ATP and Na i + were simultaneously reduced. The ATP‐dependent extra Na efflux resulting from the exclusion of K i + was abolished by 10 −4 m ‐ouabain in the sea waters. 6. The increase in the ATP‐dependent Na efflux observed in axons dialysed with 0 K i + + 10 m m ‐K + ASW was not seen in axons perfused with 310 m m ‐K i + + 450 m m ‐K + ASW. However, both experimental conditions gave rise to a similar (and small) ATP‐independent and ouabain‐insensitive efflux of Na. This indicates that the effects on the Na pump of removing K i + are not due to the simultaneous membrane depolarization. In addition, it suggests that K i + has an inhibitory effect on the Na pump, and that that effect is antagonized by Na i + and ATP. 7. The present results are consistent with the idea that the same conformation of the Na pump (and Na,K‐ATPase) can be reached by interaction with external K + after phosphorylation and with internal K + before rephosphorylation. This enzyme conformation produces an enzyme—K complex from which K + ions are not easily released unless high concentrations of ATP are present. This also stresses a non‐phosphorylating regulatory role of ATP.