Measurements of (Na+,K+)ATPase after in vitro hypoxia and reoxygenation are affected by methods of membrane preparation.
(Na+,K+ )ATPase activity was evaluated in membranes from rat hippocampal slices after in vitro hypoxia and reoxygenation. Membranes were prepared with two different methods, one using an isotonic medium and another using a hypotonic one. The changes that were found after hypoxia went into opposite directions in the two cases. Membranes prepared in a hypotonic medium are probably more suitable for these measurements. Using these membranes, hypoxia results in a slight decrease of (Na+,K+)ATPase activity and in a further decrease after reoxygenation. We also found that expressing (Na+,K+)ATPase activity as a percent of total ATPase activity is appropriate for membranes prepared under hypotonic conditions and can unveil (by reducing variability between experiments) significant changes that may be masked in small samples like ours.
Balestrino
M
M
Dipartimento di Scienze Neurologiche dell'Universita' di Genova, Italy.
Cupello
A
A
Morelli
A
A
Musante
L
L
Panfoli
I
I
D000818:Animals; D006624:Hippocampus / Q000201:enzymology / Q000378:metabolism; D002534:Hypoxia, Brain / Q000201:enzymology*; D066298:In Vitro Techniques; D007552:Isotonic Solutions; D007700:Kinetics; D008297:Male; D008566:Membranes / Q000201:enzymology; D010766:Phosphorylation; D051381:Rats; D017207:Rats, Sprague-Dawley; D000254:Sodium-Potassium-Exchanging ATPase / Q000032:analysis / Q000378:metabolism*; D013997:Time Factors
D007552:Isotonic Solutions; D000254:Sodium-Potassium-Exchanging ATPase