High affinity and low affinity ouabain binding sites in the rat heart

Area of Science:

• Cardiology
• Molecular Biology
• Biochemistry

Background:

• Cardiac glycosides like ouabain exert positive inotropic effects via specific receptors.
• Rat ventricular muscle exhibits two classes of ouabain receptors.
• Low affinity receptors are linked to Na+, K+-ATPase, but high affinity sites require further characterization.

Purpose of the Study:

• To investigate whether high affinity ouabain receptors in rat ventricular muscle are sarcolemmal Na+, K+-ATPase.
• To characterize the biochemical properties of these high affinity binding sites.

Main Methods:

• Examined [3H]ouabain binding characteristics to high affinity sites.
• Assessed the influence of ATP, Mg2+, Na+, and K+ on binding.
• Tested inhibition by other cardiac glycosides (digoxin, digitoxin) and cassaine.
• Investigated the role of sympathetic nerve terminals using 6-hydroxydopamine.
• Utilized [gamma-32P]ATP labeling to quantify Na+, K+-ATPase.

Main Results:

• High affinity [3H]ouabain binding requires ATP, Mg2+, and Na+, and is inhibited by K+.
• Digoxin, digitoxin, and cassaine inhibited binding, with cassaine being less potent.
• Destruction of sympathetic nerve terminals did not alter high affinity binding sites.
• High affinity ouabain binding sites constitute 25% of total Na+, K+-ATPase in rat ventricular muscle.

Conclusions:

• High affinity ouabain binding sites in rat ventricular muscle exhibit properties consistent with sarcolemmal Na+, K+-ATPase.
• These sites are distinct from Na+, K+-ATPase in sympathetic nerve terminals.
• A significant portion (25%) of cardiac Na+, K+-ATPase in rat ventricular muscle binds ouabain with high affinity.