Interactions of dynorphin A and related peptides with cardiac ouabain binding sites.

The effect of dynorphin A (Dyn A) and related peptides on the binding of [3H]ouabain was examined in rat cardiac sarcolemma. Scatchard analysis of [3H]ouabain binding revealed the existence of two distinct sites: a high affinity (Kd: 20.4 nM) low capacity (Bmax: 1.55 pmol/mg protein) site and a low affinity (Kd: 3695 nM) high capacity (Bmax: 39.3 pmol/mg protein) site. Dyn A-(1-13) (10 microM) interacted selectively with the low affinity [3H]ouabain binding site causing a significant decrease in the Bmax (from 39.3 to 22.2 pmol/mg protein) without altering the Kd. Dyn A-(1-13) inhibited the binding of [3H]ouabain (500 nM) with an IC50 of 2.9 microM and a maximal inhibition of 77% of the specific binding activity. The non-opioid fragments Dyn A-(2-13), Dyn A-(3-13) and Dyn A-(6-10) displayed 36%, 32% and 14% of the potency of Dyn A-(1-13) respectively; whereas 100 microM of Dyn A-(1-8), Leu-enkephalin (Leu-Enk) and selective ligands for kappa (U-50, 488H) mu [(D-Ala2-Me-Phe4.Glyol5]Enk) and delta [(D-Ser2.Thr6]Leu-Enk) opioid receptors caused little or no inhibition of [3H]ouabain binding. The relative potency of various analogs and fragments of Dyn A in inhibiting the binding of [3H]ouabain correlated well (r = 0.89-0.90) with their potency in inhibiting the binding of [3H]Dyn A-(1-13) to non-opioid Dyn sites (Dumont and Lemaire, 1996) and to block [3H]NA uptake by cardiac synaptosomes (Dumont and Lemaire, 1995). In spontaneously hypertensive rats. [3H]ouabain binding displayed a 3.8-fold enhanced sensitivity to the action of Dyn A-(1-13) an effect that correlated with the enhanced ability of the peptide to inhibit Na+/K+-ATPase and [3H]NA uptake. The results indicate that Dyn A and related peptides may modulate cardiac functions through a non-opioid interaction with the low affinity ouabain binding site.