The mechanism of action for the block of NMDA receptor channels by the opioid peptide dynorphin

Dynorphin is one of the endogenous opioids that modulates the excitability of nociceptive (pain-sensing) neurons. We have shown recently that dynorphin blocks NMDA-activated currents directly without the participation of κ- opioid receptors. In order to understand the mechanism underlying this novel action of dynorphin, we examined, in detail, the interactions between dynorphin and NMDA receptors in isolated trigeminal neurons. Dynorphin reversibly blocks NMDA-activated current (I(NMDA)). The onset and recovery of the block were determined with concentration jump experiments. The association rate (k) of dynorphin(1-17) is 4.9 x 10⁶ sec^-1 M^-1 and the dissociation rate (k) is 7.5 sec^-1. The apparent dissociation constant (K(D)) of dynorphin, calculated from these rate constants, is 1.6 μM. Dynorphin does not change the EC₅₀ of NMDA, nor the potentiating action of glycine. The binding site for dynorphin is distinct from that of Zn²⁺ or H⁺. Upon treatment with the disulfide reducing agent dithiothreitol (DTT), NMDA receptors become less susceptible to dynorphin block. The affinity of dynorphin for the modified NMDA receptors is reduced by 2.7-fold. In analyses of single NMDA channels in cell-free patches, we found that dynorphin shortens the mean open time, decreases the probability of opening of NMDA channels, but has no effect on the single channel conductance. These results suggest that dynorphin interacts with a site conformationally linked with the redox site(s) on the NMDA receptor, thus altering the gating properties of the channel.