Potentiation of NMDA receptor-mediated responses by dynorphin at low extracellular glycine concentrations

The effect of dynorphin A(1-13) on N-methyl-D-aspartate (NMDA)-activated currents was investigated in the presence of low extracellular glycine concentrations in Xenopus oocytes expressing recombinant heteromeric NMDA receptors and in cultured hippocampal neurons with the use of voltage-clamp techniques. At an extracellular added glycine concentration of 100 nM, dynorphin A(1-13) (10 mu M) greatly increased the amplitude of NMDA-activated currents for all heteromeric subunit combinations tested; on average, the potentiation was: epsilon 1/zeta 1, 3,377 +/- 1,416% (mean +/- SE); epsilon 2/zeta 1, 1,897 +/- 893%; epsilon 3/zeta 1, 4,356 +/- 846%; and epsilon 4/zeta 1, 1,783 +/- 503%. Potentiation of NMDA-activated current by dynorphin A(1-13) was concentration dependent between 0.1 and 10 mu M dynorphin A(1-13), with a half-maximal concentration value of 2.77 mu M and an apparent Hill coefficient of 2.53, for epsilon 2/zeta 1 subunits at 100 nM added extracellular glycine. Percentage potentiation by dynorphin A(1-13) was maximal at the lowest glycine concentrations tested (0.01 and 0.1 mu M), and decreased with increasing glycine concentration. No significant potentiation was observed at glycine concentrations >0.1 mu M for epsilon 1/zeta 1, epsilon 2/zeta 1, and epsilon 4/zeta 1 subunits, or at >1 mu M for epsilon 3/zeta 1 subunits. Potentiation of NMDA-activated currents by dynorphin A(1-13) was not inhibited by 1 mu M of the kappa-opioid receptor antagonist nor-binaltorphimine and potentiation was not observed with 10 mu M of the kappa-opioid receptor agonist trans-3,4-dichloro-N-methyl-N-[2-(1-pyrrolidinyl)-cyclohexyl]benzene-acetamide. Potentiation of NMDA-activated current by dynorphin A(1-13) was inhibited by the glycine antagonist kynurenic acid (50 mu M). NMDA-activated current was also potentiated at low glycine concentrations by 10 mu M dynorphin A(2-13) or (3-13), both of which have a glycine as the first amino acid, but not by 10 mu M dynorphin A(4-13), which does not have glycine as an amino acid. In hippocampal neurons, 10 mu M dynorphin A(1-13) or (2-13) potentiated steady-state NMDA-activated current in the absence of added extracellular glycine. The extracellular free glycine concentration, determined by high-performance liquid chromatography, was between 26 and 36 nM for the bathing solution in presence or absence of 10 mu M dynorphin A(1-13), (2-13), (3-13), or (4-13), and did not differ significantly among these solutions. The observations are consistent with the potentiation of NMDA-activated current at low extracellular glycine concentrations resulting from an interaction of the glycine amino acids in dynorphin A(1-13) with the glycine coagonist site on the NMDA receptor. Because dynorphin A is an endogenous peptide that can be coreleased with glutamate at glutamatergic synapses, the potentiation of NMDA receptor-mediated responses could be an important physiological regulator of NMDA receptor function at these synapses.