N¹-Dansyl-Spermine and N¹-(n-Octanesulfonyl)-Spermine, Novel Glutamate Receptor Antagonists: Block and Permeation ofN-Methyl-d-Aspartate Receptors

Abstract

The effects of several N-sulfonyl-polyamines, including N¹-dansyl-spermine (N¹-DnsSpm) andN¹-(n-octanesulfonyl)-spermine (N¹-OsSpm), were studied at recombinantN-methyl-d-aspartate (NMDA) receptors expressed in Xenopus laevis oocytes. N¹-DnsSpm and N¹-OsSpm inhibited NMDA receptors and were ∼1000-fold more potent than spermine in oocytes voltage-clamped at −70 mV. Block by N¹-DnsSpm and N¹-OsSpm was strongly voltage dependent, being more pronounced at hyperpolarized membrane potentials. With the Woodhull model of voltage-dependent channel block, the values of K_d(0) were 779 μm, 882 μm, and 7.4 mm and those of zδ were 2.58, 2.57, and 1.07 for N¹-DnsSpm, N¹-OsSpm, and spermine, respectively. This suggests that an increase in the voltage dependence of block together with an increase in affinity contributes to the increased potencies of N¹-DnsSpm and N¹-OsSpm compared with spermine. Sensitivity to N¹-DnsSpm was reduced by mutation NR1(N616Q) and was increased by mutations NR1(N616G) and NR2A(N615G). The NR1(N616G) and NR2A(N615G) mutations decreased the K_d(0) value of N¹-DnsSpm without affecting zδ, whereas the NR1(N616Q) mutation reduced zδ. These mutations may alter the accessibility of part of the polyamine binding site within the channel pore or directly alter the properties of that site. Block by N¹-DnsSpm (0.3 μm) was almost complete at −100 mV, and there was no relief of block at extreme negative membrane potentials (−100 to −200 mV) at wild-type NR1/NR2A channels. In contrast, block by N¹-DnsSpm was partially relieved at extreme negative potentials at receptors containing NR1(N616G) or NR2A(N615G), suggesting that N¹-DnsSpm can permeate these mutant channels but not wild-type NR1/NR2A channels. This is hypothesized to be due to an increase in the pore size of channels containing NR1(N616G) or NR2A(N615G), which allows passage of the bulky head group of N¹-DnsSpm. In contrast to N¹-DnsSpm, N¹-OsSpm could easily permeate wild-type NR1/NR2A channels, presumably because the head group of N¹-OsSpm can pass through the narrowest part of the channel pore.N-Sulfonyl-polyamines such as N¹-DnsSpm and N¹-OsSpm represent a new class of polyamine antagonists with which to study glutamate receptor ion channels.