Characterization of the stimulatory and inhibitory effects of polyamines on [³H]N-(1-[thienyl]cyclohexyl) piperidine binding to the N-methyl-D-aspartate receptor ionophore complex

Spermidine and spermine, as well as several other structurally related compounds, were tested in a [³H]N-(1-[thienyl]cyclohexyl) eridine ([³H]TCP) binding assay to determine the structural requirements of polyamines for activation of the N-methyl-D-aspartate-operated ion channel. Under nonequilibrium conditions, the polyamines enhanced [³H]TCP binding approximately 9-fold, with EC₅₀ values ranging from 0.8 to 60 μM. The order of potency in enhancing [³H]TCP binding was N,N'-bis(3-aminopropyl)-1,3-propanediamine > N,N'-bis-(3-aminopropyl)-ethylenediamine > spermine spermidine > N,N'-bis-(2-aminoethyl)-1,3-propanediamine. 1,3-Diaminopropane produced a partial agonistic effect, whereas putrescine, cadaverine, and 1,7-diaminoheptane were without effect at concentrations up to 1 mM. Eadie-Hofstee analysis of spermidine-induced [³H]TCP binding at equilibrium revealed a 3-fold increase in the affinity without a significant change in receptor density. This was further supported by kinetic data that showed that spermidine produced an increase in the association rate and a decrease in the dissociation rate of [³H]TCP binding to its site. Putrescine, cadaverine, and 1,3-diaminopropane antagonized the effects of spermidine by an apparently noncompetitive mechanism. Magnesium ions mimicked the effects of putrescine, suggesting the possibility that the inhibitory effects of Mg²⁺ and putrescine are mechanistically related.