Pharmacological characterization of [3H]-Ifenprodil binding to polyamine binding sites on rabbit and rat retinal homogenates: role in neuroprotection?
Polyamine binding sites (PBS) represent one of the modulatory sites on the N-methyl-D-aspartate (NMDA) receptor-channel complex. We have characterized [3H]-ifenprodil binding to the PBS on washed homogenates of rabbit and rat retinas. Specific binding of [3H]-ifenprodil (2 nM) (in the presence of 3 microM 1,3-Di [2-tolyl] guanidine HCl and 10 microM GBR12909 to block sigma sites) comprised 47-56% of the total binding. Scatchard analyses indicated interaction with apparent high- and low-affinity sites: dissociation constants (K(d)s) = 0.5-0.6 microM and apparent density of sites (Bmax) = 1.5-4.3 pmol/mg protein and K(d)s = 2.0-2.9 microM, and Bmax values = 15.8-17.8 pmol/mg protein (n = 3). Ifenprodil (Ki = 0.4-0.8 microM), eliprodil (Ki = 0.7-0.8 microM), spermine (Ki = 72-79 microM), spermidine (Ki = 283-330 microM), putrescine (Ki > 650 microM) and MK-801 (Ki > 1 mM) (n = 3-5) differentially competed for [3H]-ifenprodil binding. The biphasic competition curves for ifenprodil were resolved into two binding components: rat retinas, IC50high = 0.19 +/- 0.13 microM and IC50low = 8.7 +/- 1.3 microM; rabbit retinas, IC50high = 0.1 +/- 0.01 microM and IC50low = 16.0 +/- 7.8 microM. These studies have shown the presence of specific PBS labeled by [3H]-ifenprodil in the rabbit and rat retinas which may, in part, be responsible for mediating the neuroprotective effects of eliprodil and ifenprodil.
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Molecular Pharmacology Unit, Alcon Laboratories, Inc., Fort Worth, Texas 76134, USA.
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D000818:Animals; D001667:Binding, Competitive; D018691:Excitatory Amino Acid Antagonists / Q000494:pharmacology; D006801:Humans; D018696:Neuroprotective Agents / Q000494:pharmacology*; D010880:Piperidines / Q000378:metabolism / Q000494:pharmacology*; D011073:Polyamines / Q000378:metabolism*; D011817:Rabbits; D051381:Rats; D012160:Retina / Q000187:drug effects* / Q000378:metabolism; D014316:Tritium
D018691:Excitatory Amino Acid Antagonists; D018696:Neuroprotective Agents; D010880:Piperidines; D011073:Polyamines; D014316:Tritium; C010739:ifenprodil