Anthraquinone Polyamines: Novel Channel Blockers to Study N-Methyl-d-Aspartate Receptors

  1. Keiko Kashiwagi,
  2. Ikuko Tanaka,
  3. Maki Tamura,
  4. Hiromi Sugiyama,
  5. Tadashi Okawara,
  6. Masami Otsuka,
  7. Thomas N. Sabado,
  8. Keith Williams and
  9. Kazuei Igarashi
  1. Graduate School of Pharmaceutical Sciences, Chiba University, Chiba, Japan (K.K., I.T., M.T., H.S., K.I.); Faculty of Medical Pharmaceutical Sciences, Kumamoto University, Kumamoto, Japan (T.O., M.O.); and Department of Physiology and Pharmacology, State University of New York Downstate Medical Center, Brooklyn, New York (T.N.S., K.W.)
  1. Address correspondence to:
    Dr. Kazuei Igarashi, Graduate School of Pharmaceutical Sciences, Chiba University, 1-33 Yayoi-cho, Inage-ku, Chiba 263-8522, Japan. E-mail: iga16077{at}p.chiba-u.ac.jp

Abstract

The effects of various anthraquinone polyamines (AQP) were studied at recombinant N-methyl-d-aspartate (NMDA) receptors expressed in Xenopus laevis oocytes. The AQP derivatives had different numbers of methylene groups between the NH2 (or NH) groups in their spermidine-like tail. Thus, we termed these derivatives AQ33, AQ34, etc. All AQP derivatives inhibited responses of NR1/NR2 receptors in oocytes voltageclamped at -70 mV, with IC50 values between 4 and 22 μM. The block was strongly voltage-dependent. AQ34 and AQ33b inhibited responses of NR1/NR2 receptors but did not inhibit responses of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors expressed from GluR1 or GluR2(Q), indicating that AQ34 and AQ33b are preferential NMDA antagonists. Results of experiments using mutant NR1 and NR2 subunits identified residues that influence block by AQ34 and AQ33b. These residues are located in the outer vestibule at the selectivity filter/narrowest constriction of the channel and in the inner vestibule below the level of the selectivity filter. The results with mutant NR1 and NR2 subunits are consistent with the idea that NR1(Asn616) and NR2B(Asn616), but not NR2B(Asn615), make the narrowest constriction of NMDA channel.

Footnotes

  • This work was supported by a grant-in-aid for Scientific Research from the Ministry of Education, Culture, Sports, Science and Technology, Japan, by the Futaba Electronics Memorial Foundation, Chiba, Japan, and by the U.S. Public Health Service Grant NS35047.

  • DOI: 10.1124/jpet.103.062042.

  • ABBREVIATIONS: NMDA, N-methyl-d-aspartate; AMPA, α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid; AQP, anthraquinone polyamine; AQCC, anthraquinone-2-carboxyl chloride; MK-801, dizocilpine maleate; TB34, N1,N4,N8-tribenzylspermidine; AQ22, 2-{[2-[(anthraquinone-2-carbonyl)amino]ethyl]amino}ethylamine; AQ32, 3-{[3-[(anthraquinone-2-carbonyl)amino]propyl]amino}ethylamine; AQ23, 3-{[3-[(anthraquinone-2-carbonyl)amino]propyl]amino}ethylamine; AQ24, 4-{[2-[(anthraquinone-2-carbonyl)amino]ethyl]amino}butylamine; AQ33b, 4-(anthraquinone-2-carbonyl)-4-azaheptane-1,7-diamine; AQ33, 3-{[3-[(anthraquinone-2-carbonyl)amino]propyl]amino}propylamine; AQ34, 4-{[3-[(anthraquinone-2-carbonyl)amino]propyl]amino}butylamine.

    • Received October 24, 2003.
    • Accepted February 4, 2004.
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  1. Published online before print February 5, 2004, doi: 10.1124/jpet.103.062042 JPET June 2004 vol. 309 no. 3 884-893
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