Molecular pharmacology of the amiloride analog 3-amino-6-chloro-5-[(4- chloro-benzyl)amino]-N-[[(2,4-dimethylbenzyl)-amino]iminomethyl] -pyrazinecarboxamide (CB-DMB) as a pan inhibitor of the Na+-Ca 2+ exchanger isoforms NCX1, NCX2, and NCX3 in stably transfected cells

Agnese Secondo, Anna Pannaccione, Pasquale Molinaro, Paolo Ambrosino, Pellegrino Lippiello, Alba Esposito, Maria Cantile, Priti R. Khatri, Daniela Melisi, Gianfranco Di Renzo, Lucio Annunziato

Research output: Contribution to journalArticle

17 Citations (Scopus)

Abstract

With the help of single-cell microflorimetry, 45Ca2+ radiotracer fluxes, and patch-clamp in whole-cell configuration, we examined the effect of the amiloride derivative 3-amino-6-chloro-5-[(4-chloro-benzyl)amino]- N-[[(2,4-dimethylbenzyl)amino]-iminomethyl]-pyrazinecarboxamide (CB-DMB) on the activity of the three isoforms of the Na+/Ca2+ exchanger (NCX) and on several other membrane currents including voltage- and pHsensitive ones. This amiloride analog suppressed the bidirectional activity of all NCX isoforms in a concentration-dependent manner. The IC50 values of CB-DMB were in the nanomolar range for the outward and the inward components of the bidirectional NCX1, NCX2, and NCX3 activity. Deletion mutagenesis showed that CB-DMB inhibited NCX activity mainly at level of the f-loop but not through the interaction with Gly833 located at the level of the α2 repeat. On the other hand, CB-DMB suppressed in the micromolar range the other plasma membrane currents encoded by voltage-dependent Ca2+ channels, tetrodotoxin-sensitive Na+ channels, and pH-sensitive ASIC1a. Collectively, the data of the present study showed that CBDMB, when used in the nanomolar range, is one of the most potent compounds that can block the activity of the three NCX isoforms when they work both in the forward and in the reverse modes of operation without interfering with other ionic channels.

Original languageEnglish
Pages (from-to)212-221
Number of pages10
JournalJournal of Pharmacology and Experimental Therapeutics
Volume331
Issue number1
DOIs
Publication statusPublished - Oct 2009

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Sodium-Calcium Exchanger
Pyrazinamide
Amiloride
Protein Isoforms
Pharmacology
Tetrodotoxin
Ion Channels
Mutagenesis
Inhibitory Concentration 50
Cell Membrane
Membranes

ASJC Scopus subject areas

  • Pharmacology
  • Molecular Medicine

Cite this

Molecular pharmacology of the amiloride analog 3-amino-6-chloro-5-[(4- chloro-benzyl)amino]-N-[[(2,4-dimethylbenzyl)-amino]iminomethyl] -pyrazinecarboxamide (CB-DMB) as a pan inhibitor of the Na+-Ca 2+ exchanger isoforms NCX1, NCX2, and NCX3 in stably transfected cells. / Secondo, Agnese; Pannaccione, Anna; Molinaro, Pasquale; Ambrosino, Paolo; Lippiello, Pellegrino; Esposito, Alba; Cantile, Maria; Khatri, Priti R.; Melisi, Daniela; Di Renzo, Gianfranco; Annunziato, Lucio.

In: Journal of Pharmacology and Experimental Therapeutics, Vol. 331, No. 1, 10.2009, p. 212-221.

Research output: Contribution to journalArticle

Secondo, Agnese ; Pannaccione, Anna ; Molinaro, Pasquale ; Ambrosino, Paolo ; Lippiello, Pellegrino ; Esposito, Alba ; Cantile, Maria ; Khatri, Priti R. ; Melisi, Daniela ; Di Renzo, Gianfranco ; Annunziato, Lucio. / Molecular pharmacology of the amiloride analog 3-amino-6-chloro-5-[(4- chloro-benzyl)amino]-N-[[(2,4-dimethylbenzyl)-amino]iminomethyl] -pyrazinecarboxamide (CB-DMB) as a pan inhibitor of the Na+-Ca 2+ exchanger isoforms NCX1, NCX2, and NCX3 in stably transfected cells. In: Journal of Pharmacology and Experimental Therapeutics. 2009 ; Vol. 331, No. 1. pp. 212-221.
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T1 - Molecular pharmacology of the amiloride analog 3-amino-6-chloro-5-[(4- chloro-benzyl)amino]-N-[[(2,4-dimethylbenzyl)-amino]iminomethyl] -pyrazinecarboxamide (CB-DMB) as a pan inhibitor of the Na+-Ca 2+ exchanger isoforms NCX1, NCX2, and NCX3 in stably transfected cells

AU - Secondo, Agnese

AU - Pannaccione, Anna

AU - Molinaro, Pasquale

AU - Ambrosino, Paolo

AU - Lippiello, Pellegrino

AU - Esposito, Alba

AU - Cantile, Maria

AU - Khatri, Priti R.

AU - Melisi, Daniela

AU - Di Renzo, Gianfranco

AU - Annunziato, Lucio

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N2 - With the help of single-cell microflorimetry, 45Ca2+ radiotracer fluxes, and patch-clamp in whole-cell configuration, we examined the effect of the amiloride derivative 3-amino-6-chloro-5-[(4-chloro-benzyl)amino]- N-[[(2,4-dimethylbenzyl)amino]-iminomethyl]-pyrazinecarboxamide (CB-DMB) on the activity of the three isoforms of the Na+/Ca2+ exchanger (NCX) and on several other membrane currents including voltage- and pHsensitive ones. This amiloride analog suppressed the bidirectional activity of all NCX isoforms in a concentration-dependent manner. The IC50 values of CB-DMB were in the nanomolar range for the outward and the inward components of the bidirectional NCX1, NCX2, and NCX3 activity. Deletion mutagenesis showed that CB-DMB inhibited NCX activity mainly at level of the f-loop but not through the interaction with Gly833 located at the level of the α2 repeat. On the other hand, CB-DMB suppressed in the micromolar range the other plasma membrane currents encoded by voltage-dependent Ca2+ channels, tetrodotoxin-sensitive Na+ channels, and pH-sensitive ASIC1a. Collectively, the data of the present study showed that CBDMB, when used in the nanomolar range, is one of the most potent compounds that can block the activity of the three NCX isoforms when they work both in the forward and in the reverse modes of operation without interfering with other ionic channels.

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