title: Differential interactions of Na+ channel toxins with T-type Ca2+ channels.
abstract: Two types of voltage-dependent Ca(2+) channels have been identified in heart: high (I(CaL)) and low (I(CaT)) voltage-activated Ca(2+) channels. In guinea pig ventricular myocytes, low voltage-activated inward current consists of I(CaT) and a tetrodotoxin (TTX)-sensitive I(Ca) component (I(Ca(TTX))). In this study, we reexamined the nature of low-threshold I(Ca) in dog atrium, as well as whether it is affected by Na(+) channel toxins. Ca(2+) currents were recorded using the whole-cell patch clamp technique. In the absence of external Na(+), a transient inward current activated near -50 mV, peaked at -30 mV, and reversed around +40 mV (HP = -90 mV). It was unaffected by 30 microM TTX or micromolar concentrations of external Na(+), but was inhibited by 50 microM Ni(2+) (by approximately 90%) or 5 microM mibefradil (by approximately 50%), consistent with the reported properties of I(CaT). Addition of 30 microM TTX in the presence of Ni(2+) increased the current approximately fourfold (41% of control), and shifted the dose-response curve of Ni(2+) block to the right (IC(50) from 7.6 to 30 microM). Saxitoxin (STX) at 1 microM abolished the current left in 50 microM Ni(2+). In the absence of Ni(2+), STX potently blocked I(CaT) (EC(50) = 185 nM) and modestly reduced I(CaL) (EC(50) = 1.6 microM). While TTX produced no direct effect on I(CaT) elicited by expression of hCa(V)3.1 and hCa(V)3.2 in HEK-293 cells, it significantly attenuated the block of this current by Ni(2+) (IC(50) increased to 550 microM Ni(2+) for Ca(V)3.1 and 15 microM Ni(2+) for Ca(V)3.2); in contrast, 30 microM TTX directly inhibited hCa(V)3.3-induced I(CaT) and the addition of 750 microM Ni(2+) to the TTX-containing medium led to greater block of the current that was not significantly different than that produced by Ni(2+) alone. 1 microM STX directly inhibited Ca(V)3.1-, Ca(V)3.2-, and Ca(V)3.3-mediated I(CaT) but did not enhance the ability of Ni(2+) to block these currents. These findings provide important new implications for our understanding of structure-function relationships of I(CaT) in heart, and further extend the hypothesis of a parallel evolution of Na(+) and Ca(2+) channels from an ancestor with common structural motifs.
authors.affiliation: Excigen, Inc., Baltimore, MD 21224, USA.
authors.forename: Hui
authors.initials: H
authors.lastname: Sun
authors.forename: Diego
authors.initials: D
authors.lastname: Varela
authors.forename: Denis
authors.initials: D
authors.lastname: Chartier
authors.forename: Peter C
authors.initials: PC
authors.lastname: Ruben
authors.forename: Stanley
authors.initials: S
authors.lastname: Nattel
authors.forename: Gerald W
authors.initials: GW
authors.lastname: Zamponi
authors.forename: Normand
authors.initials: N
authors.lastname: Leblanc
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002121:Calcium Channel Blockers / Q000494:pharmacology; D020746:Calcium Channels, L-Type / Q000235:genetics / Q000502:physiology; D020747:Calcium Channels, T-Type / Q000235:genetics / Q000502:physiology*; D002460:Cell Line; D002478:Cells, Cultured; D004285:Dogs; D004347:Drug Interactions; D006801:Humans; D008012:Lidocaine / Q000494:pharmacology; D008387:Marine Toxins / Q000494:pharmacology*; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D020748:Mibefradil / Q000494:pharmacology; D008969:Molecular Sequence Data; D032383:Myocytes, Cardiac / Q000166:cytology / Q000187:drug effects / Q000502:physiology; D009532:Nickel / Q000494:pharmacology; D012530:Saxitoxin / Q000494:pharmacology; D017386:Sequence Homology, Amino Acid; D026941:Sodium Channel Blockers / Q000494:pharmacology*; D013779:Tetrodotoxin / Q000494:pharmacology; D014162:Transfection
chemical_list: C121855:CACNA1G protein, human; C479952:CACNA1H protein, human; C469757:CACNA1I protein, human; D002121:Calcium Channel Blockers; D020746:Calcium Channels, L-Type; D020747:Calcium Channels, T-Type; D008387:Marine Toxins; D026941:Sodium Channel Blockers; D020748:Mibefradil; D012530:Saxitoxin; D013779:Tetrodotoxin; D009532:Nickel; D008012:Lidocaine