title: Modification of cloned brain Na+ channels by batrachotoxin.
abstract: The effects of batrachotoxin (BTX) on cloned alpha-subunit Na+ channels were examined in CHO-K1 cells (a chinese hamster ovary cell line) transfected with rat brain NaIIA cDNA. Under whole-cell patch clamp conditions, BTX shifted the voltage dependence of the activation process by about 45 mV towards the hyperpolarizing direction and eliminated the inactivating phase of Na+ currents. Repetitive depolarizations greatly facilitated the binding of BTX with NaIIA channels while the membrane was held at -100 mV. In chloramine-T-pretreated cells, the association rate of BTX binding with the NaIIA channel was 6.5-fold faster than that in untreated cells. The estimated association rate constant for BTX binding with the open form of NaIIA channel was 1.11 x 10(6) mol-1.s-1 at room temperature. BTX-modified NaIIA channels were blocked by tetrodotoxin (TTX) in a complicated manner. First, the TTX binding to the closed state of BTX-modified NaIIA channels was not voltage dependent. The KD value of TTX was measured at 8.9 nM, which was similar to that of unmodified channels (KD = 14.2 nM). Second, the block of the open state of BTX-modified NaIIA channels by TTX was voltage dependent; depolarization reduced the potency of TTX block between -20 mV to +50 mV. Below -30 mV, the TTX affinity began to level off, probably because of the increased presence of the closed state. Unexpectedly, steady-state inactivation of BTX-modified NaIIA channels was minimal as measured by the two-pulse protocol, a phenomenon distinctly different from that found in GH3 cells.(ABSTRACT TRUNCATED AT 250 WORDS)
authors.affiliation: Department of Anesthesia Research Laboratories, Harvard Medical School, Boston, MA.
authors.forename: G K
authors.initials: GK
authors.lastname: Wang
authors.forename: S Y
authors.initials: SY
authors.lastname: Wang
mesh_terms: D000818:Animals; D001496:Batrachotoxins / Q000378:metabolism / Q000494:pharmacology*; D001566:Benzocaine / Q000494:pharmacology; D001921:Brain / Q000378:metabolism*; D016466:CHO Cells; D002460:Cell Line; D003001:Cloning, Molecular; D006224:Cricetinae; D003412:Cricetulus; D015640:Ion Channel Gating; D051381:Rats; D026941:Sodium Channel Blockers; D015222:Sodium Channels / Q000378:metabolism*; D013779:Tetrodotoxin / Q000378:metabolism
chemical_list: D001496:Batrachotoxins; D026941:Sodium Channel Blockers; D015222:Sodium Channels; D013779:Tetrodotoxin; D001566:Benzocaine