TY - JOUR
T1 - Open-channel block by the cytoplasmic tail of sodium channel β4 as a mechanism for resurgent sodium current
AU - Grieco-Calub, Tina M
AU - Malhotra, Jyoti D.
AU - Chen, Chunling
AU - Isom, Lori L.
AU - Raman, Indira M.
N1 - Funding Information:
Consistent with the role of hydrophobicity in determining the affinity of exogenous blockers, our data suggest that the uncharged sequence, LITFIL, of the wild-type β4 peptide stabilized the interaction between the blocking peptide and the pore. Upon repolarization, the electrostatic repulsion favored by the flanking lysines appeared to be counteracted by the affinity of the hydrophobic sequence for the channel, delaying unbinding. This interpretation is supported by the results with the scrambled peptide. When the hydrophobic sequence was interleaved with charged residues, rapid unbinding of the applied peptide was facilitated, producing an unusually fast rising phase of resurgent current. Given the sensitivity of the kinetics of block and unblock to the precise sequence and identity of amino acids in the synthetic peptides, it is interesting to note that, in the current protein database, the sequence LITFILKK occurs only in proteins containing the full cytoplasmic tail of the sodium channel β4 subunit.
PY - 2005/1/20
Y1 - 2005/1/20
N2 - Voltage-gated sodium channels with "resurgent" kinetics are specialized for high-frequency firing. The α subunits interact with a blocking protein that binds open channels upon depolarization and unbinds upon repolarization, producing resurgent sodium current. By limiting classical inactivation, the cycle of block and unblock shortens refractory periods. To characterize the blocker in Purkinje neurons, we briefly exposed inside-out patches to substrate-specific proteases. Trypsin and chymotrypsin each removed resurgent current, consistent with established roles for positively charged and hydrophobic/aromatic groups in blocking sodium channels. In Purkinje cells, the only known sodium channel-associated subunit that has a cytoplasmic sequence with several positive charges and clustered hydrophobic/aromatic residues is β4 (KKLITFILKKTREK; β4154-167). After enzymatic removal of block, β4154-167 fully reconstituted resurgent current, whereas scrambled or point-mutated peptides were ineffective. In CA3 pyramidal neurons, which lack β4 and endogenous block, β4154-167 generated resurgent current. Thus, β4 may be the endogenous open-channel blocker responsible for resurgent kinetics.
AB - Voltage-gated sodium channels with "resurgent" kinetics are specialized for high-frequency firing. The α subunits interact with a blocking protein that binds open channels upon depolarization and unbinds upon repolarization, producing resurgent sodium current. By limiting classical inactivation, the cycle of block and unblock shortens refractory periods. To characterize the blocker in Purkinje neurons, we briefly exposed inside-out patches to substrate-specific proteases. Trypsin and chymotrypsin each removed resurgent current, consistent with established roles for positively charged and hydrophobic/aromatic groups in blocking sodium channels. In Purkinje cells, the only known sodium channel-associated subunit that has a cytoplasmic sequence with several positive charges and clustered hydrophobic/aromatic residues is β4 (KKLITFILKKTREK; β4154-167). After enzymatic removal of block, β4154-167 fully reconstituted resurgent current, whereas scrambled or point-mutated peptides were ineffective. In CA3 pyramidal neurons, which lack β4 and endogenous block, β4154-167 generated resurgent current. Thus, β4 may be the endogenous open-channel blocker responsible for resurgent kinetics.
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U2 - 10.1016/j.neuron.2004.12.035
DO - 10.1016/j.neuron.2004.12.035
M3 - Article
C2 - 15664175
AN - SCOPUS:12344303542
VL - 45
SP - 233
EP - 244
JO - Neuron
JF - Neuron
SN - 0896-6273
IS - 2
ER -