Characterization of NaV1.6-mediated Na+ currents in smooth muscle cells isolated from mouse vas deferens.
Patch-clamp experiments were performed to investigate the behavior of voltage-activated inward currents in vas deferens myocytes from Na(V)1.6-null mice (Na(V)1.6(-/-)) lacking the expression of the Na(+) channel gene, Scn8a, and their wild-type littermates (Na(V)1.6(+/+)). Immunohistochemistry confirmed expression of Na(V)1.6 in the muscle of Na(V)1.6(+/+), but not Na(V)1.6(-/-), vas deferens. PCR analysis revealed that the only beta(1)-subunit gene expressed in Na(V)1.6(+/+) vas deferens was Scn1b. In Na(V)1.6(+/+) myocytes, the threshold for membrane currents evoked by 20 msec voltage ramps (-100 mV to 60 mV) was -38.5 +/- 4.6 mV and this was shifted to a more positive potential (-31.2 +/- 4.9 mV) by tetrodotoxin (TTX). In Na(V)1.6(-/-) myocytes, the threshold was -30.4 +/- 3.4 mV and there was no TTX-sensitive current. The Na(+) current (I(Na)) in Na(V)1.6(+/+) myocytes had a bell-shaped current-voltage relationship that peaked at approximately -10 mV. Increasing the duration of the voltage ramps beyond 20 msec reduced the peak amplitude of I(Na). I(Na) displayed both fast (tau approximately 10 msec) and slow (tau approximately 1 sec) recovery from inactivation, the magnitude of the slow component increasing with the duration of the conditioning pulse (5-40 msec). During repetitive activation (5-40 msec pulses), I(Na) declined at stimulation frequencies > 0.5 Hz and at 10 Hz <or= 50% of the current remained. These findings indicate that I(Na) is due solely to Na(V)1.6 in Na(V)1.6(+/+) myocytes. The gating properties of these channels suggest they play a major role in regulating smooth muscle excitability, particularly in response to rapid depolarizing stimuli.
Zhu
Hai-Lei
HL
Department of Pharmacology, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan.
Shibata
Atsushi
A
Inai
Tetsuichiro
T
Nomura
Masatoshi
M
Shibata
Yosaburo
Y
Brock
James A
JA
Teramoto
Noriyoshi
N
D000818:Animals; D002469:Cell Separation; D004558:Electric Stimulation; D007150:Immunohistochemistry; D015640:Ion Channel Gating; D007700:Kinetics; D008297:Male; D008564:Membrane Potentials; D051379:Mice; D008809:Mice, Inbred C3H; D018345:Mice, Knockout; D009130:Muscle, Smooth / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D032389:Myocytes, Smooth Muscle / Q000187:drug effects / Q000378:metabolism*; D062557:NAV1.6 Voltage-Gated Sodium Channel; D009419:Nerve Tissue Proteins / Q000037:antagonists & inhibitors / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D018408:Patch-Clamp Techniques; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012964:Sodium / Q000378:metabolism*; D026941:Sodium Channel Blockers / Q000494:pharmacology; D015222:Sodium Channels / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D013779:Tetrodotoxin / Q000494:pharmacology; D014649:Vas Deferens / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D062547:Voltage-Gated Sodium Channel beta-1 Subunit
D062557:NAV1.6 Voltage-Gated Sodium Channel; D009419:Nerve Tissue Proteins; D012333:RNA, Messenger; C524974:Scn1b protein, mouse; C491046:Scn8a protein, mouse; D026941:Sodium Channel Blockers; D015222:Sodium Channels; D062547:Voltage-Gated Sodium Channel beta-1 Subunit; D013779:Tetrodotoxin; D012964:Sodium