A voltage- and Ca2+-dependent big conductance K channel in cochlear spiral ligament fibrocytes.
Evidence is accruing that spiral ligament fibrocytes (SLFs) play an important role in cochlear K(+) homeostasis, but little direct physiological data is available to support this concept. Here we report the presence and characterization of a voltage- and Ca(2+)-dependent big-conductance K (BK) channel in type I SLFs cultured from the gerbil cochlea. A single-channel conductance of 298+/-5.6 pS (n=28) was measured under symmetrical K(+). Membrane potentials for half-maximal open probability (P(o)) were -67, -45 and 85 mV with cytosolic free-Ca(2+) levels of 0.7 mM, 10 microM and 1 microM, respectively (n=8-14). The Hill coefficient for Ca(2+) affinity was 1.9 at a membrane potential of 60 mV (n=6). The BK channel showed very low activity (P(o)=0.0019, n=5) under normal physiological conditions, suggesting a low resting intracellular free [Ca(2+)]. Pharmacological results fit well with the profile of classic BK channels. The estimated half-maximal inhibitory concentration and Hill coefficient for tetraethylammonium were 0.086+/-0.021 mM and 0.99, respectively (n=4-9). In whole cell recordings, the voltage-activated outward K current was inhibited 85.7+/-4.5% (n=6) by 0.1 microM iberiotoxin. A steady-state kinetic model with two open and two closed stages best described the BK gating process (tau(o1) 0.23+/-0.08 ms, tau(o2) 1.40+/-0.32 ms; tau(c1) 0.26+/-0.09 ms, tau(c2) 3.10+/-1.2 ms; n=11). RT-PCR analyses revealed a splice variant of the BK channel alpha subunit in cultured type I SLFs and freshly isolated spiral ligament tissues. The BK channel is likely to play a major role in regulating the membrane potential of type I SLFs, which may in turn influence K(+) recycling dynamics in the mammalian cochlea.
Liang
F
F
Department of Pathology and Laboratory Medicine, Medical University of South Carolina, 171 Ashley Avenue, PO Box 250908, Charleston, SC 29425, USA.
Niedzielski
A
A
Schulte
B A
BA
Spicer
S S
SS
Hazen-Martin
D J
DJ
Shen
Z
Z
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002478:Cells, Cultured; D003051:Cochlea / Q000166:cytology / Q000502:physiology*; D003053:Cochlear Duct / Q000166:cytology / Q000502:physiology*; D005260:Female; D005347:Fibroblasts / Q000502:physiology / Q000648:ultrastructure; D015870:Gene Expression / Q000502:physiology; D005849:Gerbillinae; D006706:Homeostasis / Q000502:physiology; D007150:Immunohistochemistry; D015640:Ion Channel Gating / Q000502:physiology*; D051037:Large-Conductance Calcium-Activated Potassium Channel alpha Subunits; D051036:Large-Conductance Calcium-Activated Potassium Channels; D008022:Ligaments / Q000166:cytology / Q000502:physiology*; D008297:Male; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D008854:Microscopy, Electron; D011188:Potassium / Q000378:metabolism; D026902:Potassium Channel Blockers / Q000494:pharmacology; D024681:Potassium Channels, Calcium-Activated / Q000235:genetics / Q000378:metabolism*; D019789:Tetraethylammonium / Q000494:pharmacology
D051037:Large-Conductance Calcium-Activated Potassium Channel alpha Subunits; D051036:Large-Conductance Calcium-Activated Potassium Channels; D026902:Potassium Channel Blockers; D024681:Potassium Channels, Calcium-Activated; D019789:Tetraethylammonium; D011188:Potassium; D002118:Calcium