Characterization of a stretch-activated potassium channel in chondrocytes.
Chondrocytes possess the capacity to transduce load-induced mechanical stimuli into electrochemical signals. The aim of this study was to functionally characterize an ion channel activated in response to membrane stretch in isolated primary equine chondrocytes. We used patch-clamp electrophysiology to functionally characterize this channel and immunohistochemistry to examine its distribution in articular cartilage. In cell-attached patch experiments, the application of negative pressures to the patch pipette (in the range of 20-200 mmHg) activated ion channel currents in six of seven patches. The mean activated current was 45.9 +/- 1.1 pA (n = 4) at a membrane potential of 33 mV (cell surface area approximately 240 microm(2)). The mean slope conductance of the principal single channels resolved within the total stretch-activated current was 118 +/- 19 pS (n = 6), and reversed near the theoretical potassium equilibrium potential, E(K+), suggesting it was a high-conductance potassium channel. Activation of these high-conductance potassium channels was inhibited by extracellular TEA (K(d) approx. 900 microM) and iberiotoxin (K(d) approx. 40 nM). This suggests that the current was largely carried by BK-like potassium (MaxiK) channels. To further characterize these BK-like channels, we used inside-out patches of chondrocyte membrane: we found these channels to be activated by elevation in bath calcium concentration. Immunohistochemical staining of equine cartilage samples with polyclonal antibodies to the alpha1- and beta1-subunits of the BK channel revealed positive immunoreactivity for both subunits in superficial zone chondrocytes. These experiments support the hypothesis that functional BK channels are present in chondrocytes and may be involved in mechanotransduction and chemotransduction.
Mobasheri
Ali
A
Musculoskeletal Research Group, Division of Veterinary Medicine, Faculty of Medicine and Health Sciences, University of Nottingham, Leicestershire, United Kingdom.
Lewis
Rebecca
R
Maxwell
Judith E J
JE
Hill
Claire
C
Womack
Matthew
M
Barrett-Jolley
Richard
R
D000818:Animals; D002356:Cartilage / Q000166:cytology / Q000378:metabolism*; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism / Q000648:ultrastructure; D019902:Chondrocytes / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D006736:Horses; D015640:Ion Channel Gating / Q000187:drug effects / Q000502:physiology; D051036:Large-Conductance Calcium-Activated Potassium Channels / Q000187:drug effects / Q000378:metabolism*; D040542:Mechanotransduction, Cellular / Q000502:physiology; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D009924:Organ Culture Techniques; D018408:Patch-Clamp Techniques; D011188:Potassium / Q000378:metabolism; D026902:Potassium Channel Blockers / Q000494:pharmacology; D011312:Pressure / Q000009:adverse effects; D021122:Protein Subunits / Q000187:drug effects / Q000378:metabolism; D013314:Stress, Mechanical; D016474:Weight-Bearing / Q000502:physiology
D051036:Large-Conductance Calcium-Activated Potassium Channels; D026902:Potassium Channel Blockers; D021122:Protein Subunits; D011188:Potassium