Closure of Gap Junction Channels by Arylaminobenzoates

• Published in: Molecular pharmacology 2003-06, Vol.63 (6), p.1389-1397
• Main Authors: Srinivas, Miduturu, Spray, David C
• Format: Article
• Language: English
• Subjects: Animals; Anti-Inflammatory Agents, Non-Steroidal - pharmacology; Cells, Cultured; Connexins - drug effects; Connexins - genetics; Connexins - physiology; Dose-Response Relationship, Drug; Electrophysiology; Flufenamic Acid - pharmacology; Gap Junctions - drug effects; Gap Junctions - physiology; Humans; Ion Channels - drug effects; Ion Channels - physiology; Mice; Transfection
• ISSN: 0026-895X; 1521-0111
• DOI: 10.1124/mol.63.6.1389

We determined the effect of flufenamic acid (FFA) and related derivatives on gap junction channel currents, applying the dual whole-cell patch-clamp technique to pairs of N2A neuroblastoma cells transfected with various connexins. FFA reduced gap junction channel currents in a reversible and concentration-dependent manner. Half-maximal concentrations for FFA-induced reduction of junctional conductance in cell pairs coupled by different connexins were similar (20 to 60 μM), indicating that FFA does not greatly discriminate between connexin subtypes. Hill coefficients for blockade were approximately 3, indicating a high degree of cooperativity. Analogs of FFA also reduced junctional conductance with similar potencies, whereas other unrelated chloride channel blockers had no effect. Inhibition of gap junction channels by FFA (p K a ∼ 3.8) was increased at low external pH, suggesting that the uncharged form of the drug is important for blockade. The effect of FFA did not seem to be mediated by direct binding of the drug to the pore of the gap junction channel. Internal application of high concentrations of FFA by addition to patch pipettes did not cause inhibition of channel currents. The magnitude of inhibition was neither voltage-dependent nor influenced by the nature of permeant ion. Single-channel recordings indicated that FFA reduced the channel-open probability without modifying the current amplitude and induced slow transitions between open and closed states. We propose that FFA inhibits gap junctions by inducing a conformational change in the protein upon binding to a site that is presumably located within the membrane.