Channel activity of recombinant pannexin 1 |
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Authors: | R A Romanov M F Bystrova O A Rogachevskaya S S Kolesnikov |
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Institution: | 1.Institute of Cell Biophysics,Russian Academy of Sciences,Pushchino, Moscow oblast,Russia |
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Abstract: | Mammalian taste cells of the type II release ATP, an afferent neurotransmitter, by employing unselective ATP-permeable ion
channels. The molecular identity of these channels is not known with confidence, although evidence implicates certain channel
proteins from the connexin and pannexin families as most likely candidates. Here we carried out the comparative analysis of
biophysical features and pharmacological profiles of unselective channels operative in type II cells and recombinant pannexin
1 (Panx1), which was cloned from the taste tissue and heterologously expressed in eukaryotic cells of several lines, including
HEK-293, CHO, and neuroblastoma SK-N-SH. Integral currents mediated by Panx1 hemichannels were recorded to elucidate their
kinetics characteristics, such as activation and deactivation, voltage dependence, and sensitivity to a variety of blockers,
including carbenoxolone, DIDS, and NPPB. It was shown that the heterologous expression of Panx1 in cells of each type induced
specific conductance, which exhibited outward rectification and was effectively blockable with carbenoxolone and anionic channel
blockers DIDS and NPPB. Panx1 activity was studied at the single channel level as well. As was found, transfection of HEK-293
cells with the plasmid harboring cDNA encoding Panx1 gave rise to single channel current-like events in excised patches that
were inhibited by 20 μM carbenoxolone, the relatively specific blocker of Panx1. These carbenoxolone-sensitive channels were
peculiar in that single-channel current versus membrane voltage was not linear but exhibited outward rectification. In addition,
the open-channel probability strongly increased with membrane voltage. Taken together, the data obtained here and earlier
demonstrate clearly that by their biophysical and pharmacological features, ATP-permeable channels operative in type II cells
are rather distinct from recombinant Panx1 hemichannels, thus arguing against Panx1 as the main conduit of ATP release in
taste cells. |
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