Ion permeation and selectivity of wild-type recombinant rat CNG (rOCNC1) channels expressed in HEK293 cells.
The permeation properties of adenosine 3', 5'-cyclic monophosphate (cAMP)-activated recombinant rat olfactory cyclic nucleotide-gated channels (rOCNC1) in human embryonic kidney (HEK 293) cells were investigated using inside-out excised membrane patches. The relative permeability of these rOCNC1 channels to monovalent alkali cations and organic cations was determined from measurements of the changes in reversal potential upon replacing sodium in the bathing solution with different test cations. The permeability ratio of Cl(-) relative to Na(+) (P(Cl)/P(Na)) was about 0.14, confirming that these channels are mainly permeable to cations. The sequence of relative permeabilities of monovalent alkali metal ions in these channels was P(Na) > or = P(K) > P(Li) > P(Cs) > or = P(Rb), which closely corresponds to a high-strength field sequence as previously determined for native rat olfactory receptor neurons (ORNs). The permeability sequence for organic cations relative to sodium was P(NH3OH) > P(NH4) > P(Na) > P(Tris) > P(Choline) > P(TEA), again in good agreement with previous permeability ratios obtained in native rat ORNs. Single-channel conductance sequences agreed surprisingly well with permeability sequences. These conductance measurements also indicated that, even in asymmetric bi-ionic cation solutions, the conductance was somewhat independent of current direction and dependent on the composition of both solutions. These results indicate that the permeability properties of rOCNC1 channels are similar to those of native rat CNG channels, and provide a suitable reference point for exploring the molecular basis of ion selectivity in recombinant rOCNC1 channels using site-directed mutagenesis.
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School of Physiology and Pharmacology, The University of New South Wales, Sydney 2052, Australia.
Zhu
X O
XO
Moorhouse
A J
AJ
Bieri
S
S
Cunningham
A M
AM
Barry
P H
PH
D000818:Animals; D002414:Cations, Monovalent / Q000378:metabolism*; D002460:Cell Line; D002463:Cell Membrane Permeability; D002712:Chlorides / Q000378:metabolism; D000242:Cyclic AMP / Q000494:pharmacology; D054815:Cyclic Nucleotide-Gated Cation Channels; D004553:Electric Conductivity; D006801:Humans; D007473:Ion Channels / Q000737:chemistry* / Q000235:genetics; D008564:Membrane Potentials; D008672:Metals, Alkali / Q000378:metabolism; D018408:Patch-Clamp Techniques; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry; D012964:Sodium / Q000378:metabolism; D014162:Transfection
D002414:Cations, Monovalent; D002712:Chlorides; D054815:Cyclic Nucleotide-Gated Cation Channels; D007473:Ion Channels; D008672:Metals, Alkali; D011994:Recombinant Proteins; D012964:Sodium; D000242:Cyclic AMP