Characterization of a nicotinamide-adenine dinucleotide-dependent cation channel in the CRI-G1 rat insulinoma cell line.
1. Cell-free excised membrane patches were used to examine the properties of a novel nicotinamide-adenine dinucleotide (beta-NAD+)-activated ion channel in the rat insulin-secreting cell line, CRI-G1. 2. In inside-out recordings, beta-NAD+ (0.05-1.0 mM) induced the appearance of a channel characterized by extremely slow kinetics, with mean open times in the range of seconds. The estimated EC50 for activation was 114 microM. Channel activity declined with time (run-down) following activation by beta-NAD+ in excised patches and this was not prevented by intracellular application of trypsin. 3. The single channel current-voltage relationship was linear with a conductance of 74 pS in symmetrical NaCl. The channel appears equally permeable to Na+, K+ and Cs+, exhibits an appreciable permeability to Ca2+, Mg2+ and Ba2+, but excludes anions. 4. The channel displays an unusual voltage sensitivity, with an abrupt increase in open-state probability at depolarized voltages. 5. Channel opening, in the presence of beta-NAD+, required both Ca2+ and Mg2+ to be present at the internal side of the membrane. Activation by Ca2+ required a concentration of at least 10 microM and was maximal at 0.1 mM. Ba2+ did not substitute for Ca2+ in inducing channel activity nor did it inhibit activation by Ca2+. Increasing the concentration of intracellular Mg2+ stabilized the open state of NAD(+)-activated channels. 6. The non-selective cation channel reported here differs in its gating and modulatory characteristics from non-selective cation channels described in other tissues. This channel may play a role in the pathophysiological responses of beta-cells to oxidative stress.
Herson
P S
PS
Department of Biomedical Sciences, University of Aberdeen, Foresterhill, UK.
Dulock
K A
KA
Ashford
M L
ML
D000818:Animals; D001464:Barium / Q000494:pharmacology; D002118:Calcium / Q000494:pharmacology; D004594:Electrophysiology; D007340:Insulinoma; D015640:Ion Channel Gating / Q000502:physiology; D007473:Ion Channels / Q000502:physiology*; D007515:Islets of Langerhans / Q000378:metabolism*; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D008564:Membrane Potentials / Q000502:physiology; D009243:NAD / Q000494:pharmacology*; D018384:Oxidative Stress; D018408:Patch-Clamp Techniques; D051381:Rats; D014407:Tumor Cells, Cultured
D007473:Ion Channels; D009243:NAD; D001464:Barium; D008274:Magnesium; D002118:Calcium