Ethanol Inhibition of a T-Type Ca2+ Channel Through Activity of Protein Kinase C
Article first published online: 12 MAR 2013
Copyright © 2013 by the Research Society on Alcoholism
Alcoholism: Clinical and Experimental Research
Volume 37, Issue 8, pages 1333–1342, August 2013
How to Cite
Shan, H. Q., Hammarback, J. A. and Godwin, D. W. (2013), Ethanol Inhibition of a T-Type Ca2+ Channel Through Activity of Protein Kinase C. Alcoholism: Clinical and Experimental Research, 37: 1333–1342. doi: 10.1111/acer.12098
- Issue published online: 26 JUL 2013
- Article first published online: 12 MAR 2013
- Manuscript Accepted: 2 JAN 2013
- Manuscript Received: 18 AUG 2012
- NIAAA. Grant Numbers: AA014106, AA017056, AA016852, AA016852-04S1
- Tab Williams Family
- Protein Kinase C;
- Phorbol 12-Myristate 13-Acetate;
- CaV3.2 T-Channel;
- Dorsal Root Ganglion
T-type calcium channels (T-channels) are widely distributed in the central and peripheral nervous system, where they mediate calcium entry and regulate the intrinsic excitability of neurons. T-channels are dysregulated in response to alcohol administration and withdrawal. We therefore investigated acute ethanol (EtOH) effects and the underlying mechanism of action in human embryonic kidney (HEK) 293 cell lines, as well as effects on native currents recorded from dorsal root ganglion (DRG) neurons cultured from Long-Evans rats.
Whole-cell voltage-clamp recordings were performed at 32 to 34°C in both HEK cell lines and DRG neurons. The recordings were taken after a 10-minute application of EtOH or protein kinase C (PKC) activator (phorbol 12-myristate 13-acetate [PMA]).
We recorded T-type Ca2+ currents (T-currents) from 3 channel isoforms (CaV3.1, CaV3.2, and CaV3.3) before and during administration of EtOH. We found that only 1 isoform, CaV3.2, was significantly affected by EtOH. EtOH reduced current density as well as producing a hyperpolarizing shift in steady-state inactivation of both CaV3.2 currents from HEK 293 cell lines and in native T-currents from DRG neurons that are known to be enriched in CaV3.2. A myristoylated PKC peptide inhibitor (MPI) blocked the major EtOH effects, in both the cell lines and the DRG neurons. However, PMA effects were more complex. Lower concentration PMA (100 nM) replicated the major effects of EtOH, while higher concentration PMA (1 μM) did not, suggesting that the EtOH effects operate through activation of PKC and were mimicked by lower concentration of PMA.
EtOH primarily affects the CaV3.2 isoform of T-type Ca2+ channels acting through PKC, highlighting a novel target and mechanism for EtOH effects on excitable membranes.