Activation of the Recombinant Human α7 Nicotinic Acetylcholine Receptor Significantly Raises Intracellular Free Calcium
- O. Delbono1,1,2,
- M. Gopalakrishnan3,
- M. Renganathan2,
- L. M. Monteggia3,
- M. L. Messi1 and
- J. P. Sullivan3
- Departments of 1Physiology and Pharmacology (O.D., M.L.M.) and2Internal Medicine (Gerontology) (O.D., M.R.), Bowman Gray School of Medicine of the Wake Forest University, Winston-Salem, North Carolina and 3Neuroscience Research (D-47W) (M.G., L.M.M., J.P.S.), Abbott Laboratories, Abbott Park, Illinois
Abstract
The α7 nicotinic acetylcholine receptor (nAChR) subtype, unlike other neuronal nicotinic receptors, exhibits a relatively high permeability to Ca++ ions. Although Ca++ entry through this receptor subtype has been implicated in various Ca++-dependent processes in the central nervous system, little is known about how this receptor modulates mammalian intracellular Ca++ dynamics. Intracellular Ca++responses evoked by activation of the human α7 nAChRs stably expressed in HEK-293 (human embryonic kidney) cells were studied. Inward current and intracellular Ca++ transients were recorded simultaneously in response to a fast drug application system. Current recordings under whole-cell voltage-clamp and fast ratiometric intracellular Ca++ imaging acquisition were synchronized to drug pulses. The mean peak [Ca++]i observed with 100 μM (−)-nicotine was 356 ± 48 nM (n = 8). The magnitude of the intracellular Ca++ elevation corresponds to a 20% fractional current carried by Ca++ ions. The EC50 of the intracellular Ca++ responses for (−)-nicotine, (±)-epibatidine, 1,1 dimethyl-4-phenyl-piperazinium and acetylcholine were 51, 3.5, 75 and 108 μM, respectively. These EC50 values strongly correlate with those recorded for the cationic inward current through α7 nAChR. α-Bungarotoxin, methyllcaconitine or extracellular Ca++chelation ablated (−)-nicotine-evoked increase in intracellular Ca++ concentration. This study provides evidence that cation influx through the human α7 nAChR is sufficient to mediate a significant, transient, rise in intracellular Ca++ concentration.
Footnotes
-
Send reprint requests to: Dr. Osvaldo Delbono, Dept. of Physiology and Pharmacology, Bowman Gray School of Medicine, Medical Center Boulevard, Winston-Salem, NC 27157.
-
↵1 Research in the laboratory of O.D. was supported by grants from the National Institutes of Health 2-P60AG10484, T-32-AG00182 and K01 AG00692 and from the Muscular Dystrophy Association (U.S.A.).
- Abbreviations:
- HEK293
- human embryonic kidney 293 cells
- nAChR
- nicotinic acetylcholine receptor
- EGTA
- ethylene glycol-bis(β-aminoethyl ether)-N,N,N′,N′-tetraacetic acid
- NMDA
- N-methyl-d-aspartate
- IP3
- inositol 1,4,5-triphosphate
- MLA
- methyllycaconitine
- DMPP
- 1,1 dimethyl-4-phenyl-piperazinium
- ABT-418
- (S)-3-methyl-5-(1-methyl-2-pyrrolidinyl) isoxazole
- HEPES
- N-[2-hydroxyethyl]piperazine-N′-[2-ethanesulfonic acid]
- PBS
- phosphate-buffered saline
-
- Received May 2, 1996.
- Accepted September 3, 1996.
- The American Society for Pharmacology and Experimental Therapeutics
