Calcium signalling mediated by the 9 acetylcholine receptor in a cochlear cell line from the immortomouse.
1. We have investigated the characteristics of the alpha9 acetylcholine receptor (alpha9AChR) expressed in hair cell precursors in an immortalized cell line UB/OC-2 developed from the organ of Corti of the transgenic H-2Kb-tsA58 mouse (the Immortomouse) using both calcium imaging and whole-cell recording. 2. Ratiometric measurements of fura-2 fluorescence revealed an increase of intracellular calcium concentration in cells when challenged with 10 microM ACh. The calcium increase was seen in 66 % of the cells grown at 39 degrees C in differentiated conditions. A sm aller fraction (34%) of cells grown at 33 degrees C in proliferative con ditions responded. 3. Caffeine (10mM) elevated cell calcium. In the ab sence of caffeine, the majority of imaged cells responded only once to A Ch presentations. Pretreatment with caffeine ingibited all calcium respo nses to ACh. 4. In whole-cell tight-seal recordings 10 microM ACh activa ted inward current was dependent on the extracellular calcium concentrat ion with an estimated PCa/PNa of 80 for the alpha9 receptor at physiological calcium levels. 5 . The data indicate that ACh activates a calcium-permeable channel alpha 9AChR in UB/OC-2 cells and that the channel has a significantly higher c alcium permeability than other AChRs. The results indicate that the alp ha9AChR may be able to elevate intracellular calcium levels in hair cell s both directly and via store release.
Jagger
D J
DJ
Department of Physiology, University College London, Gower Street, London WC1E 6BT, UK.
Griesinger
C B
CB
Rivolta
M N
MN
Holley
M C
MC
Ashmore
J F
JF
D000109:Acetylcholine / Q000494:pharmacology; D000818:Animals; D002110:Caffeine / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D020013:Calcium Signaling / Q000187:drug effects; D002460:Cell Line; D003051:Cochlea / Q000294:innervation / Q000378:metabolism / Q000502:physiology*; D004553:Electric Conductivity; D006198:Hair Cells, Auditory / Q000378:metabolism* / Q000502:physiology*; D051379:Mice; D008822:Mice, Transgenic; D011978:Receptors, Nicotinic / Q000502:physiology*; D013050:Spectrometry, Fluorescence
C402445:Chrna9 protein, mouse; D011978:Receptors, Nicotinic; D002110:Caffeine; D000109:Acetylcholine; D002118:Calcium