Muscarinic agonists and ATP increase the intracellular Ca2+ concentration in chick cochlear hair cells.
1. Cholinergic muscarinic agonists applied by the pressure puff method increased intracellular Ca2+ concentration in Fura-2-loaded hair cells. The Ca2+ response outlasted the agonist application. 2. The Ca2+ response induced by acetylcholine (ACh) was ACh dose dependent with a KD of 200 microM. Desensitization was negligible, and almost identical Ca2+ responses were observed when two ACh puffs were separated by 150 s. The response was blocked by d-tubocurarine (dTC). The KD of dTC blocking was 500 microM when 100 microM-ACh induced the Ca2+ response. 3. The amplitude of the ACh-induced Ca2+ responses were potentiated to 3 times the control by incubation with calcitonin gene-related peptide (CGRP; 0.1-1 microM). CGRP did not affect the resting Ca2+ concentration. Glycine (100 microM) potentiated the ACh response to 1.4 times the control, and also increased the resting Ca2+ concentration slightly. 4. The ACh-induced Ca2+ response was suppressed by atropine. It was induced in Ca2(+)-free extracellular medium, and in Ca2(+)-free medium desensitization to a second ACh stimulation was significant. The amplitude of the second Ca2+ response was 44% of the first when two ACh puffs were separated by 117 s in Ca2+ free medium. 5. Muscarine and carbamylcholine induced similar Ca2+ responses, with KD values of 130 microM for muscarine and 340 microM for carbamylcholine. Desensitization of Ca2+ responses was negligible in both agonists. 6. ATP co-exists with ACh in some presynaptic nerve terminals (Burnstock, 1981). Puff-applied ATP (100 microM) generated a Ca2+ response with a rapid rising phase and a following slow phase. In Ca2(+)-free medium the rapid phase disappeared and only the slow phase was observed. The rapid phase is due to the influx of Ca2+ ions and the slow phase is due to a release of Ca2+ ions from an intracellular reservoir. Under voltage clamp ATP induced a fast inward current and a following slow outward current. 7. Nicotine, adenosine, glycine, GABA, glutamate and bradykinin did not induce Ca2+ responses in the hair cell. 8. ACh induced hyperpolarization of the hair cell membrane under current clamp, most probably by the activation of Ca2+ activated K+ conductance. Therefore, a cholinergic muscarinic receptor may mediate the inhibitory effects of efferent innervation observed in hair cells.
Shigemoto
T
T
National Institute for Physiological Sciences, Okazaki, Japan.
Ohmori
H
H
D000109:Acetylcholine / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000494:pharmacology*; D000818:Animals; D001285:Atropine / Q000494:pharmacology; D015740:Calcitonin Gene-Related Peptide / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002217:Carbachol / Q000494:pharmacology; D002645:Chickens; D004305:Dose-Response Relationship, Drug; D005998:Glycine / Q000494:pharmacology; D006198:Hair Cells, Auditory / Q000187:drug effects / Q000378:metabolism*; D066298:In Vitro Techniques; D009116:Muscarine / Q000494:pharmacology; D010277:Parasympathomimetics / Q000494:pharmacology*; D014403:Tubocurarine / Q000494:pharmacology
D010277:Parasympathomimetics; D001285:Atropine; D009116:Muscarine; D000255:Adenosine Triphosphate; D002217:Carbachol; D015740:Calcitonin Gene-Related Peptide; D000109:Acetylcholine; D002118:Calcium; D005998:Glycine; D014403:Tubocurarine