Cholinergic muscarinic receptors in rat cochlea

doi:10.1016/0006-8993(88)90682-8

Brain Research

Volume 474, Issue 1, 22 November 1988, Pages 185-188

https://doi.org/10.1016/0006-8993(88)90682-8 Get rights and content

Abstract

Specific³H-1-quinuclidinylbenzilate (³H-1-QNB) binding to rat cochlea homogenates occurs to a homogeneous class of binding sites withK_d = 0.13 ± 0.01nMandB_max = 0.57 ± 0.07fmolper cochlea. Binding is stereoselectivity inhibited by benzetimide enantiomers. Dexetimide was more effective than levetimide in displacing³H-1-QNB from its binding sites (K_i = 4 × 10⁻¹⁰Mand 6.5 × 10⁻⁶M, respectively). Pirenzepine inhibits³H-1-QNB binding with low affinity (K_i = 2 × 10⁻⁶M), classifying the binding sites as muscarinic M₂ receptors.

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Cited by (14)

Muscarinic receptor-mediated calcium signaling in spiral ganglion neurons of the mammalian cochlea
1999, Brain Research
Using indo-1 microspectrofluorometry, we examined the effects of cholinergic agonists on the concentration of intracellular Ca²⁺ ions ([Ca²⁺]_i) in spiral ganglion neurons, isolated from rat cochleae at different stages of post-natal development (from P3 to P30). Extracellular application of acetylcholine (ACh) or carbamylcholine generated a rapid and transient increase in [Ca²⁺]_i. The ACh concentration-response curve indicated an apparent dissociation constant (K_d) of 8 μM and a Hill coefficient of 1.0. Removing extracellular free Ca²⁺ did not suppress the ACh-induced Ca²⁺ responses suggesting an intracellular Ca²⁺-release mechanism. When we compared the cholinergic response at different stages of postnatal development, there were no significant differences on the aspect of the Ca²⁺ response and the percentage of responsive neurons, which ranged between 50 and 65% per cochlear preparation. The application of muscarine triggered reversible Ca²⁺ responses similar to those observed with ACh, with an apparent K_d of 10 μM and a Hill coefficient of 1.0. The cholinergic-induced Ca²⁺ response was reversibly blocked by muscarinic antagonists with the following order of potency, atropine>4-DAMP>methoctramine>pirenzepine. Nicotine (10 to 100 μM) did not evoke Ca²⁺ responses and the nicotinic antagonist curare (10 μM) did not block the ACh-evoked responses. The present study is the first direct demonstration of functional muscarinic receptors (mAChRs) in spiral ganglion neurons. These mAChRs activated by the cholinergic lateral efferent system may participate in the regulation of the electrical activity of the afferent auditory fibers contacting the inner hair cells.
GNAZ in human fetal cochlea: expression, localization, and potential role in inner ear function
1995, Hearing Research
Dissociation of an activated α-subunit from the β-γ complex directly regulates secondary messenger proteins. To address the potential role of G proteins expressed in human fetal cochlea, degenerate oligonucleotide primers corresponding to the 3′-end of the conserved region of α-subunits were used for polymerase chain reaction amplification of reverse-transcribed total human fetal cochlear mRNAS; GNAZ and GNAQ were isolated. These two G proteins are unique among the G-protein family because they lack a typical pertussis modification site. GNAZ is expressed in high levels in neural tissue while GNAQ is ubiquitously expressed. We characterized GNAZ expression using Northern blots, tissue in-situ hybridization and immunohistochemistry techniques to elucidate the potential role of this protein in inner ear function. Our data suggest that GNAZ may play a role in maintaining the ionic balance of perilymphatic and endolymphatic cochlear fluids.
Effects of ototoxins on quinuclidinyl benzylate binding in the rat cochlea
1994, Neuroscience Letters
Ototoxins inhibit the muscarinic receptor-activated inositol phosphate synthesis in the rat cochlea. In order to study this inhibitory mechanism, we investigated the effects of the ototoxins ethacrynate, cisplatin, HgCl₂ and neomycin on [³H]quinuclidinyl benzylate binding to muscarinic receptors in adult and 12-day-old rat cochleas. The results are similar whatever the age: at concentrations that inhibit the inositol phosphate synthesis, ethacrynate is without effect. Neomycin only reduces [³H]quinuclidinyl benzylate binding at concentrations in the millimolar range. Cisplatin and HgCl₂ block the binding in a dose-dependent way. These results suggest that the block of the transduction system by cisplatin and HgCl₂ is due to direct interactions with muscarinic binding sites. Moreover, considering these data together with previous results, ethacrynate and neomycin may affect the phosphoinositide signalling pathway at targets including phosphoinositides and G proteins.
A nicotinic-like receptor mediates suppression of distortion product otoacoustic emissions by contralateral sound
1994, Hearing Research
The purpose of this investigation was to provide in vivo pharmacologic characterization of a cholinergic receptor mediating the suppressive effects of medial olivocochlear (MOC) efferent activation. MOC neurons were activated by contralateral sound and the resulting suppression of ipsilateral distortion product otoacoustic emissions (DPOAEs) was monitored before and after intracochlear perfusions of cholinergic antagonists. Results revealed a dose-dependent blockade of contralateral suppression of DPOAEs by a wide variety of nicotinic and muscarinic cholinergic receptor antagonists, as well as by non-traditional antagonists of cholinergic activity. The nicotinic antagonists, α-bungarotoxin, curare and k-bungarotoxin, and the glycine antagonist, strychnine, blocked contralateral suppression at nanomolar concentrations and demonstrated similar potencies. IC₅₀ values were 2.38 × 10⁻⁷, 2.79 × 10⁻⁷, 3.81 × 10⁻⁷ and 2.96 × 10⁻⁷ M, respectively. These agents were followed in potency by the nicotinic antagonist, trimethaphan (1.75 × 10⁻⁶ M), the M₃ muscarinic antagonist, 4-DAMP (1.88 × 10⁻⁶ M) and the GABA_A antagonist, bicuculline (2.39 × 10⁻⁶ M). Increasingly greater concentrations of the muscarinic antagonists, atropine (9.52 × 10⁻⁶ M), AF-DX 116 (2.72 × 10⁻⁵ M) and pirenzepine (8.24 × 10⁻⁴ M) were necessary to block contralateral suppression of DPOAEs. The in vivo pharmacology of this putative outer hair cell cholinergic receptor suggests that it may be a member of the nicotinic family of receptors.
Characterization of muscarinic binding sites in the adult and developing rat cochlea
1993, Neurochemistry International
The maturation of the cholinergic innervation of the rat cochlea is associated with a transient increase in the muscarinic-receptor activated inositol phosphate synthesis. In order to investigate the mechanisms involved in this transient enhancement of the inositol phosphate response, the binding properties of the cochlear muscarinic receptors were studied during rat cochlear development. Incubating the membranes from 4-day-old, 12-day-old and adult cochleas with [³H]quinuclidinyl benzylate indicates that their respective, mean concentrations of cholinoceptors are 454 ± 51 (± SEM), 39 ± 2 and 42 ± 3 fmol/mg of protein. The dissociation constants at equilibrium are 207 ± 80, 42 ± 7 and 28 ± 3 pM for the binding sites of the 4-day-old, 12-day-old and adult cochleas, respectively. Pharmacological characterization of the binding, using selective antagonists, shows that M3 cholinoceptors are expressed in developing and adult cochleas. The data demonstrate that changes in muscarinic receptor affinity and number do not correlate with the previously observed peak of the inositol phosphate metabolism. The transient enhanced inositol phosphate response is therefore not due to changes in cholinoceptors, but probably due to alterations involving the intrinsic activity of the phospholipase C and/or the efficacy of coupling of the transduction system.
Localisation of functional muscarinic receptors in the rat cochlea: evidence for efferent presynaptic autoreceptors
1993, Brain Research
In the rat cochlea, the activation of muscarinic receptors stimulates the hydrolysis of phosphoinositides but the importance of this muscarinic effect is still unknown. In order to find out about the role of the muscarinic receptors in the cochlea, we examined their functional distribution within this organ. This was achieved by measuring the formation of [³H]inositol phosphates induced by carbachol (1 mM) in two regions of the cochlea: the modiolus and the organ of Corti. At both sites, carbachol enhanced the accumulation of inositol phosphates in an atropine-sensitive way. These stimulations were completely antagonised by 4-diphenylacetoxy-N-methyl piperidine methiodide (1 μM) but unchaged by pirenzepine (1 μM). In cochleas depleted of outer hair cells by a treatment with amikacin, the carbachol-induced formation of inositol phosphates is not altered with respect to control, undamaged cochleas. Conversely, when the medial cholinergic axons which form synapses with the outer hair cells are destroyed by the section of the crossed olivocochlaer bundle the carbachol-stimulated inositol phosphates response is reduced by 35% in the organ of Corti. This section has no effect in the modiolus, despite the degeneration of some modiolar fibers. Our results show that functional muscarinic receptors are distributed both in the organ of Corti and in the modiolus. These two structures contain presumably the same class of cholinoceptor. The effects of selective destruction clearly demonstrate that a population of muscarinic receptors is located on presynaptic membranes at the level of the medial axon-outer hair cell contacts. They also point to spiral ganglion neurons and/or the Schwann cells as sites for the functional cholinoceptors in the modiolus.

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Cholinergic muscarinic receptors in rat cochlea

Abstract

Brain Research

Trends Pharmacol. Sci.

Eur. J. Pharmacol.

Co-localization of enkephaline-like and choline acetyl-transferase like immunoreactivityes in olivocochlear neurons of the guinea-pig

J. Histochem. Cytochem.

Efferent neurotransmitters

Muscarinic receptor heterogeneity

Trends Pharmacol. Sci.

Action of cholinergic and anticholinergic drugs at the cross olivocochlear bundle: hair cell function

Acta Otolaryngol.

Activities of enzymes of the cholinergic system in guinea pig cochlea

J. Histochem. Cytochem.

Differential olivocochlear projection from the lateral versus the medial zones of the superior olivary complex

J. Comp. Neurol.

Rodrigues de Miranda^J.F.