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Mouse muscle denervation increases expression of an alpha7 nicotinic receptor with unusual pharmacology
Hiroshi Tsuneki1, Ramiro Salas, John A Dani
1Division of Neuroscience, Baylor College of Medicine, One Baylor Plaza, Houston, TX 77030-3498, USA.
Abstract:
Neuronal nicotinic alpha7 subunits have been found in chick and rat skeletal muscle during development and denervation. In the present study, reverse transcriptase-polymerase chain reaction was used to detect alpha7 subunit mRNA in denervated mouse muscle. To determine whether the alpha7 subunit forms functional nicotinic acetylcholine receptors (nAChRs) in muscle, choline was used to induce a membrane depolarization because choline has been considered a specific agonist of alpha7-containing (alpha7*) nAChRs. We found, however, that choline (3-10 mM) also weakly activates muscle nAChRs. After inhibiting muscle nAChRs with a specific muscle nAChR inhibitor, alpha-conotoxin GI (alphaCTxGI), choline was used to activate the alpha7* nAChRs on muscle selectively. Four weeks after denervation, rapid application of choline (10 mM) elicited a substantial depolarization in the presence of alphaCTxGI (0.1 microM). This component of the depolarization was never present in denervated muscles obtained from mutant mice lacking the alpha7 subunit (i.e. alpha7-null mice). The depolarization component that is resistant to alphaCTxGI was antagonized by pancuronium (3-10 microM) and by a 4-oxystilbene derivative (F3, 0.1-0.5 microM) at concentrations considered highly specific for alpha7* nAChRs. Another selective alpha7 antagonist, methyllycaconitine (0.05-5 microM), did not strongly inhibit this choline-induced depolarization. Furthermore, the choline-sensitive nAChRs showed little desensitization over 10 s of application with choline (10-30 mM). These results indicate that functional alpha7* nAChRs are significantly present on denervated muscle, and that these receptors display unusual functional and pharmacological characteristics.
Insights
Functional neuronal nicotinic alpha7 receptors are present in denervated mouse muscle. These alpha7-containing receptors exhibit unique characteristics, responding to choline when muscle receptors are blocked.
Area of Science:
- Neuroscience
- Molecular Biology
- Muscle Physiology
Background:
- Neuronal nicotinic alpha7 subunits are found in developing and denervated chick and rat skeletal muscle.
- The presence and function of alpha7-containing nicotinic acetylcholine receptors (nAChRs) in denervated mouse muscle remain to be fully elucidated.
Purpose of the Study:
- To detect alpha7 subunit mRNA in denervated mouse muscle using reverse transcriptase-polymerase chain reaction.
- To determine if alpha7 subunits form functional nAChRs in denervated muscle by assessing choline-induced depolarization.
- To characterize the pharmacological and functional properties of these potential alpha7* nAChRs.
Main Methods:
- Reverse transcriptase-polymerase chain reaction (RT-PCR) to detect alpha7 subunit mRNA.
- Electrophysiological recordings of membrane depolarization in response to choline application.
- Selective blockade of muscle nAChRs using alpha-conotoxin GI (alphaCTxGI).
- Pharmacological characterization using specific antagonists like pancuronium and F3, and methyllycaconitine.
Main Results:
- Alpha7 subunit mRNA was detected in denervated mouse muscle.
- Choline induced a depolarization in denervated muscle, even after blocking muscle nAChRs with alphaCTxGI.
- This alphaCTxGI-resistant depolarization was absent in alpha7-null mice, confirming its dependence on alpha7 subunits.
- The choline-activated receptors were antagonized by pancuronium and F3, but not strongly by methyllycaconitine.
- These alpha7*-containing nAChRs showed minimal desensitization during prolonged choline application.
Conclusions:
- Functional alpha7-containing nAChRs are significantly present in denervated mouse skeletal muscle.
- These muscle-expressed alpha7* nAChRs possess distinct pharmacological and functional properties compared to other nAChRs.
- The findings suggest a potential role for alpha7* nAChRs in muscle physiology, particularly under denervation conditions.