Related Experiment Videos
Regulation of muscle AChR alpha subunit gene expression by electrical activity: involvement of protein kinase C and
A Klarsfeld1, R Laufer, B Fontaine
1U. R. A. CNRS 0210, Département des Biotechnologies, Institut Pasteur, Paris, France.
Abstract:
Using primary cultures of chicken myotubes, we investigated the involvement of protein kinase C and Ca2+ in the repression of nicotinic acetylcholine receptor (AChR) biosynthesis by electrical activity. Treatment with the Ca2+ channel blocker verapamil or the Na+ channel blocker tetrodotoxin increased alpha subunit mRNA levels 11.5- to 15-fold. The effect of tetrodotoxin was abolished in the presence of the Ca2+ ionophore A23187. Dantrolene, which blocks Ca2+ efflux from the sarcoplasmic reticulum, caused only a 1.7-fold increase in alpha subunit mRNA levels. Down regulation of protein kinase C by prolonged exposure to the phorbol ester TPA or inhibition of protein kinase C by staurosporine led to 8- to 10-fold increases in alpha subunit mRNA levels. Mature and precursor forms of AChR alpha subunit mRNA were found to vary in parallel throughout all of these treatments, suggesting that protein kinase C and Ca2+ ions may modulate AChR alpha subunit biosynthesis at the transcriptional level.
Insights
Electrical activity represses nicotinic acetylcholine receptor (AChR) biosynthesis. Protein kinase C and Ca2+ ions are involved, with their inhibition increasing AChR alpha subunit mRNA levels, suggesting transcriptional regulation.
Area of Science:
- Neuroscience
- Molecular Biology
- Cellular Physiology
Background:
- Electrical activity influences muscle cell function.
- Nicotinic acetylcholine receptors (AChRs) are crucial for neuromuscular transmission.
- Regulation of AChR biosynthesis is vital for muscle development and function.
Purpose of the Study:
- To investigate the roles of protein kinase C (PKC) and calcium ions (Ca2+) in electrical activity-mediated repression of AChR biosynthesis.
- To determine the molecular mechanisms underlying AChR regulation by electrical stimulation.
Main Methods:
- Primary chicken myotube cultures were used.
- Pharmacological agents including verapamil (Ca2+ channel blocker), tetrodotoxin (Na+ channel blocker), A23187 (Ca2+ ionophore), and dantrolene (sarcoplasmic reticulum Ca2+ efflux blocker) were applied.
- Protein kinase C was modulated using TPA (phorbol ester) and staurosporine (inhibitor).
- Alpha subunit mRNA levels for AChRs were quantified.
Main Results:
- Verapamil and tetrodotoxin significantly increased AChR alpha subunit mRNA levels (11.5- to 15-fold).
- Tetrodotoxin's effect was reversed by A23187, indicating Ca2+ involvement.
- Dantrolene showed a modest increase (1.7-fold) in mRNA levels.
- PKC downregulation or inhibition led to substantial increases in AChR alpha subunit mRNA (8- to 10-fold).
- Mature and precursor AChR alpha subunit mRNA levels changed in parallel.
Conclusions:
- Protein kinase C and Ca2+ ions play a significant role in repressing nicotinic acetylcholine receptor biosynthesis.
- These signaling pathways likely modulate AChR alpha subunit biosynthesis at the transcriptional level.
- Understanding these mechanisms is key to comprehending neuromuscular junction regulation.