Related Experiment Videos
The desensitization of the embryonic mouse muscle acetylcholine receptor depends on the cellular environment
1Laboratorio di Biofisica, Centro Ricerca Sperimentale I.R.E., Via delle Messi d’ Oro 156, I-00158 Rome, Italy.
Abstract:
The rate of desensitization of nicotinic acetylcholine (ACh) receptor (nAChR), an important characteristic of nAChR function, was studied in myotubes of the mouse C2C12 cell line at different times after fusion, by measuring the decay of ACh-evoked currents (IACh) under various patch-clamp configurations. We observed a progressive slowing of IACh decay rate (half-decay time rose from about 0.5 s to over 5 s) in myotubes of increasing size (i.e. age) under all experimental conditions, except in outside-out patches, when IACh decayed as fast as in the smallest myotubes. Single-channel conductance (about 35 pS) and open time (about 3.5 ms), measured in outside-out and cell-attached patches, were independent of myotube size. In Xenopus oocytes injected with poly(A+)RNA extracted from C2C12 myoblasts or mature myotubes, IACh decay was about 50 times slower than in myotubes. Neither cAMP-dependent nor diacylglycerol-dependent protein kinases, actin nor microtubule polymerization state influenced IACh decay. Our data indicate that the cellular environment, but not readily dialysable cytosolic factors, markedly influences the functional behaviour of nAChR.
Insights
The desensitization rate of nicotinic acetylcholine receptors (nAChR) slows with myotube age. The cellular environment, not cytosolic factors, significantly impacts nAChR function.
Area of Science:
- Neuroscience
- Cell Biology
- Biophysics
Background:
- Nicotinic acetylcholine receptors (nAChR) are crucial for neuromuscular transmission.
- nAChR desensitization is a key aspect of receptor function.
- Understanding nAChR desensitization in developing muscle cells is important.
Purpose of the Study:
- To investigate the rate of desensitization of nAChR in mouse C2C12 myotubes.
- To determine how myotube age and cellular environment affect nAChR desensitization.
- To identify factors influencing nAChR functional behavior.
Main Methods:
- Patch-clamp electrophysiology was used to measure acetylcholine (ACh)-evoked currents (IACh) in C2C12 myotubes.
- Experiments were conducted in various patch-clamp configurations (outside-out, cell-attached).
- Single-channel properties and IACh decay kinetics were analyzed.
Main Results:
- IACh decay rate progressively slowed with increasing myotube size (age), with half-decay times increasing from ~0.5s to over 5s.
- This age-dependent slowing was absent in outside-out patches, indicating a role for the cellular environment.
- Single-channel conductance and open time were independent of myotube size.
- IACh decay was significantly slower in Xenopus oocytes expressing C2C12 RNA compared to myotubes.
- Protein kinases and cytoskeletal components did not influence IACh decay.
Conclusions:
- The cellular environment, rather than readily dialyzable cytosolic factors, significantly influences nAChR desensitization kinetics.
- nAChR function is modulated by the muscle cell's developmental stage and its surrounding milieu.
- These findings provide insights into the regulation of neuromuscular transmission during muscle development.