Rodent nerve-muscle cell culture system for studies of neuromuscular junction development: Refinements and applications

Mathew P. Daniels, Brian T. Lowe, Sanjiv Shah, Jianxin Ma, Steven J. Samuelsson, Brian Lugo, Tehnaz Parakh, Chang Sub Uhm

Research output: Contribution to journalArticle

36 Citations (Scopus)

Abstract

Understanding of vertebrate neuromuscular junction (NMJ) development has been advanced by experimentation with cultures of dissociated embryonic nerve and skeletal muscle cells, particularly those derived from Xenopus and chick embryos. We previously developed a rodent (rat) nerve-muscle coculture system that is characterized by extensive induction of acetylcholine receptor (AChR) aggregation at sites of axonal contact with myotubes (Dutton et al., 1995). In this article, we report modifications of this culture system and examples of its application to the study of NMJ development: (1) We describe improved methods for the enrichment of myoblasts to give higher yields of myotubes with equal or greater purity. (2) We demonstrate lipophilic dye labelling of axons in cocultures by injection of dye into neuron aggregates and show the feasibility of studying the growth of living axons on myotubes during synapse formation. (3) We describe the preparation of a better-defined coculture system containing myotubes with purified rat motoneurons and characterize the system with respect to axon-induced AChR aggregation. (4) We demonstrate dependence of the pattern of axon-induced AChR aggregation on muscle cell species, by the use of chick-rat chimeric co-cultures. (5) We provide evidence for the role of alternatively-spliced agrin isoforms in synapse formation by using single cell RT-PCR with neurons collected from co-cultures after observation of axon-induced AChR aggregation.

Original languageEnglish
Pages (from-to)26-37
Number of pages12
JournalMicroscopy Research and Technique
Volume49
Issue number1
DOIs
Publication statusPublished - 2000 Apr 1

Fingerprint

axons
muscle cells
rodents
Receptor Aggregation
Neuromuscular Junction
acetylcholine
nerves
Coculture Techniques
Cell culture
Muscle Cells
Skeletal Muscle Fibers
Axons
Muscle
Cholinergic Receptors
Rodentia
Cell Culture Techniques
Neurons
Agglomeration
rats
Rats

Keywords

  • Acetylcholine receptor
  • Agrin
  • Axon
  • Growth cone
  • Motoneuron
  • Myotube
  • Rat
  • Synapse

ASJC Scopus subject areas

  • Anatomy
  • Histology
  • Instrumentation
  • Medical Laboratory Technology

Cite this

Rodent nerve-muscle cell culture system for studies of neuromuscular junction development : Refinements and applications. / Daniels, Mathew P.; Lowe, Brian T.; Shah, Sanjiv; Ma, Jianxin; Samuelsson, Steven J.; Lugo, Brian; Parakh, Tehnaz; Uhm, Chang Sub.

In: Microscopy Research and Technique, Vol. 49, No. 1, 01.04.2000, p. 26-37.

Research output: Contribution to journalArticle

Daniels, Mathew P. ; Lowe, Brian T. ; Shah, Sanjiv ; Ma, Jianxin ; Samuelsson, Steven J. ; Lugo, Brian ; Parakh, Tehnaz ; Uhm, Chang Sub. / Rodent nerve-muscle cell culture system for studies of neuromuscular junction development : Refinements and applications. In: Microscopy Research and Technique. 2000 ; Vol. 49, No. 1. pp. 26-37.
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