Identification of acetylcholine receptor subunits differentially expressed in singly and multiply innervated fibers of extraocular muscles.
OBJECTIVE
To identify the acetylcholine receptor (AChR) isoforms among the neuromuscular junctions (NMJs) of singly and multiply innervated fibers (SIFs and MIFs) of rat extraocular muscles (EOMs).


METHODS
EOMs were dissected from adult rats and serially sectioned. Sections were simultaneously stained for acetylcholinesterase and with an antibody to the slow myosin heavy chain to identify NMJ topography and fiber types in the same section. Synapses and subsynaptic regions of SIFs and MIFs were isolated by laser capture microdissection and the AChR subunits identified by RT-PCR.


RESULTS
The en plaque endings of SIFs expressed only the adult epsilon subunit, not the fetal gamma subunit, of the AChR, whereas the en grappe endings of the MIFs expressed only the gamma subunit, and not the epsilon subunit. Although the expression of the epsilon subunit was confined to the NMJ region of the SIFs, the gamma subunit was expressed both synaptically and extrasynaptically within the MIFs. The gamma subunit in MIFs correlated with the expression of the myogenic regulatory factor myogenin. Moreover, an unusual neuronal AChR subunit, alpha9, was found in the EOMs, but not in the limb muscles.


CONCLUSIONS
The adult epsilon and fetal gamma subunits of the AChRs are segregated into distinct synapses on distinct fiber types. The maintenance of the fetal subunit in a population of fibers is probably linked to the expression of myogenin and is a unique attribute of the EOM allotype.
Fraterman
Sven
S
Department of Cell and Developmental Biology, University of Pennsylvania School of Medicine, PA 19104, USA.
Khurana
Tejvir S
TS
Rubinstein
Neal A
NA
D000818:Animals; D007150:Immunohistochemistry; D018485:Muscle Fibers, Skeletal / Q000378:metabolism*; D018008:Myogenin / Q000378:metabolism; D009469:Neuromuscular Junction / Q000378:metabolism*; D009801:Oculomotor Muscles / Q000294:innervation*; D009882:Ophthalmic Nerve / Q000378:metabolism*; D020033:Protein Isoforms / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D011950:Receptors, Cholinergic / Q000235:genetics / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction
C503803:Myog protein, rat; D018008:Myogenin; D020033:Protein Isoforms; D012333:RNA, Messenger; D011950:Receptors, Cholinergic