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Introduction to Organization of the Neuromuscular Junction:
From Structure to Function
Biologie Cellulaire des Membranes, Institut Jacques Monod, CNRS, UMR 7592, Universités Paris 6/7, 75251 Paris-Cedex 05,
Progress in experimental science depends greatly on
the tools of study that are available. The deciphering of
the relationship between the structure and function of
the neuromuscular junction (NMJ) offers a typical example. Almost a century ago, Ramon y Cajal and his
coworkers applied silver impregnation techniques to
neuromuscular preparations, techniques first developed for the study of neurons (Ramon y Cajal, 1928).
This pioneering work set the basis for the concept of a
discontinuity between neuronal and muscular cytoplasms at the neuromuscular synapse.
Increasingly elaborate staining techniques were subsequently developed and used in conjunction with enzyme histochemistry and electrophysiology to unravel
the structural bases of chemical synaptic transmission
at the vertebrate NMJ. However, elucidation of the fine
structure of the NMJ had to await the development of
electron microscopy (EM) in the1950s. The higher resolution and rapid evolution of the various EM-associated techniques made it possible to establish the definite existence of a gap between the neuronal and muscular plasma membranes: the synaptic cleft. This
structure comprises a specialized basal lamina where
trophic factors, such as agrin and heregulin, involved
in synaptogenesis and the enzyme acetylcholinesterase
accumulate. Moreover, EM analysis revealed the two
fundamental elements of the synaptic structure: the
pre- and postsynaptic apparati (reviewed in Couteaux,
1973), which represent local differentiations of the motor nerve terminal and the sarcolemma, respectively.
These two domains are characterized by the local accumulation of specialized organelles (presynaptic vesicles, postsynaptic golgi apparatus, fundamental nuclei,
etc.) and membrane proteins (ion channels, cell adhesion molecules, cytoskeletal proteins). On the presynaptic side, the active zones contain the specialized molecular machinery for synaptic vesicle docking and
neurotransmitter release that is responsible for the
regulated secretion of acetylcholine by the nerve terminal. In front of the active zones of the nerve endings,
the crests of the postsynaptic membrane folds exhibit
extraordinarily high concentrations of a ligand-gated
Na⫹/K⫹ channel, the nicotinic acetylcholine receptor
(AChR), whereas voltage-sensitive Na ⫹ channels
(NaChs) accumulate in the depths of the postsynaptic
folds, in regions immediately adjacent to the AChRs.
These two channels are responsible for the initiation of
membrane depolarization and the generation of the
endplate potential. A number of integral and cytoskeletal proteins accumulating at the postsynaptic membrane are believed to participate in the clustering and
maintenance of AChRs and NaChs.
Synaptic transmission is a complex process requiring
a high degree of coordination between pre- and
postsynaptic structures. The first step in the study of
this process aimed at unraveling the fine structure of
the mature NMJ. The current challenge is to understand how such specializations develop and communicate at the molecular level. At this point, multiple
experimental approaches are required. This topical issue presents recent advances in the study of the morphogenesis and functioning of the NMJ obtained by the
combination of EM analysis and subcellular localization of synaptic proteins and transcripts, molecular
biology, electrophysiology, genetics, and the use of several animal models ranging from Drosophila to mammals.
Three articles in this issue deal with the morphogenesis of the NMJ. The first event in synaptogenesis
is the synaptic target recognition. Rose and Chiba et
al. (2000) present a summary of the current state of
knowledge on the chemoaffinity hypothesis and the
role of cell adhesion molecules in the initial recognition between neuronal processes and the sarcolemma
leading to the future Drosophila NMJ. The relatively
simple, well-defined pattern of synapses, the possibility of in vivo analysis, and the powerful molecular
genetics make Drosophila a choice tool for the study
of morphogenesis of the NMJ. Y. H. Koh et al. (2000)
take advantage of the Drosophila NMJ model to
study the molecular cascades and the role of synaptic
signaling molecules, such as MAGUKs, involved in
synaptic plasticity. The study of synaptogenesis in
vertebrates is confronted with a high degree of tissue
complexity. The current approach makes use of homologous recombination to generate mutant mice
that lack the molecules involved in the synaptogenesis process (reviewed in Sanes and Lichtman, 1999).
An alternative approach is the development of cellular model systems involving neuron-muscle cocultures to study in vitro synaptogenesis. M. Daniels et
al. (2000) describe the establishment and characterization of a rodent neuron-muscle coculture, and provide examples of its application to the study of mammalian NMJ development.
According to the vesicle hypothesis, quantal release
of acetylcholine (ACh) from the synaptic vesicles into
the synaptic cleft occurs upon fusion with the presynaptic membrane. This hypothesis has now become a
wide consensus. However, a number of data do not fit
into this model, instead they suggest that vesicle fusion
This Topical Issue is dedicated to the memory of Professor René Couteaux,
1909 –1999.
and quantal ACh release might be two linked-but-distinct processes. Morel and M. Isräel (2000) propose an
alternative model involving direct release of ACh
through a membrane pore. Ultrastructural evidence in
favor of this model is presented by Dunant (2000) who
makes use of freeze-fracture EM to study rapid
changes of presynaptic intramembrane particles with
regard to ACh release and vesicle fusion. Both studies
use the Torpedo electric organ, a source of abundant
cholinergic and functionally active synaptosomes.
A key enzyme in the control of cholinergic synaptic
transmission at the NMJ is the acetylcholinesterase
(AChE). Legay (2000) discusses recent data on the molecular structure and polymorphism, transcriptional
regulation and synaptic localization mechanisms, and
the implification of AChE in myasthenic syndromes.
The morphogenesis and maintenance of the postsynaptic membrane domain is one of the most intensively
studied aspects in the NMJ field. A major question
addresses the mechanisms of specific local accumulation and clustering of postsynaptic membrane molecules. Cartaud et al. (2000) discuss the Torpedo electrocyte as a model system for the study of the supramolecular organization and clustering of AChRs, and
membrane-cytoskeleton interactions, including the
dystrophin complex and rapsyn, as one of the mechanisms responsible for AChR aggregation at the
postsynaptic membrane. Caldwell (2000) discusses the
current state of knowledge on the role of cytoskeletal
and extracellular molecules in NaCh clustering at the
troughs of the mammalian postsynaptic folds.
Transcriptional regulation is another mechanism responsible for the local accumulation of synaptic proteins. RNAs encoding several synaptic integral and
cytoskeletal proteins accumulate at the NMJ (reviewed
in Sanes and Lichtman, 1999). Gramolini et al. (2000)
present recent data on the compartmentalized expression and postsynaptic localization of the dystrophinrelated protein utrophin in skeletal muscle fibers.
These studies also provide the basis for the development of a potential therapeutic strategy for Duchenne
muscular dystrophy aimed at replacing dystrophin by
extrasynaptically overexpressed utrophin. Finally, in
my contribution, (Kordeli, 2000) I discuss the current
state of knowledge of the spectrin-based skeleton that
coexists with the dystrophin-associated protein complex at the postsynaptic membrane and is thought to
participate in the organization of the postsynaptic folds
and the clustering of synaptic proteins.
Caldwell JH. 2000. Clustering of sodium channels at the neurocuscular junction. Microsc Res Tech 49:84 – 89.
Cartaud J, Cartaud, Annie A, Kordeli, Ekaternini E, Ludosky MA,
Marchard S, Stetzkowski-Marden F. 2000. Torpedo electrocyte: a
model system to study membrane-cytoskeleton interactions at the
postsynaptic membrane. Microsc Res Tech 49:73– 83.
Couteaux R. 1973. Motor endplate structure. In Bourne GH, editor.
Structure and function of muscle, vol 2. New York: Academic Press,
p. 483–530.
Daniels MP, Lowe BT, Shah S, Ma J, Samuelsson SJ, Steven J, Lugo
B, Parakh T, Uhm C-S. 2000. Rodent nerve muscle cell culture
system for studies of neuromuscular junction development: refinements and applications. Microsc Res Tech 49:26 –37.
Demian R, Chiba A. 2000. Synaptic target recognition at Drosophila
neuromuscular junctions. Microsc Res Tech 49:3–13.
Dunant Y. 2000. Quantal acetylcholine release: vesicle fusion or intramembrane particles? Microsc Res Tech 49:38 – 46.
Gramolini A, Wu J, Jasmin BJ. 2000. Regulation and functional
significance of utrophin expression at the mammalian neuromuscular synapse. Microsc Res Tech 49:90 –100.
Koh YH, Gramates LS, Budnik V. 2000. Drosophila larval euromuscular junction: molecular components and mechanisms underlying
synaptic plasticity. Microsc Res Tech 49:14 –25.
Kordeli E. 2000. Spectrin-based skeleton at the postsynaptic membrane of the neuromuscular junction. Microsc Res Tech 49:101–108.
Legay C. 2000. Why so many forms of acetylcholinesterase?. Microsc
Res Tech 49:56 –72.
Morel N, Israël M. 2000. Role of mediatophore in connection with
proteins of the active zone in synaptic transmission. Microsc Res
Tech 49:47–55.
Ramon y Cajal S. 1928. (Reprinted 1991). Degeneration and regeneration of the nervous system. London: Oxford University Press.
Sanes JR, Lichtman JW. 1999. Development of the vertebrate neuromuscular junction. Annu Rev Neurosci 22:389 – 442.
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