Synaptic development: insights from Drosophila

https://doi.org/10.1016/j.conb.2007.01.001Get rights and content

In Drosophila, the larval neuromuscular junction is particularly tractable for studying how synapses develop and function. In contrast to vertebrate central synapses, each presynaptic motor neuron and postsynaptic muscle cell is unique and identifiable, and the wiring circuit is invariant. Thus, the full power of Drosophila genetics can be brought to bear on a single, reproducibly identifiable, synaptic terminal. Each individual neuromuscular junction encompasses hundreds of synaptic neurotransmitter release sites housed in a chain of synaptic boutons. Recent advances have increased our understanding of the mechanisms that shape the development of both individual synapses — that is, the transmitter release sites including active zones and their apposed glutamate receptor clusters — and the whole synaptic terminal that connects a pre- and post-synaptic cell.

Introduction

The synapse is the primary site of communication between neurons and the fundamental unit of function in the nervous system. Synapse formation involves bidirectional signals between pre- and post-synaptic cells that lead to the development of specialized structures for the release and detection of neurotransmitter. Whereas the basic mechanism of synapse formation is probably genetically determined, the function, maturation and stability of synapses are dynamically regulated during development. Such developmental plasticity not only underlies the refinement of neural circuits, but similar mechanisms might function throughout life to mediate activity-dependent synaptic change. As such, identifying molecular pathways that shape synapse development is a central goal of the developmental neurobiologist. In this review, we describe recent work from the Drosophila neuromuscular junction (NMJ) that provides insights into synaptic development.

The Drosophila NMJ is a favorite model system for studies of the synapse. First, it has the advantage of the power and elegance of modern Drosophila genetics. In addition to the obvious benefits of generating and analyzing mutants, synaptic studies are greatly aided by the ability to control gene function in a temporal and tissue-restricted manner. Such techniques facilitate the precise manipulation of circuits, including the differential regulation of gene function in adjacent target cells of a single motor neuron for the study of synaptic competition.

Second, the NMJ is accessible to various experimental techniques. Electrophysiology, FM (frequency-modulated) dye labeling, Ca2+ imaging and behavioral studies all probe the physiological function of the synapse. Immunohistochemistry, electron microscopy and live imaging provide a clear view of the structural and molecular anatomy of the synapse.

Third, the Drosophila NMJ is glutamatergic. As a result, its molecular constituents — and potentially its developmental mechanisms — resemble vertebrate, central glutamatergic synapses more closely than does the vertebrate cholinergic NMJ.

Fourth, each Drosophila NMJ is unique and identifiable. In each segmental unit of the neuromuscular system, 32 identified motor neurons synapse with 30 identified postsynaptic muscle cells in a stereotyped pattern. Not only are the cells stereotyped, but the arborization pattern and synaptic strength are also roughly stereotyped. Thus, multiple iterations of an identified NMJ can be analyzed in a single fly, and that same NMJ can be reliably compared from fly to fly. This single-synapse resolution enables subtle changes to be observed and characterized when examining mutants.

Last, despite its stereotyped circuitry, the Drosophila NMJ shows robust plasticity. Changes in the environment, neuronal activity and gene function all lead to modification of synaptic structure and function during development. Thus, the Drosophila NMJ combines many of the best features of the simple, genetically tractable Caenorhabditis elegans model with the more complex, physiologically accessible mouse model.

Before describing recent advances, we wish to highlight an important semantic issue. Researchers in the field (including us) have an unfortunate tendency to use the term ‘synapse’ to describe two very different structures. First, the whole synaptic connection formed between a motor neuron and muscle is often referred to as a synapse. Such a structure comprises a branched chain of synaptic boutons formed by the motor neuron and typically surrounded by an elaborate membranous compartment made by the muscle. For an average NMJ, this connection can include 20–50 synaptic boutons. Within each bouton, however, are multiple presynaptic release sites, termed active zones, where synaptic vesicles cluster and fuse. Opposite each active zone, postsynaptic glutamate receptors cluster to sense the released transmitter. This dyad of active zone and glutamate receptor cluster is also called a synapse, and is more akin to the usual definition of vertebrate glutamatergic synapses. Because each bouton contains approximately ten active zones, each motor neuron can form upwards of 500 such synapses with a single postsynaptic cell.

We highlight this semantic point because the development of each structure is probably controlled by very different molecular mechanisms. As such, we address each structure separately in this review. We refer to the chain of boutons formed between motor neuron and muscle as the ‘synaptic terminal’, and reserve the term ‘synapse’ for individual active zones and their apposed receptor cluster.

Section snippets

Active zones at the synapse

For many years, studies of motor neuron morphology at the NMJ have relied on antibodies that recognize neuronal membrane proteins, the neuronal cytoskeleton and/or synaptic vesicle proteins. By using these reagents, numerous mutants have been characterized that alter synaptic terminal morphology. These tools are inadequate, however, for studies of individual synapses at the NMJ. Recently, the identification of the first active-zone protein in Drosophila, Bruchpilot (BRP), along with the

Trans-synaptic signals control synaptic terminal development

The synaptic terminal encompasses all of the synaptic contacts between a motor neuron and postsynaptic muscle cell. Development of the Drosophila NMJ synaptic terminal is regulated by trans-synaptic signals, including Wingless (Wg), a member of the Wnt family, and Glass bottom boat (Gbb), a member of the bone morphogenetic protein (BMP) and transforming growth factor-β (TGFβ) family [17]. At the NMJ, Wg and Gbb act like classical morphogens, stimulating a signal to the nucleus that presumably

Synaptic terminal stability

Synaptic terminal morphology is shaped by both the maintenance and the formation of synaptic contacts. Indeed, many developing neuronal circuits initially form exuberant synaptic contacts that are later refined by selective elimination and stabilization [58]. At the Drosophila larval NMJ, synapse elimination does not regulate connectivity as it does at the vertebrate NMJ. Synaptic boutons can, however, be lost during development. This loss is visualized as a ‘footprint’ — that is, a bouton or

Conclusions

Analysis of synaptic development at the Drosophila NMJ has matured in recent years. The traditional metric for analyzing NMJ development has been to count the number of synaptic boutons. Although this is still an important descriptor of a developing synapse, as a single measure it is inadequate. As our understanding of cell biological and signal transduction mechanisms grows, novel genes that shape synaptic development should be placed within the rich context of developmental mechanisms at work

Update

Three recent papers provide additional insights into the molecular mechanisms underlying synaptic development. Pielage et al. demonstrate that the selective disruption of postsynaptic spectrin leads to abnormally large active zones whose spacing is altered. They propose that a postsynaptic spectrin–actin lattice acts as a scaffold to organize pre- and post-synaptic development [64]. Work from Schmid et al. investigates the role of ionotropic glutamate receptors for synaptic development [65].

References and recommended reading

Papers of particular interest, published within the period of review, have been highlighted as:

  • • of special interest

  • •• of outstanding interest

Acknowledgements

We thank Richard Daniels, Ethan Graf, EJ Brace and Chunlai Wu for helpful comments on this manuscript. CAC is supported by an award from Paralyzed Veterans of America, and AD is supported by awards from the National Institutes of Health (NS051453 and NS043171) and the Keck Foundation.

References (67)

  • N.M. Viquez et al.

    The B′ protein phosphatase 2A regulatory subunit well-rounded regulates synaptic growth and cytoskeletal stability at the Drosophila neuromuscular junction

    J Neurosci

    (2006)
  • C. Ruiz-Canada et al.

    New synaptic bouton formation is disrupted by misregulation of microtubule stability in aPKC mutants

    Neuron

    (2004)
  • B.A. Eaton et al.

    LIM kinase1 controls synaptic stability downstream of the type II BMP receptor

    Neuron

    (2005)
  • J.M. Rawson et al.

    The heparan sulfate proteoglycans Dally-like and Syndecan have distinct functions in axon guidance and visual-system assembly in Drosophila

    Curr Biol

    (2005)
  • J. Ashley et al.

    Fasciclin II signals new synapse formation through amyloid precursor protein and the scaffolding protein dX11/Mint

    J Neurosci

    (2005)
  • B. Marie et al.

    Dap160/intersectin scaffolds the periactive zone to achieve high-fidelity endocytosis and normal synaptic growth

    Neuron

    (2004)
  • D.K. Dickman et al.

    Altered synaptic development and active zone spacing in endocytosis mutants

    Curr Biol

    (2006)
  • H.I. Wan et al.

    Highwire regulates synaptic growth in Drosophila

    Neuron

    (2000)
  • J. D'Souza et al.

    Formation of the retinotectal projection requires Esrom, an ortholog of PAM (protein associated with Myc)

    Development

    (2005)
  • A. DiAntonio et al.

    Ubiquitination-dependent mechanisms regulate synaptic growth and function

    Nature

    (2001)
  • B.D. McCabe et al.

    Highwire regulates presynaptic BMP signaling essential for synaptic growth

    Neuron

    (2004)
  • C.A. Collins et al.

    Highwire restrains synaptic growth by attenuating a MAP kinase signal

    Neuron

    (2006)
  • G.N. Patrick

    Synapse formation and plasticity: recent insights from the perspective of the ubiquitin proteasome system

    Curr Opin Neurobiol

    (2006)
  • F. Zalfa et al.

    mRNPs, polysomes or granules: FMRP in neuronal protein synthesis

    Curr Opin Neurobiol

    (2006)
  • Y. Goda et al.

    Mechanisms of synapse assembly and disassembly

    Neuron

    (2003)
  • J. Pielage et al.

    Presynaptic spectrin is essential for synapse stabilization

    Curr Biol

    (2005)
  • S. Hebbar et al.

    Pruning of motor neuron branches establishes the DLM innervation pattern in Drosophila

    J Neurobiol

    (2004)
  • S. Sanyal et al.

    AP-1 functions upstream of CREB to control synaptic plasticity in Drosophila

    Nature

    (2002)
  • F. Hannan et al.

    Second messenger systems underlying plasticity at the neuromuscular junction

    Int Rev Neurobiol

    (1999)
  • A. Schmid et al.

    Non-NMDA-type glutamate receptors are essential for maturation but not for initial assembly of synapses at Drosophila neuromuscular junctions

    J Neurosci

    (2006)
  • The Fly Neuromuscular Junction: Structure and Function, Second Edition. Edited by Budnik V and Ruiz-Cañada C. Academic...
  • R.J. Kittel et al.

    Bruchpilot promotes active zone assembly, Ca2+ channel clustering, and vesicle release

    Science

    (2006)
  • D.A. Wagh et al.

    Bruchpilot, a protein with homology to ELKS/CAST, is required for structural integrity and function of synaptic active zones in Drosophila

    Neuron

    (2006)
  • Cited by (203)

    View all citing articles on Scopus
    View full text