Molecular mechanism of active zone organization at vertebrate neuromuscular junctions

Abstract

Organization of presynaptic active zones is essential for development, plasticity, and pathology of the nervous system. Recent studies indicate a trans-synaptic molecular mechanism that organizes the active zones by connecting the pre- and the postsynaptic specialization. The presynaptic component of this trans-synaptic mechanism is comprised of cytosolic active zone proteins bound to the cytosolic domains of voltage-dependent calcium channels (P/Q-, N-, and L-type) on the presynaptic membrane. The postsynaptic component of this mechanism is the synapse organizer (laminin β2) that is expressed by the postsynaptic cell and accumulates specifically on top of the postsynaptic specialization. The pre- and the postsynaptic components interact directly between the extracellular domains of calcium channels and laminin β2 to anchor the presynaptic protein complex in front of the postsynaptic specialization. Hence, the presynaptic calcium channel functions as a scaffolding protein for active zone organization and as an ion-conducting channel for synaptic transmission. In contrast to the requirement of calcium influx for synaptic transmission, the formation of the active zone does not require the calcium influx through the calcium channels. Importantly, the active zones of adult synapses are not stable structures and require maintenance for their integrity. Furthermore, aging or diseases of the central and peripheral nervous system impair the active zones. This review will focus on the molecular mechanisms that organize the presynaptic active zones and summarize recent findings at the neuromuscular junctions and other synapses.

Fig. 1

(a) A schematic diagram of the vertebrate neuromuscular junction (left) and a close-up of one active zone area (right). Solid arrows represent interactions, and the dotted arrow indicates a functional link. Horizontal lines show pre- and post-synaptic membranes. A junctional fold is indicated as a trough on the postsynaptic side. The space between these lines represents the synaptic cleft. The gray triangle depicts the electron dense material of presynaptic active zones detected by electron microscopy. The size of the proteins and the synaptic cleft are not in scale. VDCC voltage-dependent calcium channels. (b) The active zone protein Bassoon forms puncta and labels the active zones in NMJs of adult mice. (c) The synaptic vesicle associated protein synapsin1 shows a diffuse distribution pattern that visualizes the nerve morphology. Note the difference in the distribution patterns of these two presynaptic proteins. Bassoon, synapsin1, and acetylcholine receptors (AChRs) were visualized at the NMJs of sternomastoid muscles of postnatal day 36 mice by fluorescent immunohistochemistry using anti-Bassoon antibodies (green in b), anti-synapsin1 antibodies (green in c), and Alexa Fluor 594-labeled α-bungarotoxin (red). Many Bassoon puncta are localized above the postsynaptic junctional folds, which were visualized as bright lines of α-bungarotoxin staining. Scale bar 1βm.