In Vivo Calcium Signaling during Synaptic Refinement at the Drosophila Neuromuscular Junction

Area of Science:

• Developmental Neuroscience
• Molecular Neurobiology
• Synaptic Plasticity

Background:

• Neural activity is crucial for refining synaptic connections during development.
• Low-frequency calcium (Ca2+) oscillations are implicated in synaptic pruning in various systems.
• Molecular mechanisms underlying activity-dependent synaptic refinement remain incompletely understood.

Purpose of the Study:

• To investigate the role of Ca2+ oscillations in activity-dependent synaptic pruning at Drosophila neuromuscular junctions (NMJs).
• To elucidate the molecular signaling network involved in growth cone filopodia exploration and retraction.
• To understand how neurons respond to muscle-derived chemorepellents like Sema2a during synapse formation.

Main Methods:

• In vivo imaging of Drosophila embryonic growth cone filopodia.
• Analysis of episodic and oscillatory Ca2+ signals in motoneuron filopodia and synaptic boutons.
• Genetic manipulation to assess the roles of phosphatases (calcineurin, PP1) and kinases (CaMKII, PKA) in synaptic refinement.

Main Results:

• Motoneuron filopodia repeatedly contacted off-target muscle fibers before retracting, with Ca2+ signals observed in both motile and stabilized contacts.
• Ca2+ transients matured into regular low-frequency (0.03 Hz) oscillations during late embryogenesis.
• Genetic evidence points to a presynaptic Ca2+-dependent signaling network involving calcineurin, PP1, CaMKII, and PKA, which modulates the response to Sema2a.

Conclusions:

• Oscillatory Ca2+ signals are a key component of the molecular pathway underlying synaptic network refinement in Drosophila.
• A complex presynaptic signaling network involving kinases and phosphatases orchestrates the removal of off-target contacts in response to muscle chemorepellents.
• This study provides insights into the activity-dependent mechanisms establishing precise synaptic connectivity during development.