Postsynaptic Calcium Extrusion at the Mouse Neuromuscular Junction Alkalinizes the Synaptic Cleft

Area of Science:

• Neuroscience
• Physiology

Background:

• Extracellular pH significantly influences neurotransmission by affecting transmitter release and receptor activity.
• Synaptic cleft pH can either acidify or alkalize due to vesicular release or ion transport mechanisms.
• The pH dynamics at the mammalian neuromuscular junction (NMJ) under physiological conditions are not well understood.

Purpose of the Study:

• To investigate extracellular pH transients in the synaptic cleft of the mammalian NMJ.
• To identify the sources and mechanisms underlying these pH changes during neurotransmission.

Main Methods:

• Utilized viral expression of the pH-sensitive probe pHusion-Ex in mouse muscle to monitor synaptic cleft pH.
• Employed GCaMP3 to track postsynaptic intracellular calcium (Ca²⁺) dynamics.
• Manipulated nerve stimulation parameters (frequency and duration) and inhibited the plasma membrane Ca²⁺ ATPase (PMCA).

Main Results:

• Brief nerve stimulation (5s at 50 Hz) induced significant and prolonged alkalization, dependent on postsynaptic Ca²⁺ release.
• Sustained stimulation (20s at 50 Hz) resulted in a prolonged net acidification of the synaptic cleft.
• Postsynaptic Ca²⁺ liberation positively correlated with cleft alkalization, and PMCA inhibition reduced this alkalization.

Conclusions:

• Cholinergic synapses at the mouse NMJ typically alkalize due to activity-induced release of intracellular Ca²⁺.
• Under strenuous stimulation, the NMJ exhibits net acidification.
• These bidirectional pH changes are use-dependent and provide new insights into synaptic plasticity and potential therapeutic targets.