Neurotransmission: Getting vesicles ready for action

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CAPS (Ca²⁺-dependent activator protein of secretion) proteins play an important role in priming synaptic vesicles for exocytosis and neurotransmitter release.

The release of neurotransmitters from synaptic vesicles in response to stimulation is crucial for information transfer between neurons. Before exocytosis takes place synaptic vesicles must be primed, yet the molecular mechanisms responsible for this process are only partially understood. Jockusch et al. have now identified a central role for CAPS (Ca²⁺-dependent activator protein of secretion) proteins — previously linked to the exocytosis of large dense-core vesicles — in synaptic-vesicle priming.

The authors cultured glutamatergic hippocampal neurons from wild-type, CAPS1-knockout (KO), CAPS2-KO and CAPS1;CAPS2 double-knockout (CAPS DKO) mice. The neurons formed synapses with themselves (autapses), allowing the researchers to analyze their synaptic electrophysiology. Neurons that lacked CAPS1 or both CAPS proteins demonstrated reduced evoked excitatory postsynaptic currents (EPSCs), which correlated with a reduction in the size of the 'readily releasable' pool of primed vesicles.

To determine the cause of the reduced EPSCs, the authors carried out several experiments. There were no major changes in synaptic morphology or in the total number of synapses in the mutant cultures, and when neurotransmitters were applied exogenously the postsynaptic responses were normal. The amplitude of miniature EPSCs (mEPSCs) was similar in CAPS1-KO and CAPS DKO cultures, but the frequency of the mEPSCs was reduced to a greater extent in CAPS DKO neurons, indicating that vesicle priming rather than vesicle filling was defective.

Next, the authors showed that high-frequency stimulation of the neurons overcame the effects of the absence of the CAPS proteins, augmenting the EPSC amplitude to wild-type levels and increasing the readily releasable vesicle pool in the CAPS DKO neurons. Similarly, mimicking the effects of high-frequency stimulation by increasing intracellular Ca²⁺ levels with calcimycin caused CAPS DKO neurons to spontaneously release glutamate to the same extent as wild-type neurons.

What is the relationship between the CAPS proteins and other proteins that have been linked to vesicle priming, such as Munc13-1? The authors showed that Munc13-1 levels are not altered by the loss of CAPS proteins. Treatment of CAPS DKO neurons with PDBU, which causes Munc13-dependent priming, increased EPSC amplitude, suggesting that the residual release capacity that is observed in the absence of CAPS proteins is mediated by Munc13-1. Overexpressing CAPS1 in neurons that lacked Munc13-1, or overexpressing Munc13-1 in CAPS DKO neurons, did not rescue the mutant phenotypes, suggesting that both proteins act in the same pathway or work together to prime vesicles.

This study furthers our knowledge of the protein interactions that are responsible for synaptic-vesicle exocytosis. The next challenge will be to understand how the CAPS proteins work together with Munc13-1 and other members of the priming complex to mediate this process.

Katherine Whalley

References

Jockusch, W. J. et al. CAPS-1 and CAPS-2 are essential synaptic vesicle priming proteins. Cell 131, 796–808 (2007) | Article | PubMed |