Serine-Arginine Rich Protein Kinase 2 regulates number of docked vesicles confirmed by FIB-SEM

Isabelle Paulussen1, Alexander Mueller2, Julia Betzin2, Susanne Schoch2, Dirk Dietrich1

1 Experimental Neurophysiology, Department of Neurosurgery, University Clinic Bonn, Germany
2 Institute of Neuropathology and Department of Epileptology, University Clinic Bonn, Germany

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Serine-Arginine Rich Protein Kinase 2 (SRPK2) was recently identified as a novel kinase in mammalian neurons. Cultured neurons overexpressing SRPK2 revealed a 32% (±11%, n=6) increase in stimulated transmitter release compared to control (n=6) by using a vesicle release assay (FM4-64 dye). On the other hand, knock-down of SRPK2 significantly decreased the release by 17% (±8%, n=6). Thus, SRPK2 levels regulate synaptic release probability (Pr) in hippocampal neurons. However, the exact mechanism how SRPK2 controls Pr is still not understood. Pr could be increased by SRPK2 either by an enhanced vesicular release probability or by an increased number of readily releasable vesicles per synapse. To differentiate between these possibilities, we used focused ion beam-scanning electron microscopy and counted the number of docked vesicles per synapse in 3D reconstructions. Neurons overexpressing SRPK2 displayed a 57% (±10%, n=6) increase in the number of docked vesicles (from 5 to 8 docked vesicles) which can thus fully account for the observed enhancement of release. SRPK2 also increased the active zone as assessed by measuring the postsynaptic density (PSD) area (from 0.039±0.003 µm2 to 0.069±0.009 µm2, n=5 and n=6). SRPK2 knock-down did not change the number of docked vesicles (4 docked vesicles, n=5) or the PSD area (0.035±0.001 µm2, n=5).

This data suggests that maintenance of basal synaptic ultrastructure is not dependent on SRPK2, but that SRPK2 might play an important role in triggering a series of actions resulting in increased pre- and postsynaptic specialization required for mediating an enhanced transmission in presynaptic plasticity paradigms.