A Single Amino Acid Mutation in SNAP-25 Induces Anxiety-Related Behavior in Mouse

Masakazu Kataoka,Minesuke Yokoyama,Yohko Nakahara,Masami Takahashi,Hitoshi Miyaoka,Reiko Kuwahara,Rika Suzuki-Migishima,Itsuko Nihonmatsu,Shigeru Watanabe,Shiro Ikegami,Saori Yamamori,Sadahiro Azuma,Yuko Katoh-Fukui,Taku Sato,Kaoru Inokuchi,Eiji Suzuki

doi:10.1371/journal.pone.0025158

Abstract

Synaptosomal-associated protein of 25 kDa (SNAP-25) is a presynaptic protein essential for neurotransmitter release. Previously, we demonstrate that protein kinase C (PKC) phosphorylates Ser187 of SNAP-25, and enhances neurotransmitter release by recruiting secretory vesicles near to the plasma membrane. As PKC is abundant in the brain and SNAP-25 is essential for synaptic transmission, SNAP-25 phosphorylation is likely to play a crucial role in the central nervous system. We therefore generated a mutant mouse, substituting Ser187 of SNAP-25 with Ala using “knock-in” technology. The most striking effect of the mutation was observed in their behavior. The homozygous mutant mice froze readily in response to environmental change, and showed strong anxiety-related behavior in general activity and light and dark preference tests. In addition, the mutant mice sometimes exhibited spontaneously occurring convulsive seizures. Microdialysis measurements revealed that serotonin and dopamine release were markedly reduced in amygdala. These results clearly indicate that PKC-dependent SNAP-25 phosphorylation plays a critical role in the regulation of emotional behavior as well as the suppression of epileptic seizures, and the lack of enhancement of monoamine release is one of the possible mechanisms underlying these defects.

Highlights

Synaptic transmission requires neurotransmitter release from presynaptic nerve terminals
We found that DA and 5-HT release in the amygdala were markedly reduced despite the fact that neurotransmitter content were not decreased in the mutant mice
The expression level of Synaptosomal-associated protein of 25 kDa (SNAP-25) in Snap25S187A/S187A mice decreased to about 50% of that in WT mice and certain aspects of the phenotypes might be uniquely derived from reduced expression

Summary

Introduction

Synaptic transmission requires neurotransmitter release from presynaptic nerve terminals. Three SNARE proteins, VAMP-2/ synaptobrevin 2 in the synaptic vesicle membrane, and SNAP-25 and syntaxin 1 in synaptic plasma membrane, play crucial roles in the exocytotic release of neurotransmitters [1,2,3,4]. Neurotransmitter release is enhanced by the activation of protein kinase C (PKC) [8,9]. We showed that PKC activation induced enhancement of Ca2+-dependent release of dopamine (DA) and acetylcholine (ACh) from PC12 cells, and Ser187 was phosphorylated in these conditions [10,11]. In adrenal chromaffin cells and insulin secreting cells, PKC activation enhanced exocytotic release of these hormones by increasing the size of the readily releasable vesicle pool and the highly Ca2+-sensitive vesicle pool (HCSP), and phosphorylation of SNAP-25 at Ser187 was essential for these effects [13,14,15,16]. The physiological roles of phosphorylation at this site are still obscure [18,22]

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