Differential Effects of SNAP-25 Deletion on Ca2+-Dependent and Ca2+-Independent Neurotransmission

Department of Neuroscience, The University of Texas Southwestern Medical Center, Dallas, Texas 75390-9111, USA.
Journal of Neurophysiology (Impact Factor: 2.89). 09/2007; 98(2):794-806. DOI: 10.1152/jn.00226.2007
Source: PubMed


At the synapse, SNAP-25, along with syntaxin/HPC-1 and synaptobrevin/VAMP, forms SNARE N-ethylmaleimide-sensitive factor [soluble (NSF) attachment protein receptor] complexes that are thought to catalyze membrane fusion. Results from neuronal cultures of synaptobrevin-2 knockout (KO) mice showed that loss of synaptobrevin has a more severe effect on calcium-evoked release than on spontaneous release or on release evoked by hypertonicity. In this study, we recorded neurotransmitter release from neuronal cultures of SNAP-25 KO mice to determine whether they share this property. In neurons lacking SNAP-25, as those deficient in synaptobrevin-2, we found that approximately 10-12% of calcium-independent excitatory and inhibitory neurotransmitter release persisted. However, in contrast to synaptobrevin-2 knockouts, this remaining readily releasable pool in SNAP-25-deficient synapses was virtually insensitive to calcium-dependent-evoked stimulation. Although field stimulation reliably evoked neurotransmitter release in synaptobrevin-2 KO neurons, responses were rare in neurons lacking SNAP-25, and unlike synaptobrevin-2-deficient synapses, SNAP-25-deficient synapses did not exhibit facilitation of release during high-frequency stimulation. This severe loss of evoked exocytosis was matched by a reduction, but not a complete loss, of endocytosis during evoked stimulation. Moreover, synaptic vesicle turnover probed by FM-dye uptake and release during hypertonic stimulation was relatively unaffected by the absence of SNAP-25. This last difference indicates that in contrast to synaptobrevin, SNAP-25 does not directly function in endocytosis. Together, these results suggest that SNAP-25 has a more significant role in calcium-secretion coupling than synaptobrevin-2.

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    • "Neurons were used for experiments between 14 to 18 days in vitro. Dissociated hippocampal cultures from SNAP25 knockout mice and their wild-type littermates were generated from E17-20 embryos and were plated on poly-d-lysine coated coverslips as described previously (Bronk et al., 2007). Neurons were used for experiments 14–18 days in vitro. "
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    • "However, we found via DKO mice the roles of STX1 in Ca2+-triggered synchronous release and in neuronal viability. The phenotypes of the DKO neurons were similar to SNAP-25−/− neurons in several apparent aspects such as embryonic lethality, reduced neuron survival in culture and smaller amplitude of spontaneous events [10], [35], [36]. Cleavage of STX1 or SNAP-25 by botulinum neurotoxin induces degeneration of neurons, suggesting that STX1 and SNAP-25 cooperate to support neuron survival [37]. "
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    • "Although neurons from their wild-type littermates showed swift responses to Reelin application (Figure 4B), neurons lacking SNAP-25 (SNAP25 À/À ) showed no response to Reelin and had an overall lower mEPSC frequency (Figure 4C) (Bronk et al., 2007). Here, it is important to note that SNAP-25 deficient synapses respond to other secretagogues such as hypertonic sucrose, ionomycin, or a-latrotoxin (Bronk et al., 2007; Deá k et al., 2009). These data indicate that Reelin causes an increase in SV fusion frequency that requires the function of the plasma membrane-associated SNARE, SNAP-25, in agreement with an earlier study suggesting a SNAP-25-dependent role for Reelin in presynaptic function (Hellwig et al., 2011). "
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