Cell-Specific Loss of SNAP25 from Cortical Projection Neurons Allows Normal Development but Causes Subsequent Neurodegeneration

Abstract

Synaptosomal associated protein 25 kDa (SNAP25) is an essential component of the SNARE complex regulating synaptic vesicle fusion. SNAP25 deficiency has been implicated in a variety of cognitive disorders. We ablated SNAP25 from selected neuronal populations by generating a transgenic mouse (B6-Snap25tm3mcw (Snap25-flox)) with LoxP sites flanking exon5a/5b. In the presence of Cre-recombinase, Snap25-flox is recombined to a truncated transcript. Evoked synaptic vesicle release is severely reduced in Snap25 conditional knockout (cKO) neurons as shown by live cell imaging of synaptic vesicle fusion and whole cell patch clamp recordings in cultured hippocampal neurons. We studied Snap25 cKO in subsets of cortical projection neurons in vivo (L5-Rbp4-Cre; L6-Ntsr1-Cre; L6b-Drd1a-Cre). cKO neurons develop normal axonal projections, but axons are not maintained appropriately, showing signs of swelling, fragmentation and eventually complete absence. Onset and progression of degeneration are dependent on the neuron type, with L5 cells showing the earliest and most severe axonal loss. Ultrastructural examination revealed that cKO neurites contain autophagosome/lysosome-like structures. Markers of inflammation such as Iba1 and lipofuscin are increased only in adult cKO cortex. Snap25 cKO can provide a model to study genetic interactions with environmental influences in several disorders.

A role for the cortex in sleep-wake regulation

“…Figure 3). This result is in line with the observation that Rbp4 expression is not restricted to the layer 5 of the neocortex, but is also present in granule cells of the hippocampal dentate gyrus 25 , which is involved in generating theta activity during REM sleep 28,29 .…”

“…Transgenic animals (Rbp4-Cre;Ai14;Snap25 fl/fl ) did not show obvious physical, neurological or behavioural abnormalities but their body weight was lower compared to controls (CTR: 23.4g, HOM: 20.9g, t(13)=2.87, p=0.013) as reported previously 25 . The electrophysiological signals revealed typical signatures of wakefulness and sleep states.…”

“…To induce a cortex-wide reduction in the output from layer 5 pyramidal neurons, we generated a transgenic mouse line, in which a subpopulation (~ 15-30 %) of pyramidal cells in layer 5 of the neocortex lacked the key t-SNARE protein SNAP25 (Rbp4-Cre;Ai14;Snap25 fl/fl ) 25 . While Rbp4-Cre is known as a pan layer 5 driver line 26 , it additionally presents a strong Cre-expression in hippocampal dentate gyrus, but was only sparsely expressed in the hypothalamus, typically considered a major brain area involved in sleep-wake state transitions (Suppl.…”

“…Figure 2). Cre-dependent excision of exon 5a/5b leads to reduced length gene transcript and non-detectable levels of SNAP25 protein in Cre-positive neurons 25 . As well known, ablation of SNAP25 virtually abolishes calcium-evoked neurotransmitter release from the respective neurons 25,27 , which renders the cells carrying the mutation functionally silenced.…”

Genetic inactivation of synaptosomal‐associated protein 25 (SNAP‐25) in adult hippocampal neural progenitors impairs pattern discrimination learning but not survival or structural maturation of newborn dentate granule cells

“…Likewise, recent studies using transgenic mice that allow for selective induction of tetanus toxin expression in Emx1-derived neurons, (which represent >95% of principal neurons in the dorsal telencephalon), demonstrate remarkable assembly and maintenance of excitatory synapses in the absence of glutamatergic neurotransmission in most principle cells of the hippocampal trisynaptic circuit during early postnatal development (Sando et al, 2017). Pertinent to the current research, a recent study has found that conditional loss of SNAP-25 from long-range projecting cortical neurons from embryonic stages allows normal axonal projections but results in eventual cell death months after silencing, although DGCs appeared to remain intact (Hoerder-Suabedissen et al, 2018). Our findings add to this literature to demonstrate the ability of adult-generated DGCs to survive, undergo structural maturation and form anatomical efferent connections when AP-dependent neurotransmission is blocked at the early hippocampal progenitor stage in adult mice.…”