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β-site amyloid precursor protein cleaving enzyme 1 (BACE1) inhibition impairs synaptic plasticity via seizure protein 6 (SZE6)
β-site amyloid precursor protein cleaving enzyme 1 (BACE1) inhibition impairs synaptic plasticity via seizure protein 6 (SZE6)
Alzheimer’s disease (AD) is the most prevalent neurodegenerative disorder among the elderly. Amyloid-β is thought to be one of the causative factors for AD, which is produced by BACE1 (Beta-secretase) initiated sequential proteolytic cleavage of APP. BACE1 inhibition is one of the promising therapeutic approaches for AD. Currently, several BACE1 inhibitors are undergoing Phase 2/3 clinical trials. However, prolonged BACE1 inhibition interferes structural and functional synaptic plasticity in mice, most likely due to the interrupted metabolism of BACE1 substrates. Seizure protein 6 (SEZ6) is predominantly cleaved by BACE1. Furthermore, Sez6 null mice share some phenotype with BACE1-inhibited mice including reduced dendritic spine density in cortex and diminished performance in hippocampal-dependent behavioral tests. In order to shed more light on the function of SEZ6, we analyzed the dendritic spine structure and synaptic plasticity in constitutive (Sez6-/-:GFP-M) and conditional (Sez6cKO/cKO:SlickV) Sez6 KO mice. In vivo two photon microscopy data showed that lack of SEZ6 induces a dose dependent alteration of dendritic spine density and morphology in adult mice. To rule out developmental deficits and identify which SEZ6 proteolytic fragments are involved we monitored spine density in Sez6cKO/cKO:SlickV mice. The tamoxifen-inducible recombinase CreERT2 and eYFP are co-expressed in a small subset of neurons in SlickV mice. By applying tamoxifen, Sez6 was knockout specifically in eYFP positive neurons in adult mice. It caused a small but significant spine density reduction. Electrophysiological field recordings in hippocampus CA1 region showed that SEZ6 is involved in synaptic transmission and LTP mainly due to post-synaptic mechanism. To study the dendritic spine plasticity in Sez6-/-:GFP-M mice, we repeatedly imaged the apical tufts of layer V pyramidal neurons in the cerebral cortex in both normal condition and environmental enrichment condition by intravital two-photon microscopy. Sez6-/- mice does not show alerted dendritic spine plasticity in base line condition, but they have deficits in conditions that boost spine plasticity like environmental enrichment. Then, we investigated whether SEZ6 is involved in BACE1-inhibition-induced synaptic alteration. We applied a diet mixed with NB-360 to Sez6-/-:GFP-M and Sez6cKO/cKO:SlickV mice. NB-360 is a novel blood-brain barrier penetrable BACE1 inhibitor. Immunoblotting analysis showed that NB-360 strongly suppressed SEZ6 and APP cleavage similar to Bace1 knockout. To study the impact of long-term pharmacological inhibition of BACE1 in Sez6-/-:GFP-M mice, we repeatedly imaged the apical tufts of layer V pyramidal neurons in the cerebral cortex for 7 weeks using intravital two-photon microscopy. Although 3-week treatment of NB-360 caused a significant but reversible reduction of density of total dendritic spines, persistent spines (persisting ⩾ 7 days) and new gained spines in control mice, the same treatment did not affect dendritic spine dynamics in Sez6-/-:GFP-M mice. To rule out developmental deficits, we monitored spine dynamics upon NB-360 treatment in Sez6cKO/cKO:SlickV mice. Chronic NB-360 treatment did not alter spine plasticity in the neurons lacking cell-autonomous SEZ6. Finally, electrophysiological field recordings in hippocampal CA1 region showed that LTP is reduced in chronic NB-360 treated WT mice and vehicle treated Sez6-/- mice, but NB-360 treatment did not interfere with LTP in Sez6-/- mice. Our data suggest that SEZ6 has a pivotal role in maintaining normal dendritic spine structure and function. Furthermore, SEZ6 is involved in BACE1-inhibitor-induced structural and functional synaptic alterations.
BACE1, BACE inhibitor, SEZ6, AD
Zhu, Kaichuan
2018
English
Universitätsbibliothek der Ludwig-Maximilians-Universität München
Zhu, Kaichuan (2018): β-site amyloid precursor protein cleaving enzyme 1 (BACE1) inhibition impairs synaptic plasticity via seizure protein 6 (SZE6). Dissertation, LMU München: Faculty of Chemistry and Pharmacy
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Abstract

Alzheimer’s disease (AD) is the most prevalent neurodegenerative disorder among the elderly. Amyloid-β is thought to be one of the causative factors for AD, which is produced by BACE1 (Beta-secretase) initiated sequential proteolytic cleavage of APP. BACE1 inhibition is one of the promising therapeutic approaches for AD. Currently, several BACE1 inhibitors are undergoing Phase 2/3 clinical trials. However, prolonged BACE1 inhibition interferes structural and functional synaptic plasticity in mice, most likely due to the interrupted metabolism of BACE1 substrates. Seizure protein 6 (SEZ6) is predominantly cleaved by BACE1. Furthermore, Sez6 null mice share some phenotype with BACE1-inhibited mice including reduced dendritic spine density in cortex and diminished performance in hippocampal-dependent behavioral tests. In order to shed more light on the function of SEZ6, we analyzed the dendritic spine structure and synaptic plasticity in constitutive (Sez6-/-:GFP-M) and conditional (Sez6cKO/cKO:SlickV) Sez6 KO mice. In vivo two photon microscopy data showed that lack of SEZ6 induces a dose dependent alteration of dendritic spine density and morphology in adult mice. To rule out developmental deficits and identify which SEZ6 proteolytic fragments are involved we monitored spine density in Sez6cKO/cKO:SlickV mice. The tamoxifen-inducible recombinase CreERT2 and eYFP are co-expressed in a small subset of neurons in SlickV mice. By applying tamoxifen, Sez6 was knockout specifically in eYFP positive neurons in adult mice. It caused a small but significant spine density reduction. Electrophysiological field recordings in hippocampus CA1 region showed that SEZ6 is involved in synaptic transmission and LTP mainly due to post-synaptic mechanism. To study the dendritic spine plasticity in Sez6-/-:GFP-M mice, we repeatedly imaged the apical tufts of layer V pyramidal neurons in the cerebral cortex in both normal condition and environmental enrichment condition by intravital two-photon microscopy. Sez6-/- mice does not show alerted dendritic spine plasticity in base line condition, but they have deficits in conditions that boost spine plasticity like environmental enrichment. Then, we investigated whether SEZ6 is involved in BACE1-inhibition-induced synaptic alteration. We applied a diet mixed with NB-360 to Sez6-/-:GFP-M and Sez6cKO/cKO:SlickV mice. NB-360 is a novel blood-brain barrier penetrable BACE1 inhibitor. Immunoblotting analysis showed that NB-360 strongly suppressed SEZ6 and APP cleavage similar to Bace1 knockout. To study the impact of long-term pharmacological inhibition of BACE1 in Sez6-/-:GFP-M mice, we repeatedly imaged the apical tufts of layer V pyramidal neurons in the cerebral cortex for 7 weeks using intravital two-photon microscopy. Although 3-week treatment of NB-360 caused a significant but reversible reduction of density of total dendritic spines, persistent spines (persisting ⩾ 7 days) and new gained spines in control mice, the same treatment did not affect dendritic spine dynamics in Sez6-/-:GFP-M mice. To rule out developmental deficits, we monitored spine dynamics upon NB-360 treatment in Sez6cKO/cKO:SlickV mice. Chronic NB-360 treatment did not alter spine plasticity in the neurons lacking cell-autonomous SEZ6. Finally, electrophysiological field recordings in hippocampal CA1 region showed that LTP is reduced in chronic NB-360 treated WT mice and vehicle treated Sez6-/- mice, but NB-360 treatment did not interfere with LTP in Sez6-/- mice. Our data suggest that SEZ6 has a pivotal role in maintaining normal dendritic spine structure and function. Furthermore, SEZ6 is involved in BACE1-inhibitor-induced structural and functional synaptic alterations.