miR-218-2 regulates cognitive functions in the hippocampus through complement component 3-dependent modulation of synaptic vesicle release.

microRNA-218 (miR-218) has been linked to several cognition related neurodegenerative and neuropsychiatric disorders. However, whether miR-218 plays a direct role in cognitive functions remains unknown. Here, using the miR-218 knockout (KO) mouse model and the sponge/overexpression approaches, we showed that miR-218-2 but not miR-218-1 could bidirectionally regulate the contextual and spatial memory in the mice. Furthermore, miR-218-2 deficiency induced deficits in the morphology and presynaptic neurotransmitter release in the hippocampus to impair the long term potentiation. Combining the RNA sequencing analysis and luciferase reporter assay, we identified complement component 3 (C3) as a main target gene of miR-218 in the hippocampus to regulate the presynaptic functions. Finally, we showed that restoring the C3 activity in the miR-218-2 KO mice could rescue the synaptic and learning deficits. Therefore, miR-218-2 played an important role in the cognitive functions of mice through C3, which can be a mechanism for the defective cognition of miR-218 related neuronal disorders.

Synaptotagmin-7 deficiency induces mania-like behavioral abnormalities through attenuating GluN2B activity.

Synaptotagmin-7 (Syt7) probably plays an important role in bipolar-like behavioral abnormalities in mice; however, the underlying mechanisms for this have remained elusive. Unlike antidepressants that cause mood overcorrection in bipolar depression, N-methyl-d-aspartate receptor (NMDAR)-targeted drugs show moderate clinical efficacy, for unexplained reasons. Here we identified Syt7 single nucleotide polymorphisms (SNPs) in patients with bipolar disorder and demonstrated that mice lacking Syt7 or expressing the SNPs showed GluN2B-NMDAR dysfunction, leading to antidepressant behavioral consequences and avoidance of overcorrection by NMDAR antagonists. In human induced pluripotent stem cell (iPSC)-derived and mouse hippocampal neurons, Syt7 and GluN2B-NMDARs were localized to the peripheral synaptic region, and Syt7 triggered multiple forms of glutamate release to efficiently activate the juxtaposed GluN2B-NMDARs. Thus, while Syt7 deficiency and SNPs induced GluN2B-NMDAR dysfunction in mice, patient iPSC-derived neurons showed Syt7 deficit-induced GluN2B-NMDAR hypoactivity that was rescued by Syt7 overexpression. Therefore, Syt7 deficits induced mania-like behaviors in mice by attenuating GluN2B activity, which enabled NMDAR antagonists to avoid mood overcorrection.

Synaptotagmin-7 is a key factor for bipolar-like behavioral abnormalities in mice.

The pathogenesis of bipolar disorder (BD) has remained enigmatic, largely because genetic animal models based on identified susceptible genes have often failed to show core symptoms of spontaneous mood cycling. However, pedigree and induced pluripotent stem cell (iPSC)-based analyses have implicated that dysfunction in some key signaling cascades might be crucial for the disease pathogenesis in a subpopulation of BD patients. We hypothesized that the behavioral abnormalities of patients and the comorbid metabolic abnormalities might share some identical molecular mechanism. Hence, we investigated the expression of insulin/synapse dually functioning genes in neurons derived from the iPSCs of BD patients and the behavioral phenotype of mice with these genes silenced in the hippocampus. By these means, we identified synaptotagmin-7 (Syt7) as a candidate risk factor for behavioral abnormalities. We then investigated Syt7 knockout (KO) mice and observed nocturnal manic-like and diurnal depressive-like behavioral fluctuations in a majority of these animals, analogous to the mood cycling symptoms of BD. We treated the Syt7 KO mice with clinical BD drugs including olanzapine and lithium, and found that the drug treatments could efficiently regulate the behavioral abnormalities of the Syt7 KO mice. To further verify whether Syt7 deficits existed in BD patients, we investigated the plasma samples of 20 BD patients and found that the Syt7 mRNA level was significantly attenuated in the patient plasma compared to the healthy controls. We therefore concluded that Syt7 is likely a key factor for the bipolar-like behavioral abnormalities.

Application of induced pluripotent stem cells to understand neurobiological basis of bipolar disorder and schizophrenia.

The etiology of neuropsychiatric disorders, such as schizophrenia and bipolar disorder, usually involves complex combinations of genetic defects/variations and environmental impacts, which hindered, for a long time, research efforts based on animal models and patients' non-neuronal cells or post-mortem tissues. However, the development of human induced pluripotent stem cell (iPSC) technology by the Yamanaka group was immediately applied to establish cell research models for neuronal disorders. Since then, techniques to achieve highly efficient differentiation of different types of neural cells following iPSC modeling have made much progress. The fast-growing iPSC and neural differentiation techniques have brought valuable insights into the pathology and neurobiology of neuropsychiatric disorders. In this article, we first review the application of iPSC technology in modeling neuronal disorders and discuss the progress in the accompanying neural differentiation. Then, we summarize the progress in iPSC-based research that has been accomplished so far regarding schizophrenia and bipolar disorder.

[Effect of FGF-21 on learning and memory ability and antioxidant capacity in brain tissue of D-galactose-induced aging mice].

This study aims to investigate the effects of fibroblast growth factor 21 (FGF-21) on learning and memory abilities and antioxidant capacity of D-galactose-induced aging mice. Kunming mice (37.1 +/- 0.62) g were randomly divided into normal control group, model group and FGF-21 high, medium and low dose groups (n = 8). Each group was injected in cervical part subcutaneously with D-galactose 180 mg x kg(-1) x d(-1) once a day for 8 weeks. At the same time, FGF-21-treated mice were administered with FGF-21 by giving subcutaneous injection in cervical part at the daily doses of 5, 2 and 1 mg x kg(-1) x d(-1). The normal control group was given with normal saline by subcutaneous injection in cervical part. At seventh week of the experiment, the learning and memory abilities of mice were determined by water maze and jumping stand tests. At the end of the experiment, the mice were sacrificed and the cells damage of hippocampus was observed by HE staining in each group. Reactive oxygen species (ROS), malondialdehyde (MDA), superoxide dismutase (SOD), glutathione peroxidase (GPx), catalase (CAT) and total antioxidant capacity (T-AOC) in the brain of mice were determined. The results showed that different doses of FGF-21 could reduce the time reaching the end (P < 0.01 or P < 0.05) and the number of touching blind side (P < 0.01 or P < 0.05) in the water maze comparing with the model group. It could also prolong the latency time (P < 0.05) and decrease the number of errors (P < 0.01 or P < 0.05) in the step down test. The result of HE staining showed that FGF-21 could significantly reduce brain cell damage in the hippocampus. The ROS and MDA levels of three different doses FGF-21 treatment group reduced significantly than that of the model group [(5.58 +/- 1.07), (7.78 +/- 1.92), (9.03 +/- 1.77) vs (12.75 +/- 2.02) pmol (DCF) x min(-1) x mg(-1), P < 0.01 or P < 0.05], [(2.92 +/- 0.71), (4.21 +/- 0.81), (4.41 +/- 0.97) vs (5.62 +/- 0.63) nmol x mg(-1) (protein), P < 0.01]. Comparing with the model group, the activities of SOD, GPx, CAT and T-AOC of the three different doses FGF-21 treatment groups were also improved in a dose-dependent manner. This study demonstrates that FGF-21 can ameliorate learning and memory abilities of D-galactose induced aging mice, improve the antioxidant abilities in brain tissue and delay brain aging. This finding provides a theoretical support for clinical application of FGF-21 as a novel therapeutics for preventing aging.

Synaptotagmin-1 interacts with PI(4,5)P2 to initiate synaptic vesicle docking in hippocampal neurons.

Synaptic vesicle (SV) docking is a dynamic multi-stage process that is required for efficient neurotransmitter release in response to nerve impulses. Although the steady-state SV docking likely involves the cooperation of Synaptotagmin-1 (Syt1) and soluble N-ethylmaleimide-sensitive factor attachment protein receptors (SNAREs), where and how the docking process initiates remains unknown. Phosphatidylinositol-4,5-bisphosphate (PI(4,5)P2) can interact with Syt1 and SNAREs to contribute to vesicle exocytosis. In the present study, using the CRISPRi-mediated multiplex gene knockdown and 3D electron tomography approaches, we show that in mouse hippocampal synapses, SV docking initiates at ∼12 nm to the active zone (AZ) by Syt1. Furthermore, we demonstrate that PI(4,5)P2 is the membrane partner of Syt1 to initiate SV docking, and disrupting their interaction could abolish the docking initiation. In contrast, the SNARE complex contributes only to the tight SV docking within 0-2 nm. Therefore, Syt1 interacts with PI(4,5)P2 to loosely dock SVs within 2-12 nm to the AZ in hippocampal neurons.

Author Correction: CRISPR interference-based specific and efficient gene inactivation in the brain.

In the version of this article initially published, the affiliation for Jian Zhang and Shuangli Mi was incomplete. In addition to the Key Laboratory of Genomics and Precision Medicine, they are also affiliated with the University of Chinese Academy of Sciences, Beijing, China. In Supplementary Fig. 1h,l, the molecular mass marker accompanying Snap25 was labeled 58 kDa; the correct value is 25 kDa. In Supplementary Fig. 9b,c, the top panel was labeled Syt1, with molecular mass markers ranging from 46 to 100 kDa; it is actually Snap25, with molecular mass markers ranging from 17 to 46 kDa. The errors have been corrected in the HTML and PDF versions of the article.

CRISPR interference-based specific and efficient gene inactivation in the brain.

CRISPR-Cas9 has been demonstrated to delete genes in postmitotic neurons. Compared to the establishment of proliferative cell lines or animal strains, it is more challenging to acquire a highly homogeneous consequence of gene editing in a stable neural network. Here we show that dCas9-based CRISPR interference (CRISPRi) can efficiently silence genes in neurons. Using a pseudotarget fishing strategy, we demonstrate that CRISPRi shows superior targeting specificity without detectable off-target activity. Furthermore, CRISPRi can achieve multiplex inactivation of genes fundamental for neurotransmitter release with high efficiency. By developing conditional CRISPRi tools targeting synaptotagmin I (Syt1), we modified the excitatory to inhibitory balance in the dentate gyrus of the mouse hippocampus and found that the dentate gyrus has distinct regulatory roles in learning and affective processes in mice. We therefore recommend CRISPRi as a useful tool for more rapid investigation of gene function in the mammalian brain.