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Objective: To investigate the whole brain input and output projection of dopamine neuron in the ventral tegmental areal (VTA), analyse the whole brain upstream and downstream neural circuits connections. Methods: The mixture of helper viruses AAV-EF1a-DIO-GT and AAV-EF1a-DIO-G (1:1) was injected into the VTA of DAT-cre transgenic mice under stereotaxic apparatus. After two weeks, recombinant rabies virus (RV) EnVA-RV-mCherry was microinjected into VTA, One week later, the RV virus completed retrograde transsynaptic infection and fully expressed the fluorescent protein, prepare whole brain frozen section, scan the sections with an automatic scanning fluorescence microscope. The anterograde tracing virus AAV-EF1a-DIO-GFP was microinjected into the VTA of DAT-cre transgenic mice under stereotaxic apparatus. After two weeks, virus and fluorescent protein were fully expressed, prepare whole brain frozen sections, the sections containing VTA were immunofluorescence stained with (tyrosine hydroxylase)TH anti- body, scan the sections with an automatic scanning fluorescence microscope. Results: RV retrograde transsynapse tracing results shows, the neurons expressed red fluorescent protein carried by RV virus distributed in many region of whole brain. Mainly includes the anterior cortex, striatum, nucleus accumbens (NAC), preoptic area of hypothalamus (POH), lateral hypothalamus (LH), paraventricular nucleus of hypothalamus (PVN), amygdala, ventral tegmental area (VTA), substantia nigra (SN), dorsal raphe (DR), parbrachial nucleus (PBN) and habenula (HB). Anterograde tracing results shows, the fibers expressing GFP were mainly concentrated in the medial prefrontal cortex (mPFC), striatum, nucleus accumbens (NAc), lateral septal nucleus (LS), amygdala, and lateral hypothalamus (LH). Conclusion: The up- stream input projections of VTA dopamine neurons were widely distributed in the whole brain, including the anterior cortex, basal gan- glia area, hypothalamus area, limbic system, and many nuclear in the midbrain. The downstream output projections of VTA dopamine neurons were mainly concentrated in the nucleus accumbens (NAC) and striatum of the basal ganglia, but also some projection in the medial prefrontal cortex(mPFC) and hypothalamus. [ABSTRACT FROM AUTHOR]

BACKGROUND: The pathogenesis of Alzheimer’s disease is closely related to abnormal mitochondrial dynamics. Our previous research demonstrated that Fasudil has neuroprotective effect. However, it is needed to explore whether Fasudil has beneficial effect on regulating mitochondiral dynamics. OBJECTIVE: To investigate the effect and mechanism of ROCK inhibitor on cognitive function and neuronal apoptosis in Alzheimer’s disease mice. METHODS: Amyloid precursor protein/presenilin-1 (APP/PS1) transgenic mice were randomly divided into the Fasudil group (25 mg/kg/d) and normal saline group (equivalent volume normal saline), and wild-type C57BL/6 mice at the same age and gender served as normal controls (same volume normal saline). Administration in each group was given via intraperitoneal injection once daily for 2 months. Spatial cognition of mice was detected by Morris water maze test and Y maze test. Nissl staining was used to observe and analyze the number and morphology of neurons. TUNEL staining was applied to observe neuronal apoptosis. The protein levels of NeuN, Bax, Bcl-2, Cleaved Caspase-3, dynamin-related protein 1 (DRP1), mitochondrial fission protein 1 (FIS1), optic atrophic protein 1 (OPA1), mitofusin 1 (Mfn1), and mitofusin 2 (Mfn2) in hippocampus tissue were determined by western blot test. The expression of NeuN and DRP1 was detected by immunofluorescence staining. RESULTS AND CONCLUSION: Fasudil ameliorated cognitive impairment and improved loss of learning, memory and exploration function in APP/PS1 mice. Compared with the normal control group, the number of survived neurons was decreased, the apoptotic rate of neurons was increased, the expression of Bax and Cleaved Caspase-3 was increased, but the expression of NeuN and Bcl-2 was decreased in APP/PS1 mice. These changes were all strongly reversed by a 2-month treatment of Fasudil. Fasudil markedly down-regulated the expression of DRP1 and FIS1, but remarkably up-regulated the expression of OPA1, Mfn1, and Mfn2. These findings indicate that Fasudil significantly improves the spatial cognitive function in APP/PS1 mice, which may be related to restoring the mitochondrial fission and fusion imbalance so as to inhibit neuronal apoptosis. [ABSTRACT FROM AUTHOR]

[Objectives] The study aimed to investigate the expression of Nesfatin-1 in reproductive axis and its role in puberty of female rats. [Methods] Female Sprague-Dawley(SD) rats were used as experimental materials, and immunohistochemical technique was used to observe the location of Nesfatin-1/Nucb2 protein on hypothalamus-pituitary-ovarian axis (HPOA) at adult female rats. Real-time fluorescent quantitative PCR(RT-qPCR) method was adopted to analyze the level of Nesfatin-1/Nucb2, Tac2, Gnrh mRNA in the hypothalamus during the different physiological stages(infant, prepuberty, puberty and adult) in female rats. The Nesfatin-1 was intracerebroventricularly injected into the hypothalamus in the prepubertal (25 day-old) female rats. When the vulva opened, the hypothalamus, pituitary, ovary and serum were collected to analyze the transcription levels of Gnrh, ERα mRNA in hypothalamus, GnrhR, FSHβ, LHβ mRNA in pituitary and FSHR,LHR mRNA in ovarian by RT-qPCR, and the level of serum E2 using ELISA. [ Results]The results showed that Nesfatin-1 was mainly distributed in the arcuate nucleus, supraoptic nucleus, para ventricular nucleus, lateral hypothalamic area, anterior pituitary, oocytes, corpus luteum, interstitial cells and granular cells. During the different physiological stages, the expression level of Nesfatin-1/Nucb2, Tac2 and Gnrh mRNA in the hypothalamus of rats increased firstly and then decreased. The expression levels of Nesfatin-1/Nucb2 mRNA were positively correlated with Tac2 mRNA (r=0.940, P<0.001), and the highest expression level at prepuberty was found compared to other periods(P<0.01). Injection of Nesfatin-1 into the lateral ventricle at 25 day-old rats could advance the onset of the puberty about 10 days, and significantly increased the weight of ovaries, the concentration of E2 in serum and the expression levels of Gnrh, GnrhR, LHβ,LHR,FSHβ and ERα mRNA(P<0.05)in reproductive axis, but not changed the expression of FSHR mRNA. [Conclusions] Nesfatin-1 is widely expressed in HPOA. Nesfatin-1 promotes the onset of puberty by up-regulating the expression of Gnrh mRNA and its receptor and reproduction-related genes. [ABSTRACT FROM AUTHOR]