Additional file 1: Figure S1. of Lrp5/6 are required for cerebellar development and for suppressing TH expression in Purkinje cells via β-catenin

The earliest stage of defective cerebellar morphology in Lrp5/6 dCKO mice. (a-d) Nissl staining shows that failure of cerebellar foliation is not evident in the dCKO mice at P0 (b) but is observed at P3 (d) compared to control mice (a, c). (e-h) Pax6 expression in control and Lrp5/6 dCKO mice at P0 (e, g) and P3 (f, h). EGL, external granule layer. Scale bars, 500 μm in (d) and applies to (a-c), and 25 μm in (h) and applies to (e-g). (PDF 400 kb

<i>Atg5</i>- and <i>Atg7</i>-dependent autophagy in dopaminergic neurons regulates cellular and behavioral responses to morphine

<p>The molecular basis of chronic morphine exposure remains unknown. In this study, we hypothesized that macroautophagy/autophagy of dopaminergic neurons would mediate the alterations of neuronal dendritic morphology and behavioral responses induced by morphine. Chronic morphine exposure caused <i>Atg5</i> (autophagy-related 5)- and <i>Atg7</i> (autophagy-related 7)-dependent and dopaminergic neuron-specific autophagy resulting in decreased neuron dendritic spines and the onset of addictive behaviors. In cultured primary midbrain neurons, morphine treatment significantly reduced total dendritic length and complexity, and this effect could be reversed by knockdown of <i>Atg5</i> or <i>Atg7</i>. Mice deficient for <i>Atg5</i> or <i>Atg7</i> specifically in the dopaminergic neurons were less sensitive to developing a morphine reward response, behavioral sensitization, analgesic tolerance and physical dependence compared to wild-type mice. Taken together, our findings suggested that the <i>Atg5</i>- and <i>Atg7</i>-dependent autophagy of dopaminergic neurons contributed to cellular and behavioral responses to morphine and may have implications for the future treatment of drug addiction.</p

Additional file 2: Table S1. of Oxytocin is implicated in social memory deficits induced by early sensory deprivation in mice

Gender effects in the behavioral tests. (DOCX 48 kb

Additional file 3: Figure S2. of Oxytocin is implicated in social memory deficits induced by early sensory deprivation in mice

OXT levels in the brain is elevated after intranasal administration of OXT. (JPG 919 kb

Serotonin (5-HT) neuron-specific inactivation of Cadherin-13 impacts 5-HT system formation and cognitive function

Genome-wide screening approaches identified the cell adhesion molecule Cadherin-13 (CDH13) as a risk factor for neurodevelopmental disorders, nevertheless the contribution of CDH13 to the disease mechanism remains obscure. CDH13 is involved in neurite outgrowth and axon guidance during early brain development and we previously provided evidence that constitutive CDH13 deficiency influences the formation of the raphe serotonin (5-HT) system by modifying neuron-radial glia interaction. Here, we dissect the specific impact of CDH13 on 5-HT system development and function using a 5-HT neuron-specific Cdh13 knockout mouse model (conditional Cdh13 knockout, Cdh13 cKO). Our results show that exclusive inactivation of CDH13 in 5-HT neurons selectively increases 5-HT neuron density in the embryonic dorsal raphe, with persistence into adulthood, and serotonergic innervation of the developing prefrontal cortex. At the behavioral level, adult Cdh13 cKO mice display delayed acquisition of several learning tasks and a subtle impulsive-like phenotype, with decreased latency in a sociability paradigm alongside with deficits in visuospatial memory. Anxiety-related traits were not observed in Cdh13 cKO mice. Our findings further support the critical role of CDH13 in the development of dorsal raphe 5-HT circuitries, a mechanism that may underlie specific clinical features observed in neurodevelopmental disorders

The rostromedial tegmental nucleus is essential for non-rapid eye movement sleep

<div><p>The rostromedial tegmental nucleus (RMTg), also called the GABAergic tail of the ventral tegmental area, projects to the midbrain dopaminergic system, dorsal raphe nucleus, locus coeruleus, and other regions. Whether the RMTg is involved in sleep–wake regulation is unknown. In the present study, pharmacogenetic activation of rat RMTg neurons promoted non-rapid eye movement (NREM) sleep with increased slow-wave activity (SWA). Conversely, rats after neurotoxic lesions of 8 or 16 days showed decreased NREM sleep with reduced SWA at lights on. The reduced SWA persisted at least 25 days after lesions. Similarly, pharmacological and pharmacogenetic inactivation of rat RMTg neurons decreased NREM sleep. Electrophysiological experiments combined with optogenetics showed a direct inhibitory connection between the terminals of RMTg neurons and midbrain dopaminergic neurons. The bidirectional effects of the RMTg on the sleep–wake cycle were mimicked by the modulation of ventral tegmental area (VTA)/substantia nigra compacta (SNc) dopaminergic neuronal activity using a pharmacogenetic approach. Furthermore, during the 2-hour recovery period following 6-hour sleep deprivation, the amount of NREM sleep in both the lesion and control rats was significantly increased compared with baseline levels; however, only the control rats showed a significant increase in SWA compared with baseline levels. Collectively, our findings reveal an essential role of the RMTg in the promotion of NREM sleep and homeostatic regulation.</p></div

Pharmacogenetic inhibition of ventral tegmental area (VTA)/substantia nigra compacta (SNc) dopaminergic (DAergic) neurons in tyrosine hydroxylase (TH)-Cre mice.

<p>Schematic representation of Cre-dependent adeno-associated virus (AAV) vectors expressing hM3Dq/hM4Di-mCherry under the control of human synapsin (hSyn) promoter. WPRE, woodchuck hepatitis virus post-transcriptional regulatory element; CNO, clozapine-N-oxide; IP, intraperitoneal. (B) Microinjection of AAV-DIO-hM4Di-mCherry into the VTA induced hM4Di expression (red). ml, medial lemniscus; SNc, substantia nigra, compacta; SNr, substantia nigra, reticular part. (C) Representative photomicrographs of the VTA depicting mCherry expression (red), TH (green), and 4,6-diamidino-2-phenylindole (DAPI, blue) immunoreactivity and merge images (yellow) from a TH-Cre mouse microinjected with Cre-dependent AAV vectors containing hM4Di. (D) Statistics of the coexpression of TH and mCherry immunofluorescence of VTA neurons (<i>n</i> = 3, per group). (E) Drawings of superimposed AAV-injected area in the VTA (left, <i>n</i> = 8) and SNc (right, <i>n</i> = 8) of TH-Cre mice with different colors. (F, H) Representative photomicrographs of mCherry expression after microinjection of AAVs containing hM4Di into the VTA (F) and SNc (H). (G, I) CNO (G, I; top) and saline (G, I; bottom) did not induce c-Fos expression in hM4Di-expressing DAergic neurons in VTA (G) or SNc (I). (J) In TH-Cre mice, the firing property of a recorded VTA hM4Di/mCherry-positive DAergic neuron with hyperpolarization-activated cation current (<i>I</i>_h) (top) and CNO perfusion inhibited its firing rate (bottom). Underlying data can be found in <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.2002909#pbio.2002909.s001" target="_blank">S1 Data</a>.</p

Pharmacogenetic inhibition of ventral tegmental area (VTA)/substantia nigra compacta (SNc) dopaminergic (DAergic) neurons promoted non-rapid eye movement (NREM) sleep in tyrosine hydroxylase (TH)-Cre mice.

<p>(A, C) Time course changes of NREM and REM sleep and wakefulness after administration of saline or clozapine-N-oxide (CNO) in hM4Di-expressing TH-Cre mice in the VTA (A, <i>n</i> = 8) and SNc (C, <i>n</i> = 8) DAergic neurons. (B, D) Total sleep–wake amounts during 09:00–13:00 hours (B, VTA) or 09:00–12:00 hours (D, SNc) after administration of saline or CNO. (E–H) Number of NREM sleep bouts with different duration (E, G) and average power density of NREM sleep (F, H) in 4 hours (E, F, VTA) or 3 hours (G, H, SNc) after saline or CNO injection in TH-Cre mice. *<i>p</i> < 0.05, **<i>p</i> < 0.01 versus saline by paired <i>t</i> tests. CNO (1.0 mg/kg) was given by intraperitoneal (IP) injection at 09:00 hours. The horizontal open and filled bars on the <i>x</i>-axes indicate the 12-hour light period and the 12-hour dark period, respectively. Underlying data can be found in <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.2002909#pbio.2002909.s001" target="_blank">S1 Data</a>.</p

Rats with RMTg lesions by ibotenic acid for 8 days showed a deficit in rebound of non-rapid eye movement (NREM) sleep quality in response to 6-hour sleep deprivation (SD).

<p>(A, B) Time course of NREM sleep in control (A) and RMTg lesioned rats (B). Sleep was deprived from 13:00 to 19:00 hours. (C, D) Amount of NREM sleep (C) and mean duration of NREM sleep (D) during 19:00–21:00 hours following 6-hour SD. (E, F) Time course of slow-wave activity (SWA) (expressed as percentage of delta power [0.5–4 Hz] in NREM sleep per hour) from 19:00 to 07:00 hours following 6-hour SD in control (E) and RMTg lesioned rats (F). *<i>p</i> < 0.05, **<i>p</i> < 0.01 versus corresponding baseline using 2-way ANOVA followed by paired <i>t</i> tests. Control (<i>n</i> = 6); lesion (<i>n</i> = 4). The horizontal open and filled bars on the <i>x</i>-axes indicate the 12-hour light and the 12-hour dark period, respectively. Underlying data can be found in <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.2002909#pbio.2002909.s001" target="_blank">S1 Data</a>.</p

Pharmacogenetic activation of rostromedial tegmental nucleus (RMTg) neurons increased non-rapid eye movement (NREM) sleep in rats.

<p>(A, B) Sleep–wake quantities following saline and clozapine-N-oxide (CNO) injection, including average hourly (A) and total sleep–wake amounts (B) during the post-injection period (09:00–16:00 hours). (C–G) Sleep–wake architecture during the post-injection period (09:00–16:00 hours), including conversions between S (NREM sleep), W (wakefulness), and R (REM sleep) stages (C), episode numbers of sleep–wake stages (D), number of NREM and REM sleep bouts with different duration (E, G), and mean duration of sleep–wake stages (F). (H, I) Average slow-wave activity (SWA) of NREM sleep (H) and electroencephalogram (EEG) power of REM sleep in the frequency range of 6–10 Hz (I) during the post-injection period (09:00–16:00 hours) following saline and CNO injection. *<i>p</i> < 0.05, **<i>p</i> < 0.01 versus saline by paired <i>t</i> tests. CNO (0.3 mg/kg) was given by intraperitoneal (IP) injection at 09:00 hours (<i>n</i> = 9). The horizontal open and filled bars on the <i>x</i>-axes indicate the 12-hour light period and the 12-hour dark period, respectively. Underlying data can be found in <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.2002909#pbio.2002909.s001" target="_blank">S1 Data</a>.</p