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•A new freely available ImageJ-based plugin called DiAna is presented.•DiAna proposes two 3D procedures for image segmentation.•DiAna proposes 3D automated object-based ...co-localization analysis.•We introduce a novel method for statistical significance of object-based co-localization.•DiAna has extended functions for 3D distance analysis and 3D measurements.
We present a new plugin for ImageJ called DiAna, for Distance Analysis, which comes with a user-friendly interface. DiAna proposes robust and accurate 3D segmentation for object extraction. The plugin performs automated object-based co-localization and distance analysis. DiAna offers an in-depth analysis of co-localization between objects and retrieves 3D measurements including co-localizing volumes and surfaces of contact. It also computes the distribution of distances between objects in 3D. With DiAna, we furthermore introduce an original method, which allows for estimating the statistical significance of object co-localization. DiAna offers a complete and intuitive 3D image analysis tool for biologists.
Huntington's disease is an autosomal dominant neurodegenerative disease caused by abnormal polyglutamine expansion in huntingtin (Exp-HTT) leading to degeneration of striatal neurons. Altered brain ...cholesterol homeostasis has been implicated in Huntington's disease, with increased accumulation of cholesterol in striatal neurons yet reduced levels of cholesterol metabolic precursors. To elucidate these two seemingly opposing dysregulations, we investigated the expression of cholesterol 24-hydroxylase (CYP46A1), the neuronal-specific and rate-limiting enzyme for cholesterol conversion to 24S-hydroxycholesterol (24S-OHC). CYP46A1 protein levels were decreased in the putamen, but not cerebral cortex samples, of post-mortem Huntington's disease patients when compared to controls. Cyp46A1 mRNA and CYP46A1 protein levels were also decreased in the striatum of the R6/2 Huntington's disease mouse model and in SThdhQ111 cell lines. In vivo, in a wild-type context, knocking down CYP46A1 expression in the striatum, via an adeno-associated virus-mediated delivery of selective shCYP46A1, reproduced the Huntington's disease phenotype, with spontaneous striatal neuron degeneration and motor deficits, as assessed by rotarod. In vitro, CYP46A1 restoration protected SThdhQ111 and Exp-HTT-expressing striatal neurons in culture from cell death. In the R6/2 Huntington's disease mouse model, adeno-associated virus-mediated delivery of CYP46A1 into the striatum decreased neuronal atrophy, decreased the number, intensity level and size of Exp-HTT aggregates and improved motor deficits, as assessed by rotarod and clasping behavioural tests. Adeno-associated virus-CYP46A1 infection in R6/2 mice also restored levels of cholesterol and lanosterol and increased levels of desmosterol. In vitro, lanosterol and desmosterol were found to protect striatal neurons expressing Exp-HTT from death. We conclude that restoring CYP46A1 activity in the striatum promises a new therapeutic approach in Huntington's disease.
Dysfunctions in brain cholesterol homeostasis have been extensively related to brain disorders. The main pathway for brain cholesterol elimination is its hydroxylation into 24S-hydroxycholesterol by ...the cholesterol 24-hydrolase, CYP46A1. Increasing evidence suggests that CYP46A1 has a role in the pathogenesis and progression of neurodegenerative disorders, and that increasing its levels in the brain is neuroprotective. However, the mechanisms underlying this neuroprotection remain to be fully understood. Huntington's disease is a fatal autosomal dominant neurodegenerative disease caused by an abnormal CAG expansion in huntingtin's gene. Among the multiple cellular and molecular dysfunctions caused by this mutation, altered brain cholesterol homeostasis has been described in patients and animal models as a critical event in Huntington's disease. Here, we demonstrate that a gene therapy approach based on the delivery of CYP46A1, the rate-limiting enzyme for cholesterol degradation in the brain, has a long-lasting neuroprotective effect in Huntington's disease and counteracts multiple detrimental effects of the mutated huntingtin. In zQ175 Huntington's disease knock-in mice, CYP46A1 prevented neuronal dysfunctions and restored cholesterol homeostasis. These events were associated to a specific striatal transcriptomic signature that compensates for multiple mHTT-induced dysfunctions. We thus explored the mechanisms for these compensations and showed an improvement of synaptic activity and connectivity along with the stimulation of the proteasome and autophagy machineries, which participate to the clearance of mutant huntingtin (mHTT) aggregates. Furthermore, BDNF vesicle axonal transport and TrkB endosome trafficking were restored in a cellular model of Huntington's disease. These results highlight the large-scale beneficial effect of restoring cholesterol homeostasis in neurodegenerative diseases and give new opportunities for developing innovative disease-modifying strategies in Huntington's disease.
The vagus nerve serves as an interoceptive relay between the body and the brain. Despite its well-established role in feeding behaviors, energy metabolism, and cognitive functions, the intricate ...functional processes linking the vagus nerve to the hippocampus and its contribution to learning and memory dynamics remain still elusive.
Here, we investigated whether and how the gut-brain vagal axis contributes to hippocampal learning and memory processes at behavioral, functional, cellular, and molecular levels. Our results indicate that the integrity of the vagal axis is essential for long-term recognition memories, while sparing other forms of memory. In addition, by combing multi-scale approaches, our findings show that the gut-brain vagal tone exerts a permissive role in scaling intracellular signaling events, gene expressions, hippocampal dendritic spines density as well as functional long-term plasticities (LTD and LTP). These results highlight the critical role of the gut-brain vagal axis in maintaining the spontaneous and homeostatic functions of hippocampal ensembles and in regulating their learning and memory functions.
In conclusion, our study provides comprehensive insights into the multifaceted involvement of the gut-brain vagal axis in shaping time-dependent hippocampal learning and memory dynamics. Understanding the mechanisms underlying this interoceptive body-brain neuronal communication may pave the way for novel therapeutic approaches in conditions associated with cognitive decline, including neurodegenerative disorders.
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•The gut-brain vagal axis contributes to long-term recognition memories.•The gut-brain vagal axis is dispensable for short-term memories.•The vagal axis regulates molecular and signaling dynamics in the hippocampus.•The gut-brain vagal tone shapes the structural density of hippocampal dendritic spines.•The gut-brain vagal tone ensures physiological forms of synaptic plasticity (LTD and LTP).
Monoaminergic modulation of cortical and thalamic inputs to the dorsal striatum (DS) is crucial for reward-based learning and action control. While dopamine has been extensively investigated in this ...context, the synaptic effects of serotonin (5-HT) have been largely unexplored. Here, we investigated how serotonergic signaling affects associative plasticity at glutamatergic synapses on the striatal projection neurons of the direct pathway (dSPNs). Combining chemogenetic and optogenetic approaches reveals that impeding serotonergic signaling preferentially gates spike-timing-dependent long-term depression (t-LTD) at thalamostriatal synapses. This t-LTD requires dampened activity of the 5-HT4 receptor subtype, which we demonstrate controls dendritic Ca2+ signals by regulating BK channel activity, and which preferentially localizes at the dendritic shaft. The synaptic effects of 5-HT signaling at thalamostriatal inputs provide insights into how changes in serotonergic levels associated with behavioral states or pathology affect striatal-dependent processes.
•5-HT signaling controls t-LTD in striatal projection neurons of the direct pathway•5-HT signaling shapes dendritic Ca2+ signals•5-HT-mediated regulation of t-LTD biases thalamostriatal synapses•Gating of t-LTD requires dampened activity of the 5-HT4R subtype, which modulates BK channel function
Cavaccini and Gritti et al. combine chemogenetic and optogenetic approaches to show that serotonergic signaling provides a control mechanism of synaptic plasticity at thalamic inputs to the striatum.
Repeated cocaine exposure produces new spine formation in striatal projection neurons (SPNs) of the nucleus accumbens. However, an acute exposure to cocaine can trigger long-lasting synaptic ...plasticity in SPNs leading to behavioral alterations. This raises the intriguing question as to whether a single administration of cocaine could enduringly modify striatal connectivity.
A three-dimensional morphometric analysis of presynaptic glutamatergic boutons and dendritic spines was performed on SPNs 1 hour and 1 week after a single cocaine administration. Time-lapse two-photon microscopy in adult slices was used to determine the precise molecular-events sequence responsible for the rapid spine formation.
A single injection triggered a rapid synaptogenesis and persistent increase in glutamatergic connectivity in SPNs from the shell part of the nucleus accumbens, specifically. Synapse formation occurred through clustered growth of active spines contacting pre-existing axonal boutons. Spine growth required extracellular signal-regulated kinase activation, while spine stabilization involved transcription-independent protein synthesis driven by mitogen-activated protein kinase interacting kinase-1, downstream from extracellular signal-regulated kinase. The maintenance of new spines driven by mitogen-activated protein kinase interacting kinase-1 was essential for long-term connectivity changes induced by cocaine in vivo.
Our study originally demonstrates that a single administration of cocaine is able to induce stable synaptic rewiring in the nucleus accumbens, which will likely influence responses to subsequent drug exposure. It also unravels a new functional role for cocaine-induced extracellular signal-regulated kinase pathway independently of nuclear targets. Finally, it reveals that mitogen-activated protein kinase interacting kinase-1 has a pivotal role in cocaine-induced connectivity.
Background Activation of the extracellular signal-regulated kinase (ERK) in the striatum is crucial for long-term behavioral alterations induced by drugs of abuse. In response to cocaine, ERK ...phosphorylation (i.e., activation) is restricted to medium-sized spiny neurons expressing dopamine D1 receptor (D1R) and depends on a concomitant stimulation of D1R and glutamate N -methyl-D-aspartate receptor (NMDAR). However, the mechanisms responsible for this activation, especially the respective contribution of D1R and NMDAR, remain unknown. Methods We studied striatal neurons in culture stimulated with D1R agonist and/or glutamate and wild-type or genetically modified mice treated with cocaine. Biochemical, immunohistochemical, and imaging studies were performed. Mice were also subjected to behavioral experiments. Results Stimulation of D1R cannot activate ERK by itself but potentiates glutamate-mediated calcium influx through NMDAR that is responsible for ERK activation. Potentiation of NMDAR by D1R depends on a cyclic adenosine monophosphate–independent signaling pathway, which involves tyrosine phosphorylation of the NR2B subunit of NMDAR by Src family kinases. We also demonstrate that the D1R/Src family kinases/NR2B pathway is responsible for ERK activation by cocaine in vivo. Inhibition of this pathway abrogates cocaine-induced locomotor sensitization and conditioned place preference. Conclusions Our results show that potentiation of NR2B-containing NMDAR by D1R is necessary and sufficient to trigger cocaine-induced ERK activation. They highlight a new cyclic adenosine monophosphate–independent pathway responsible for the integration of dopamine and glutamate signals by the ERK cascade in the striatum and for long-term behavioral alterations induced by cocaine.
Resolution, high signal intensity and elevated signal to noise ratio (SNR) are key issues for biologists who aim at studying the localisation of biological structures at the cellular and subcellular ...levels using confocal microscopy. The resolution required to separate sub-cellular biological structures is often near to the resolving power of the microscope. When optimally used, confocal microscopes may reach resolutions of 180 nm laterally and 500 nm axially, however, axial resolution in depth is often impaired by spherical aberration that may occur due to refractive index mismatches. Spherical aberration results in broadening of the point-spread function (PSF), a decrease in peak signal intensity when imaging in depth and a focal shift that leads to the distortion of the image along the z-axis and thus in a scaling error. In this study, we use the novel mounting medium CFM3 (Citifluor Ltd., UK) with a refractive index of 1.518 to minimize the effects of spherical aberration. This mounting medium is compatible with most common fluorochromes and fluorescent proteins. We compare its performance with established mounting media, harbouring refractive indices below 1.500, by estimating lateral and axial resolution with sub-resolution fluorescent beads. We show furthermore that the use of the high refractive index media renders the tissue transparent and improves considerably the axial resolution and imaging depth in immuno-labelled or fluorescent protein labelled fixed mouse brain tissue. We thus propose to use those novel high refractive index mounting media, whenever optimal axial resolution is required.
Drug addiction is defined as a compulsive pattern of drug-seeking- and taking- behavior, with recurrent episodes of abstinence and relapse, and a loss of control despite negative consequences. ...Addictive drugs promote reinforcement by increasing dopamine in the mesocorticolimbic system, which alters excitatory glutamate transmission within the reward circuitry, thereby hijacking reward processing. Within the reward circuitry, the striatum is a key target structure of drugs of abuse since it is at the crossroad of converging glutamate inputs from limbic, thalamic and cortical regions, encoding components of drug-associated stimuli and environment, and dopamine that mediates reward prediction error and incentive values. These signals are integrated by medium-sized spiny neurons (MSN), which receive glutamate and dopamine axons converging onto their dendritic spines. MSN primarily form two mostly distinct populations based on the expression of either DA-D1 (D1R) or DA-D2 (D2R) receptors. While a classical view is that the two MSN populations act in parallel, playing antagonistic functional roles, the picture seems much more complex. Herein, we review recent studies, based on the use of cell-type-specific manipulations, demonstrating that dopamine differentially modulates dendritic spine density and synapse formation, as well as glutamate transmission, at specific inputs projecting onto D1R-MSN and D2R-MSN to shape persistent pathological behavioral in response to drugs of abuse. We also discuss the identification of distinct molecular events underlying the detrimental interplay between dopamine and glutamate signaling in D1R-MSN and D2R-MSN and highlight the relevance of such cell-type-specific molecular studies for the development of innovative strategies with potential therapeutic value for addiction. Because drug addiction is highly prevalent in patients with other psychiatric disorders when compared to the general population, we last discuss the hypothesis that shared cellular and molecular adaptations within common circuits could explain the co-occurrence of addiction and depression. We will therefore conclude this review by examining how the nucleus accumbens (NAc) could constitute a key interface between addiction and depression.
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