Activation of Type 1 Cannabinoid Receptor (CB1R) promotes neurogenesis in murine subventricular zone cell cultures

The endocannabinoid system has been implicated in the modulation of adult neurogenesis. Here, we describe the effect of type 1 cannabinoid receptor (CB1R) activation on self-renewal, proliferation and neuronal differentiation in mouse neonatal subventricular zone (SVZ) stem/progenitor cell cultures. Expression of CB1R was detected in SVZ-derived immature cells (Nestin-positive), neurons and astrocytes. Stimulation of the CB1R by (R)-(+)-Methanandamide (R-m-AEA) increased self-renewal of SVZ cells, as assessed by counting the number of secondary neurospheres and the number of Sox2+/+ cell pairs, an effect blocked by Notch pathway inhibition. Moreover, R-m-AEA treatment for 48 h, increased proliferation as assessed by BrdU incorporation assay, an effect mediated by activation of MAPK-ERK and AKT pathways. Surprisingly, stimulation of CB1R by R-m-AEA also promoted neuronal differentiation (without affecting glial differentiation), at 7 days, as shown by counting the number of NeuN-positive neurons in the cultures. Moreover, by monitoring intracellular calcium concentrations ([Ca2+](i)) in single cells following KCl and histamine stimuli, a method that allows the functional evaluation of neuronal differentiation, we observed an increase in neuronal-like cells. This proneurogenic effect was blocked when SVZ cells were co-incubated with R-m-AEA and the CB1R antagonist AM 251, for 7 days, thus indicating that this effect involves CB1R activation. In accordance with an effect on neuronal differentiation and maturation, R-m-AEA also increased neurite growth, as evaluated by quantifying and measuring the number of MAP2-positive processes. Taken together, these results demonstrate that CB1R activation induces proliferation, self-renewal and neuronal differentiation from mouse neonatal SVZ cell cultures.Fundacao para a Ciencia e a Tecnologia - Portugal [POCTI/SAU-NEU/68465/2006, PTDC/SAU-NEU/104415/2008, PTDC/SAU-NEU/101783/2008, POCTI/SAU-NEU/110838/2009]; Fundacao Calouste Gulbenkian [96542]; Fundacao para a Ciencia e Tecnologiainfo:eu-repo/semantics/publishedVersio

Ceruloplasmin Protects Against Rotenone-Induced Oxidative Stress and Neurotoxicity

To clarify the neuroprotective property of ceruloplasmin and the pathogenesis of aceruloplasminemia, we generated ceruloplasmin-deficient (CP−/−) mice on the C57BL/10 genetic background and further treated them with a mitochondrial complex I inhibitor, rotenone. There was no iron accumulation in the brains of CP−/− mice at least up to 60 weeks of age. Without rotenone treatment, CP−/− mice showed slight motor dysfunction compared with CP+/+ mice, but there were no detectable differences in the levels of oxidative stress markers between these two groups. A low dose of rotenone did not affect the mitochondrial complex I activity in our mice, however, it caused a significant change in motor behavior, neuropathology, or the levels of oxidative stress markers in CP−/− mice, but not in CP+/+ mice. Our data support that ceruloplasmin protects against rotenone-induced oxidative stress and neurotoxicity, probably through its antioxidant properties independently of its function of iron metabolism

Abnormal Brain Iron Homeostasis in Human and Animal Prion Disorders

Neurotoxicity in all prion disorders is believed to result from the accumulation of PrP-scrapie (PrPSc), a β-sheet rich isoform of a normal cell-surface glycoprotein, the prion protein (PrPC). Limited reports suggest imbalance of brain iron homeostasis as a significant associated cause of neurotoxicity in prion-infected cell and mouse models. However, systematic studies on the generality of this phenomenon and the underlying mechanism(s) leading to iron dyshomeostasis in diseased brains are lacking. In this report, we demonstrate that prion disease–affected human, hamster, and mouse brains show increased total and redox-active Fe (II) iron, and a paradoxical increase in major iron uptake proteins transferrin (Tf) and transferrin receptor (TfR) at the end stage of disease. Furthermore, examination of scrapie-inoculated hamster brains at different timepoints following infection shows increased levels of Tf with time, suggesting increasing iron deficiency with disease progression. Sporadic Creutzfeldt-Jakob disease (sCJD)–affected human brains show a similar increase in total iron and a direct correlation between PrP and Tf levels, implicating PrPSc as the underlying cause of iron deficiency. Increased binding of Tf to the cerebellar Purkinje cell neurons of sCJD brains further indicates upregulation of TfR and a phenotype of neuronal iron deficiency in diseased brains despite increased iron levels. The likely cause of this phenotype is sequestration of iron in brain ferritin that becomes detergent-insoluble in PrPSc-infected cell lines and sCJD brain homogenates. These results suggest that sequestration of iron in PrPSc–ferritin complexes induces a state of iron bio-insufficiency in prion disease–affected brains, resulting in increased uptake and a state of iron dyshomeostasis. An additional unexpected observation is the resistance of Tf to digestion by proteinase-K, providing a reliable marker for iron levels in postmortem human brains. These data implicate redox-iron in prion disease–associated neurotoxicity, a novel observation with significant implications for prion disease pathogenesis

Developmental consequences of perinatal cannabis exposure: behavioral and neuroendocrine effects in adult rodents

Cannabis is the most commonly used illicit drug among pregnant women. Since the endocannabinoid system plays a crucial role in brain development, maternal exposure to cannabis derivatives might result in long-lasting neurobehavioral abnormalities in the exposed offspring. It is difficult to detect these effects, and their underlying neurobiological mechanisms, in clinical cohorts, because of their intrinsic methodological and interpretative issues. The present paper reviews relevant rodent studies examining the long-term behavioral consequences of exposure to cannabinoid compounds during pregnancy and/or lactation. Maternal exposure to even low doses of cannabinoid compounds results in atypical locomotor activity, cognitive impairments, altered emotional behavior, and enhanced sensitivity to drugs of abuse in the adult rodent offspring. Some of the observed behavioral abnormalities might be related to alterations in stress hormone levels induced by maternal cannabis exposure. There is increasing evidence from animal studies showing that cannabinoid drugs are neuroteratogens which induce enduring neurobehavioral abnormalities in the exposed offspring. Several preclinical findings reviewed in this paper are in line with clinical studies reporting hyperactivity, cognitive impairments and altered emotionality in humans exposed in utero to cannabis. Conversely, genetic, environmental and social factors could also influence the neurobiological effects of early cannabis exposure in humans

Interleukin-6 promoter polymorphism interacts with pain and life stress influencing depression phenotypes

Interleukin-6 (IL-6) has emerged as a potent biomarker for depression as its elevated plasma levels in patients with clinical depression have been confirmed by meta-analyses. Increased plasma IL-6 concentration was associated with various psychological stress factors and physical disorders accompanied by pain. Another modulator of the IL-6 level is rs1800795, a promoter polymorphism in the IL-6 gene which is able to influence its expression rate. Therefore, we examined in a Hungarian population sample of 1053 volunteers with European origins if rs1800795 polymorphism can affect depression symptoms measured by Zung Self-rating Depression Scale (ZSDS), and Brief Symptom Inventory (BSI). We also investigated the interactions of the polymorphism with reported painful physical conditions and Recent Negative Life Events (RLE) measured by the List of Life Threatening Experiences. Rs1800795 significantly interacted with both RLE and painful condition on depressive symptoms measured by ZSDS and BSI using different heritability models, while no main effects of the polymorphism were identified. After correction for multiple testing only the rs1800795 x RLE interaction effect (recessive model) remained significant on the BSI score, while both RLE and painful conditions significantly interacted on the ZSDS. In conclusion, the functional IL-6 rs1800795 polymorphism in interaction with various stress factors increases the risk of depression and has a greater impact on symptoms measured by the ZSDS. Thus, IL-6 and other cytokines may be more relevant in the development of somatic symptoms compared to affective signs of depression, delineating a specific genotype-phenotype relationship in this heterogeneous disorder

Paradoxical Role of Prion Protein Aggregates in Redox-Iron Induced Toxicity

Imbalance of iron homeostasis has been reported in sporadic Creutzfeldt-Jakob-disease (sCJD) affected human and scrapie infected animal brains, but the contribution of this phenotype to disease associated neurotoxicity is unclear.Using cell models of familial prion disorders, we demonstrate that exposure of cells expressing normal prion protein (PrP(C)) or mutant PrP forms to a source of redox-iron induces aggregation of PrP(C) and specific mutant PrP forms. Initially this response is cytoprotective, but becomes increasingly toxic with time due to accumulation of PrP-ferritin aggregates. Mutant PrP forms that do not aggregate are not cytoprotective, and cells show signs of acute toxicity. Intracellular PrP-ferritin aggregates induce the expression of LC3-II, indicating stimulation of autophagy in these cells. Similar observations are noted in sCJD and scrapie infected hamster brains, lending credence to these results. Furthermore, phagocytosis of PrP-ferritin aggregates by astrocytes is cytoprotective, while culture in astrocyte conditioned medium (CM) shows no measurable effect. Exposure to H(2)O(2), on the other hand, does not cause aggregation of PrP, and cells show acute toxicity that is alleviated by CM.These observations suggest that aggregation of PrP in response to redox-iron is cytoprotective. However, subsequent co-aggregation of PrP with ferritin induces intracellular toxicity unless the aggregates are degraded by autophagosomes or phagocytosed by adjacent scavenger cells. H(2)O(2), on the other hand, does not cause aggregation of PrP, and induces toxicity through extra-cellular free radicals. Together with previous observations demonstrating imbalance of iron homeostasis in prion disease affected brains, these observations provide insight into the mechanism of neurotoxicity by redox-iron, and the role of PrP in this process

Role of the Cellular Prion Protein in Oligodendrocyte Precursor Cell Proliferation and Differentiation in the Developing and Adult Mouse CNS

There are numerous studies describing the signaling mechanisms that mediate oligodendrocyte precursor cell (OPC) proliferation and differentiation, although the contribution of the cellular prion protein (PrPc) to this process remains unclear. PrPc is a glycosyl-phosphatidylinositol (GPI)-anchored glycoprotein involved in diverse cellular processes during the development and maturation of the mammalian central nervous system (CNS). Here we describe how PrPc influences oligodendrocyte proliferation in the developing and adult CNS. OPCs that lack PrPc proliferate more vigorously at the expense of a delay in differentiation, which correlates with changes in the expression of oligodendrocyte lineage markers. In addition, numerous NG2-positive cells were observed in cortical regions of adult PrPc knockout mice, although no significant changes in myelination can be seen, probably due to the death of surplus cells

Prolonged oral cannabinoid administration prevents neuroinflammation, lowers β-amyloid levels and improves cognitive performance in Tg APP 2576 mice

Background: Alzheimer’s disease (AD) brain shows an ongoing inflammatory condition and non-steroidal antiinflammatories diminish the risk of suffering the neurologic disease. Cannabinoids are neuroprotective and antiinflammatory agents with therapeutic potential. Methods: We have studied the effects of prolonged oral administration of transgenic amyloid precursor protein (APP) mice with two pharmacologically different cannabinoids (WIN 55,212-2 and JWH-133, 0.2 mg/kg/day in the drinking water during 4 months) on inflammatory and cognitive parameters, and on 18F-fluoro-deoxyglucose (18FDG) uptake by positron emission tomography (PET). Results: Novel object recognition was significantly reduced in 11 month old Tg APP mice and 4 month administration of JWH was able to normalize this cognitive deficit, although WIN was ineffective. Wild type mice cognitive performance was unaltered by cannabinoid administration. Tg APP mice showed decreased 18FDG uptake in hippocampus and cortical regions, which was counteracted by oral JWH treatment. Hippocampal GFAP immunoreactivity and cortical protein expression was unaffected by genotype or treatment. In contrast, the density of Iba1 positive microglia was increased in Tg APP mice, and normalized following JWH chronic treatment. Both cannabinoids were effective at reducing the enhancement of COX-2 protein levels and TNF-a mRNA expression found in the AD model. Increased cortical b-amyloid (Ab) levels were significantly reduced in the mouse model by both cannabinoids. Noteworthy both cannabinoids enhanced Ab transport across choroid plexus cells in vitro. Conclusions: In summary we have shown that chronically administered cannabinoid showed marked beneficial effects concomitant with inflammation reduction and increased Ab clearanceThis work was supported by the Spanish Ministry of Science and Technology (SAF 2005-02845 to M.L.C). A.M.M-M. was recipient a fellowship from the Ministry of Education and Scienc

The endocannabinoid system in mental disorders: Evidence from human brain studies

Mental disorders have a high prevalence compared with many other health conditions and are the leading cause of disability worldwide. Several studies performed in the last years support the involvement of the endocannabinoid system in the etiopathogenesis of different mental disorders. The present review will summarize the latest information on the role of the endocannabinoid system in psychiatric disorders, specifically depression, anxiety, and schizophrenia. We will focus on the findings from human brain studies regarding alterations in endocannabinoid levels, cannabinoid receptors and endocannabinoid metabolizing enzymes in patients suffering mental disorders. Studies carried out in humans have consistently demonstrated that the endocannabinoid system is fundamental for emotional homeostasis and cognitive function. Thus, deregulation of the different elements that are part of the endocannabinoid system may contribute to the pathophysiology of several mental disorders. However, the results reported are controversial. In this sense, different alterations in gene and/or protein expression of CB1 receptors have been shown depending on the technical approach used or the brain region studied. Despite the current discrepancies regarding cannabinoid receptors changes in depression and schizophrenia, present findings point to the endocannabinoid system as a pivotal neuromodulatory pathway relevant in the pathophysiology of mental disorders.This study was supported by the Spanish Ministry of Economy and Competitiveness (SAF2015-67457-R, MINECO/FEDER), the Plan Estatal de I+D+i 2013-2016, the Instituto de Salud Carlos III-Subdirección General de Evaluación y Fomento de la Investigación, Spanish Ministry of Economy, FEDER (PI13/01529) and the Basque Government (IT616/13). I I-L is a recipient of a Predoctoral Fellowship from the Basque Government. E F-Z is a recipient of a Predoctoral Fellowship from the University of Cantabria. CM is a recipient of a Postdoctoral Marie Skłodowska-Curie Individual Fellowship (H2020-MSCA-IF-2016, ID 747487)

Whole organisms or pure compounds? entourage effect versus drug specificity

As the therapeutic use of sacred plants and fungi becomes increasingly accepted by Western medicine, a tug of war has been taking place between those who advocate the traditional consumption of whole organisms and those who defend exclusively the utilization of purified compounds. The attempt to reduce organisms to single active principles is challenged by the sheer complexity of traditional medicine. Ayahuasca, for example, is a concoction of at least two plant species containing multiple psychoactive substances with complex interactions. Similarly, cannabis contains dozens of psychoactive substances whose specific combinations in different strains correspond to different types of therapeutic and cognitive effects. The “entourage effect” refers to the synergistic effects of the multiple compounds present in whole organisms, which may potentiate clinical efficacy while attenuating side effects. In opposition to this view, mainstream pharmacology is adamant about the need to use purified substances, presumably more specific and safe. In this chapter, I will review the evidence on both sides to discuss the scientific, economic, and political implications of this controversy. The evidence indicates that it is time to embrace the therapeutic complexity of psychedelics.2019-07-3