Cannabinoids Induce Cell Death and Promote P2X7 Receptor Signaling in Retinal Glial Progenitors in Culture

Freitas, Hércules Rezende; Isaac, Alinny Rosendo; Silva, Thayane Martins; Diniz, Geyzzara Oliveira Ferreira; Dabdab, Yara dos Santos; Bockmann, Eduardo Cosendey; Guimarães, Marília Zaluar P.; Calaza, Karin da Costa; Mello, Fernando G. de; Ventura, Ana Lúcia Marques; Reis, Ricardo Augusto de Melo; França, Guilherme Rapozeiro

doi:10.1007/s12035-019-1537-y

Cited by 12 publications

(24 citation statements)

References 82 publications

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“…In total, 607 cells were analyzed from 30- and 45-day cerebral organoids, in which 245 cells (40%) were responsive to 50 mM KCl, while only 5 cells (1%) were activated by 100 μM ATP. The area under the curve was quantified for those cultured flat cells (Figures 5A,M), or neuron-like cells (Figures 5G,N), which responded to 100 μM ATP or 50 mM KCl, as previously done (Freitas et al, 2019). This percentage agrees with the previously addressed protein content, which indicated the presence of more neuronal than glial cells in organoids at this differentiation stage, several of which are young and still unresponsive.…”

Section: Resultsmentioning

confidence: 99%

Human Cerebral Organoids and Fetal Brain Tissue Share Proteomic Similarities

Nascimento

Saia-Cereda

Sartore

et al. 2019

Front. Cell Dev. Biol.

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The limited access to functional human brain tissue has led to the development of stem cell-based alternative models. The differentiation of human pluripotent stem cells into cerebral organoids with self-organized architecture has created novel opportunities to study the early stages of the human cerebral formation. Here we applied state-of-the-art label-free shotgun proteomics to compare the proteome of stem cell-derived cerebral organoids to the human fetal brain. We identified 3,073 proteins associated with different developmental stages, from neural progenitors to neurons, astrocytes, or oligodendrocytes. The major protein groups are associated with neurogenesis, axon guidance, synaptogenesis, and cortical brain development. Glial cell proteins related to cell growth and maintenance, energy metabolism, cell communication, and signaling were also described. Our data support the variety of cells and neural network functional pathways observed within cell-derived cerebral organoids, confirming their usefulness as an alternative model. The characterization of brain organoid proteome is key to explore, in a dish, atypical and disrupted processes during brain development or neurodevelopmental, neurodegenerative, and neuropsychiatric diseases.

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Section: Resultsmentioning

confidence: 99%

Human Cerebral Organoids and Fetal Brain Tissue Share Proteomic Similarities

Nascimento

Saia-Cereda

Sartore

et al. 2019

Front. Cell Dev. Biol.

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“…Recent data have shown that progenitors from the embryonic avian retina unresponsive to ATP ( Figure 2A ) differentiate into neurons (activated by KCl or glutamate) or glia (activated by ATP) in a cannabinoid-rich environment ( Figure 2B ). Calcium shifts induced by ATP in avian Müller glia increased up to 30% in early retinal cells when cannabinoid receptors CB1 and CB2 are activated by the cannabinoid agonist WIN 55,212-2 ( Figure 2C ) ( Freitas et al, 2019 ). Alternatively, a decrease in the number of glutamate-, GABA, and KCl-responsive cells (neurons and progenitors) was reported.…”

Section: Retina a Central Modelmentioning

confidence: 99%

“…Emerging calcium shifts induced by ATP are exclusive to Müller glia (green trace), primed by the early cannabinoid receptor activation (with WIN 55,212-2). On the other hand, neurons are activated by KCl and glutamate (red trace) ( Freitas et al, 2019 ) (C) . Instead of differentiating, some progenitors die (D) due to the activation of death-inducing pathways [bottom panel: live (calcein, green) and dead (ethidium homodimer-1, red)].…”

Section: Retina a Central Modelmentioning

confidence: 99%

Cell Calcium Imaging as a Reliable Method to Study Neuron–Glial Circuits

2020

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Complex dynamic cellular networks have been studied in physiological and pathological processes under the light of single-cell calcium imaging (SCCI), a method that correlates functional data based on calcium shifts operated by different intracellular and extracellular mechanisms integrated with their cell phenotypes. From the classic synaptic structure to tripartite astrocytic model or the recent quadripartite microglia added ensemble, as well as other physiological tissues, it is possible to follow how cells signal spatiotemporally to cellular patterns. This methodology has been used broadly due to the universal properties of calcium as a second messenger. In general, at least two types of receptor operate through calcium permeation: a fast-acting ionotropic receptor channel and a slow-activating metabotropic receptor, added to exchangers/transporters/pumps and intracellular Ca 2+ release activated by messengers. These prototypes have gained an enormous amount of information in dynamic signaling circuits. SCCI has also been used as a method to associate phenotypic markers during development and stage transitions in progenitors, stem, vascular cells, neuro- and glioblasts, neurons, astrocytes, oligodendrocytes, and microglia that operate through ion channels, transporters, and receptors. Also, cancer cells or inducible cell lines from human organoids characterized by transition stages are currently being used to model diseases or reconfigure healthy cells in terms of the expression of calcium-binding/permeable molecules and shed light on therapy.

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“…Increased differentiation might be assessed in the future not only by analyzing the voltage channels after ATP stimulation, but also by measuring the response of Cannabinoid, P2 £ 7 or glycine receptors. 71,72 Even if we observed neural electrophysiological features, the expression of the pan-neuronal marker TUJ1 together with the development of a stable resting membrane potential and soma polarization suggest that the differentiation towards a neuronal phenotype has initiated, but is not yet completed after 48h. Glycosylation levels can change in the neural stem cells along with the fate conversion, and glycosylation can prime the astroglia towards a ganglion cell fate.…”

Section: Discussionmentioning

confidence: 84%

Section: Discussionmentioning

confidence: 99%