Features of the in vitro expression profile of hippocampal neurogenic niche cells during optogenetic stimulation

Abstract

In the central nervous system of mammals, there are specialized areas in which neurogenesis — neurogenic niches — is observed in the postnatal period. It is believed that astrocytes in the composition of neurogenic niches play a significant role in the regulation of neurogenesis, and therefore they are considered as a promising “target” for the possible control of neurogenesis, including the use of optogenetics. In the framework of this work, we formed an in vitro model of a neurogenic niche, consisting of cerebral endothelial cells, astrocytes and neurospheres. Astrocytes in the neurogenic niche model expressed canalorodopsin ChR2 and underwent photoactivation. The effect of photoactivated astrocytes on the expression profile of neurogenic niche cells was evaluated using immunocytochemical analysis methods. It was found that intact astrocytes in the composition of the neurogenic niche contribute to neuronal differentiation of stem cells, as well as the activation of astroglia expressing photosensitive proteins, changes the expression of molecules characterized by intercellular interactions of pools of resting and proliferating cells in the composition of the neurogenic niche with the participation of NAD+ (Cx43, CD38, CD157), lactate (MCT1). In particular, the registered changes reflect a violation of the paracrine intercellular interactions of two subpopulations of cells, one of which acts as a source of NAD+, and the second as a consumer of NAD+ to ensure the processes of intracellular signal transduction; a change in the mechanisms of lactate transport due to aberrant expression of the lactate transporter MCT1 in cells forming a pool of cells developing along the neuronal path of differentiation. In general, with photostimulation of niche astrocytes, the total proliferative activity increases mainly due to neural progenitor cells, but not neural stem cells. Thus, optogenetic activation of astrocytes can become a promising tool for controlling the activity of neurogenesis processes and the formation of a local proneurogenic microenvironment in an in vitro model of a neurogenic niche.

Current progress and challenges in the development of brain tissue models: How to grow up the changeable brain in vitro?

“…Even simple static models are still useful for studying intercellular communications in the brain tissue. 17 , 28 , 219 , 263 – 265 Application of microfluidic systems opens new opportunities for the establishment of changeable microenvironment, reconstruction of brain fluids flow and cells behaviors that are controlled by fluids movement and their chemical composition (e.g. fluidic shear stress enabling BMECs to develop and to function with higher efficacy), achievement of better barrier structural integrity as well as continuous monitoring of brain cells metabolism and performing the high throughput high content imaging and analysis.…”

“…7. In addition to all above mentioned characteristics, the models aimed to reproduce key events in neurogenesis and neurogenesis-coupled brain plasticity should consider the following additional tasks: 7.1. establishment of multicellular ensembles made of NSCs/NPCs and their progeny as well as fully-differentiated cells (astrocytes, BMECs, CPECs, EpCs), tight integration of functionally and phenotypically distinct compartments (neurogenic niche, NVU/brain parenchyma, and associated brain tissue barriers) on a chip mimicking the regulation of neurogenesis in (patho)physiological conditions 31 , 135 , 281 – 283 ; 7.2. reproduction of ECM composition, oxygen and nutrients supply for better cell viability and functionality, reproduction of metabolic plasticity and local humoral microenvironment supporting cell-to-cell communications, establishment of conditions supportive for neurogenesis and angiogenesis 204 , 284 – 286 ; 7.3. reconstitution of the chip microarchitecture, fluids exchange and establishment of chemical gradients that are supportive for NSCs/NPCs maintenance due to dynamic changes in the concentrations of local and “systemic” regulatory molecules (neurotransmitters, gliotransmitters, growth factors, cytokines, alarmins, metabolites) produced by the cells themselves, infused into the microfluidic device artificially, or embedded into the ECM and scaffolds 13 , 287 ; 7.4. achieving the controllable and reproducible recruitment, proliferation, differentiation, apoptosis of NSCs/NPCs, migration of neuroblasts, maturation of newly-formed neurons and their functional integration within the NVU/brain parenchyma compartment 242 , 244 , 247 , 264 , 288 , 289 ; 7.5. establishment of the molecular machinery responsible for dynamic/phasic changes in the tissue (for instance, circadian/diurnal rhythms and/or mitochondrial dynamics) that are important for determining the stem cells fate or barrier functions 136 , 290 ; 7.6. achievement of prolonged viability of the neurogenic niche in vitro model enabling long-lasting recording of “developmental” and plastic changes in the brain tissue. …”

“…7.4. achieving the controllable and reproducible recruitment, proliferation, differentiation, apoptosis of NSCs/NPCs, migration of neuroblasts, maturation of newly-formed neurons and their functional integration within the NVU/brain parenchyma compartment 242 , 244 , 247 , 264 , 288 , 289 ;…”

Blood–Brain Barrier and Neurovascular Unit In Vitro Models for Studying Mitochondria-Driven Molecular Mechanisms of Neurodegeneration

“…Using BMECs but not ECs of other origin as a part of the models is more physiological to reproduce the main mechanisms of BBB development and functioning. As we have shown before, specific metabolic properties of BMECs makes them an interesting object for managing the barrier permeability in BBB models, or even in tissue models based on the cerebral endothelial monolayer (i.e., neurogenic niche in vitro model) [ 123 , 139 , 140 , 141 , 142 , 143 , 144 ].…”