Unraveling ChR2-driven stochastic Ca2+ dynamics in astrocytes – A call for new interventional paradigms

Abstract Control of astrocytes via modulation of Ca2+ oscillations using techniques like optogenetics can prove to be crucial in therapeutic intervention of a variety of neurological disorders. However, a systematic study quantifying the effect of optogenetic stimulation in astrocytes is yet to be performed. Here, we propose a novel stochastic Ca2+dynamics model that incorporates the light sensitive component – channelrhodopsin 2 (ChR2). Utilizing this model, we studied the effect of various pulsed light stimulation paradigms on astrocytes for select variants of ChR2 (wild type, ChETA, and ChRET/TC) in both an individual and a network of cells. Our results exhibited a consistent pattern of Ca2+ activity among individual cells in response to optogenetic stimulation, i.e., showing steady state regimes with increased Ca2+ basal level and Ca2+ spiking probability. Furthermore, we performed a global sensitivity analysis to assess the effect of stochasticity and variation of model parameters on astrocytic Ca2+ dynamics in the presence and absence of light stimulation, respectively. Results indicated that directing variants towards the first open state of the photo-cycle of ChR2 (o1) enhances spiking activity in astrocytes during optical stimulation. Evaluation of the effect of astrocytic ChR2 expression (heterogeneity) on Ca2+ signaling revealed that the optimal stimulation paradigm of a network does not necessarily coincide with that of an individual cell. Simulation for ChETA-incorporated astrocytes suggest that maximal activity of a single cell reduced the spiking probability of the network of astrocytes at higher degrees of ChR2 expression efficiency due to an elevation of basal Ca2+ beyond physiological levels. Collectively, the framework presented in this study provides valuable information for the selection of light stimulation paradigms that elicit optimal astrocytic activity using existing ChR2 constructs, as well as aids in the engineering of future optogenetic constructs. Author summary Optogenetics – an avant-garde technique involves targeted delivery of light sensitive ion channels to cells. Channelrhodopsin 2 (ChR2), an algal derived light sensitive ion channel has extensively been used in neuroscience to manipulate various cell types in a guided and controlled manner. Despite being predominantly used in neurons, recent advancements have led to the expansion of the application of optogenetics in non-neuronal cell types, like astrocytes. These cells play a key role in various aspects of the central nervous system and alteration of their signaling is associated with various disorders, including epilepsy, stroke and Alzheimer’s disease. Hence, invaluable information for therapeutic intervention can be obtained from using optogenetics to regulate astrocytic activity in a strategic manner. Here, we propose a novel computational model to assess astrocytic response to optogenetic stimulation which implicitly accounts for the stochastic character of Ca2+ signaling in this cell type. We identified light stimulation paradigms suitable for eliciting astrocytic Ca2+ response within physiological levels in widely-used ChR2 variants and identified highly sensitive parameters in ChR2 kinetics conducive for higher probability in Ca2+ spiking. Overall, the results of this model can be used to boost astrocyte light-induced behavior prediction and the development of improved future optogenetic constructs.