Molecular approaches for manipulating astrocytic signaling in vivo

Astrocytes are the predominant glial type in the central nervous system and play important roles in assisting neuronal function and network activity. Astrocytes exhibit complex signaling systems that are essential for their normal function and the homeostasis of the neural network. Altered signaling in astrocytes is closely associated with neurological and psychiatric diseases, suggesting tremendous therapeutic potential of these cells. To further understand astrocyte function in health and disease, it is important to study astrocytic signaling in vivo. In this review, we discuss molecular tools that enable the selective manipulation of astrocytic signaling, including the tools to selectively activate and inactivate astrocyte signaling in vivo. Lastly, we highlight a few tools in development that present strong potential for advancing our understanding of the role of astrocytes in physiology, behavior, and pathology

Challenging the Dogma: Reevaluating the Role of Astrocyte Calcium Signaling in Physiology

Gliotransmission represents one of the most important conceptual shifts in neuroscience in the past several decades. Gliotransmission refers to the process whereby glial cells release specific neurotransmitters that modulate synaptic transmission via pre- and postsynaptic activation of G-protein coupled (GPCR) and ionotropic receptors. Astrocytes release glutamate, ATP or D-serine upon activation of a wide range of Gq-GPCRs to modulate neuronal activity in the numerous regions of the brain. These "gliotransmitters" modulate spontaneous and evoked neuronal activity at both excitatory and inhibitory synapses, and affect heterosynaptic depression and long term potentiation in the hippocampus. The theory of astrocyte Ca2+-dependent modulation of neuronal activity is built upon the hypothesis that activation of Gq-GPCRs on astrocytes leads to Inositol trisphosphate (IP3) receptor-mediated calcium increases that trigger the release of gliotransmitters. However, evidence supporting this hypothesis has primarily relied on non-physiological methods for increasing or decreasing glial Ca2+. For example, uncaging of Ca2+ or chelation of intracellular Ca2+ in astrocytes affects synaptic transmission. However the usage of these methods does not specifically target IP3 receptor-mediated Ca2+ increases, considered the primary source of Ca2+ changes in astrocytes for gliotransmission. To probe the role of IP3 receptors (IP3R) in Ca2+-dependent release of gliotransmitters from astrocytes, we have used the IP3R type 2 (IP3R2) knockout mouse model. Through a combination of Ca2+ imaging and electrophysiology experiments we found that deletion of IP3R2 in astrocytes blocks intracellular Ca2+ increases in astrocytes in response to Gq GCPR activation. Further, that lack of IP3R-mediated Ca2+ increases does not affect excitatory synaptic transmission of both CA1 and CA3 pyramidal neurons in the hippocampus. Analysis of IP3R2 conditional knockout mice reveals specific behavioral changes in acoustic startle response and spatial learning in the Morris Water Maze. These novel findings represent a departure from the established theory of gliotransmission and are a significant step forward in our understanding of the role of astrocytic IP3R-mediated Ca2+ increases in physiology

Loss of IP3 Receptor-Dependent Ca2+ Increases in Hippocampal Astrocytes Does Not Affect Baseline CA1 Pyramidal Neuron Synaptic Activity

Astrocytes in the hippocampus release calcium (Ca2+) from intracellular stores intrinsically and in response to activation of Gq-linked G-protein coupled receptors (GPCRs) through the binding of inositol 1,4,5-trisphosphate (IP3) to its receptor (IP3R). Astrocyte Ca2+ has been deemed necessary and sufficient to trigger the release of gliotransmitters, such as ATP and glutamate, from astrocytes to modulate neuronal activity. Several lines of evidence suggest that IP3R Type 2 (IP3R2) is the primary IP3R expressed by astrocytes. In order to determine if IP3R2 is the primary functional IP3R responsible for astrocytic Ca2+ increases, we conducted experiments using an IP3R2 knockout mouse model (IP3R2 KO). We show for the first time that lack of IP3R2 blocks both spontaneous and Gq-linked GPCR mediated increases in astrocyte Ca2+. Furthermore, neuronal Gq-linked GPCR Ca2+ increases remain intact, suggesting that IP3R2 does not play a major functional role in neuronal calcium store release or may not be expressed in neurons. Additionally, we show that lack of IP3R2 in the hippocampus does not affect baseline excitatory neuronal synaptic activity as measured by spontaneous EPSC (sEPSC) recordings from CA1 pyramidal neurons. Whole cell recordings of the tonic NMDA receptor (NMDA-R) mediated current indicates that ambient glutamate levels are also unaffected in the IP3R2 KO. These data show that IP3R2 is the key functional IP3R driving Gq-linked GPCR mediated Ca2+ increases in hippocampal astrocytes and that removal of astrocyte Ca2+ increases does not significantly affect excitatory neuronal synaptic activity or ambient glutamate levels

Astrocyte IP3R2-dependent Ca2+ signaling is not a major modulator of neuronal pathways governing behavior

Calcium-dependent release of gliotransmitters by astrocytes is reported to play a critical role in synaptic transmission and be necessary for long-term potentiation (LTP), long-term depression (LTD) and other forms of synaptic modulation that are correlates of learning and memory. Further, physiological processes reported to be dependent on Ca2+ fluxes in astrocytes include functional hyperemia, sleep, and regulation of breathing. The preponderance of findings indicate that most, if not all, receptor dependent Ca2+ fluxes within astrocytes are due to release of Ca2+ through IP3 receptor/channels in the endoplasmic reticulum. Findings from several laboratories indicate that astrocytes only express IP3 receptor type 2 (IP3R2) and that a knockout of IP3R2 obliterates the GPCR-dependent astrocytic Ca2+ responses. Assuming that astrocytic Ca2+ fluxes play a critical role in synaptic physiology, it would be predicted that elimination of astrocytic Ca2+ fluxes would lead to marked changes in behavioral tests. Here, we tested this hypothesis by conducting a broad series of behavioral tests that recruited multiple brain regions, on an IP3R2 conditional knockout mouse model. We present the novel finding that behavioral processes are unaffected by lack of astrocyte IP3R-mediated Ca2+ signals. IP3R2 cKO animals display no change in anxiety or depressive behaviors, and no alteration to motor and sensory function. Morris water maze testing, a behavioral correlate of learning and memory, was unaffected by lack of astrocyte IP3R2-mediated Ca2+-signaling. Therefore, in contrast to the prevailing literature, we find that neither receptor-driven astrocyte Ca2+ fluxes nor, by extension, gliotransmission is likely to be a major modulating force on the physiological processes underlying behavior

Functional Roles of Astrocyte Calcium Elevations: From Synapses to Behavior

Astrocytes are fundamental players in the regulation of synaptic transmission and plasticity. They display unique morphological and phenotypical features that allow to monitor and to dynamically respond to changes. One of the hallmarks of the astrocytic response is the generation of calcium elevations, which further affect downstream cellular processes. Technical advances in the field have allowed to spatially and to temporally quantify and qualify these elevations. However, the impact on brain function remains poorly understood. In this review, we discuss evidences of the functional impact of heterogeneous astrocytic calcium events in several brain regions, and their consequences in synapses, circuits, and behavior.Foundation for Science and Technology (FCT) fellowships (SFRH/BPD/97281/2013 to JO, SFRH/BD/101298/2014 to SG-G, IF/00328/2015 to JO, IF/01079/2014 to LP); Marie Curie Fellowship FP7-PEOPLE- 2010-IEF 273936 and BIAL Foundation Grant 207/14 to JO and 427/14 to LP; Northern Portugal Regional Operational Programme (NORTE 2020), under the Portugal 2020 Partnership Agreement, through the European Regional Development Fund (FEDER; NORTE-01-0145-FEDER-000013); FEDER funds, through the Competitiveness Factors Operational Programme (COMPETE), and National Funds, through the FCT (POCI-01- 0145-FEDER-007038)info:eu-repo/semantics/publishedVersio

What Is the Role of Astrocyte Calcium in Neurophysiology?

Astrocytes comprise approximately half of the volume of the adult mammalian brain and are the primary neuronal structural and trophic supportive elements. Astrocytes are organized into distinct nonoverlapping domains and extend elaborate and dense fine processes that interact intimately with synapses and cerebrovasculature. The recognition in the mid 1990s that astrocytes undergo elevations in intracellular calcium concentration following activation of G protein-coupled receptors by synaptically released neurotransmitters demonstrated not only that astrocytes display a form of excitability but also that astrocytes may be active participants in brain information processing. The roles that astrocytic calcium elevations play in neurophysiology and especially in modulation of neuronal activity have been intensely researched in recent years. This review will summarize the current understanding of the function of astrocytic calcium signaling in neurophysiological processes and discuss areas where the role of astrocytes remains controversial and will therefore benefit from further study

Astrocytes: orchestrating synaptic plasticity?

Synaptic plasticity is the capacity of a preexisting connection between two neurons to change in strength as a function of neural activity. Because synaptic plasticity is the major candidate mechanism for learning and memory, the elucidation of its constituting mechanisms is of crucial importance in many aspects of normal and pathological brain function. In particular, a prominent aspect that remains debated is how the plasticity mechanisms, that encompass a broad spectrum of temporal and spatial scales, come to play together in a concerted fashion. Here we review and discuss evidence that pinpoints to a possible non-neuronal, glial candidate for such orchestration: the regulation of synaptic plasticity by astrocytes.Comment: 63 pages, 4 figure