Ca²⁺transients in astrocyte fine processes occur via Ca²⁺influx in the adult mouse hippocampus

Abstract

Astrocytes display complex morphologies with an array of fine extensions extending from the soma and the primary thick processes. Until the use of genetically encoded calcium indicators (GECIs) selectively expressed in astrocytes, Ca signaling was only examined in soma and thick primary processes of astrocytes where Ca -sensitive fluorescent dyes could be imaged. GECI imaging in astrocytes revealed a previously unsuspected pattern of spontaneous Ca transients in fine processes that has not been observed without chronic expression of GECIs, raising potential concerns about the effects of GECI expression. Here, we perform two-photon imaging of Ca transients in adult CA1 hippocampal astrocytes using a new single-cell patch-loading strategy to image Ca -sensitive fluorescent dyes in the cytoplasm of fine processes. We observed that astrocyte fine processes exhibited a high frequency of spontaneous Ca transients whereas astrocyte soma rarely showed spontaneous Ca oscillations similar to previous reports using GECIs. We exploited this new approach to show these signals were independent of neuronal spiking, metabotropic glutamate receptor (mGluR) activity, TRPA1 channels, and L- or T-type voltage-gated calcium channels. Removal of extracellular Ca almost completely and reversibly abolished the spontaneous signals while IP R2 KO mice also exhibited spontaneous and compartmentalized signals, suggesting they rely on influx of extracellular Ca . The Ca influx dependency of the spontaneous signals in patch-loaded astrocytes was also observed in astrocytes expressing GCaMP3, further highlighting the presence of Ca influx pathways in astrocytes. The mechanisms underlying these localized Ca signals are critical for understanding how astrocytes regulate important functions in the adult brain. GLIA 2016;64:2093-2103.

Disentangling astroglial physiology with a realistic cell model in silico

“…The underlying molecular machinery appears to engage Ca 2+ -dependent Ca 2+ release controlled by inositol 1,4,5-trisphosphate (IP 3 ) and possibly ryanodine receptor-channels displaying a highly non-linear (sometimes bell-shaped) dependence between channel activity and Ca 2+ concentration 50 . Recently, local Ca 2+ transients (but not global waves) have been documented in astroglia lacking IP 3 receptors 51,52 . In any such cases, intracellular Ca 2+ signal propagation must be tightly controlled by the local Ca 2+ buffering capacity 53 , the feature comprehensively explored in nerve cells.…”

Transient Opening of the Mitochondrial Permeability Transition Pore Induces Microdomain Calcium Transients in Astrocyte Processes

“…Indeed, interstitial cells of Cajal (ICC) in the gastrointestinal tract and submucosal cells in the colon exhibit rhythmic, spontaneous Ca 2+ oscillations generated by Ca 2+ exchange between ER Ca 2+ stores and mitochondria (Ward et al, 2000; Yoneda et al, 2002) that are blocked when the electrochemical gradient across the inner mitochondrial membrane or IP 3 -dependent release of Ca 2+ from ER stores are inhibited (Ward et al, 2000). Recent results suggest that activity in some astrocyte microdomains is dependent on extracellular Ca 2+ influx (Rungta et al, 2016; Srinivasan et al, 2015), suggesting that mitochondrial Ca 2+ is subject to homeostatic control or that there are also plasma membrane channels/transporters in these domains that enable Ca 2+ influx independent of mitochondria.…”

Astrocyte Calcium Responses to Sensory Input: Influence of Circuit Organization and Experimental Factors

“…These essential functions involve activation of calcium signaling pathways within the cytosol of the astrocyte, which can be activated intrinsically (Nett et al, 2002; Srinivasan et al, 2015; Rungta et al, 2016), or in response to neuronal activity (Di Castro et al, 2011; Haustein et al, 2014). Defining the quantitative relationship between neuronal activity and astrocyte calcium signaling has proven difficult.…”