Microglial Calcium Waves During the Hyperacute Phase of Ischemic Stroke

Abstract

Background and Purpose: Ischemic injury triggers multiple pathological responses in the brain tissue, including spreading depolarizations across the cerebral cortex (cortical spreading depolarizations [CSD]). Microglia have been recently shown to play a significant role in the propagation of CSD. However, the intracellular responses of myeloid cells during ischemic stroke have not been investigated. Methods: We have studied intracellular calcium activity in cortical microglia in the stroke model of the middle cerebral artery occlusion, using the murine Polr2a-based and Cre-dependent GCaMP5 and tdTomato reporter (PC::G5-tdT). High-speed 2-photon microscopy through cranial windows was employed to record signals from genetically encoded indicators of calcium. Inflammatory stimuli and pharmacological inhibition were used to modulate microglial calcium responses in the somatosensory cortex. Results: In vivo imaging revealed periodical calcium activity in microglia during the hyperacute phase of ischemic stroke. This activity was more frequent during the first 6 hours after occlusion, but the amplitudes of calcium transients became larger at later time points. Consistent with CSD nature of these events, we reproducibly triggered comparable calcium transients with microinjections of potassium chloride (KCl) into adjacent cortical areas. Furthermore, lipopolysaccharide-induced peripheral inflammation, mimicking sterile inflammation during ischemic stroke, produced significantly greater microglial calcium transients during CSD. Finally, in vivo pharmacological analysis with CRAC (calcium release-activated channel) inhibitor CM-EX-137 demonstrated that CSD-associated microglial calcium transients after KCl microinjections are mediated at least in part by the CRAC mechanism. Conclusions: Our findings demonstrate that microglia participate in ischemic brain injury via previously undetected mechanisms, which may provide new avenues for therapeutic interventions.

Mathematical modelling of human P2X-mediated plasma membrane electrophysiology and calcium dynamics in microglia

“…Activation of P2X 4 R and P2X 7 R results in intricate kinks for 1mM and 3mM of ATP in rat microglia [ 38 ]. Other work on microglia and astrocytes reveal kink-style behaviour of [ 16 , 45 – 47 ] and such dynamics are also observed in other animal and human cell lines including those experimentally reported in [ 48 – 53 ]. Note that no kink appears for ATP < 1mM because the initial rise in current is dominated by current flow through the P2X 7 .…”

“…Microglia are involved in many neurodegenerative disorders, such as Parkinson’s and Alzheimer’s diseases, and are massively activated during brain injuries such as stroke [ 3 , 15 , 16 ]. There are many known hurdles to overcome in the study of microglia for drug discovery strategies [ 17 – 19 ].…”

Microglia sense neuronal activity via GABA in the early postnatal hippocampus

“…It also became evident that only a subpopulation of microglia responds to a given neurotransmitter, implying a large heterogeneity in their chemosensitivity. Recent evidence suggests that neuronal activity is sensed by microglia (Umpierre et al, 2020;Umpierre and Wu, 2020;Liu et al, 2021), raising the possibility that intrinsic neurotransmitter receptors enable microglia to sense synaptic activity. An experimental approach characterizing microglial Ca 2+ responses triggered by neurotransmitters in situ and in vivo would require an experimental model that robustly monitors microglial Ca 2+ elevations, such as a mouse model expressing a genetically encoded Ca 2+ indicator (GECI) specifically in microglia at developmental and adult stages.…”

Brain acute ischemia mechanisms: implications to experimental and clinical treatment

“…Классическое объяснение медленных волн деполяризации включает в себя выход ионов калия из разрушенных клеток и выделение эксайтотоксичных медиаторов [46]. Распространение волн медленной деполяризации захватывает и глиальные клетки, вызывая в них опустошение кальциевых депо, однако последствия их участия пока точно не изучены [59]. В результате нарушения регуляции депо астроциты могут достаточно активно выделять кальций в межклеточное пространство [60].…”