Orai, RyR, and IP3R channels cooperatively regulate calcium signaling in brain mid-capillary pericytes

Abstract

Pericytes are multifunctional cells of the vasculature that are vital to brain homeostasis, yet many of their fundamental physiological properties, such as Ca2+ signaling pathways, remain unexplored. We performed pharmacological and ion substitution experiments to investigate the mechanisms underlying pericyte Ca2+ signaling in acute cortical brain slices of PDGFRβ-Cre::GCaMP6f mice. We report that mid-capillary pericyte Ca2+ signalling differs from ensheathing type pericytes in that it is largely independent of L- and T-type voltage-gated calcium channels. Instead, Ca2+ signals in mid-capillary pericytes were inhibited by multiple Orai channel blockers, which also inhibited Ca2+ entry triggered by endoplasmic reticulum (ER) store depletion. An investigation into store release pathways indicated that Ca2+ transients in mid-capillary pericytes occur through a combination of IP3R and RyR activation, and that Orai store-operated calcium entry (SOCE) is required to sustain and amplify intracellular Ca2+ increases evoked by the GqGPCR agonist endothelin-1. These results suggest that Ca2+ influx via Orai channels reciprocally regulates IP3R and RyR release pathways in the ER, which together generate spontaneous Ca2+ transients and amplify Gq-coupled Ca2+ elevations in mid-capillary pericytes. Thus, SOCE is a major regulator of pericyte Ca2+ and a target for manipulating their function in health and disease.

L-Type Ca2+channels and TRPC3 channels shape brain pericyte Ca2+signaling and hemodynamics throughout the arteriole to capillary networkin vivo

“…S2C, D) or in vivo (Fig. 5E), similar to a recent report in brain slices (26). However, Pyr3 reduced the amplitude of Ca 2+ events within the soma of capillary pericytes in brain slices (Fig.…”

“…GCaMP6s positive capillary pericytes were identified more than 5 branches from the penetrating arteriole. Recent studies of capillary pericyte Ca 2+ events in brain slices have found conflicting effects of L-type Ca 2+ channel blockers and no effect of Pyr3 on Ca 2+ event frequency (25,26). Therefore, we conducted both brain slice pharmacology experiments with nimodipine (1 µM) and Pyr3 (3 µM; Fig.…”

“…Both ensheathing and capillary pericytes express a broad range of Ca 2+ channels and vasomodulator receptors that can trigger fluctuations in intracellular Ca 2+ . Elevated intracellular Ca 2+ in ensheathing pericytes causes cellular contraction (25,26), and this is highly mediated by L-type VGCC activity (7,15,22,26). In distal capillary pericytes, the activity of VGCC appears to be less robust than in ensheathing pericyte s, since VGCC blockers mildly reduce or do not affect spontaneous Ca 2+ signaling in capillary pericytes in brain slices (25,26), and these channels are not activated during pressure-induced capillary constriction unlike upstream ensheathing pericytes and smooth muscle cells (22).…”

“…Elevated intracellular Ca 2+ in ensheathing pericytes causes cellular contraction (25,26), and this is highly mediated by L-type VGCC activity (7,15,22,26). In distal capillary pericytes, the activity of VGCC appears to be less robust than in ensheathing pericyte s, since VGCC blockers mildly reduce or do not affect spontaneous Ca 2+ signaling in capillary pericytes in brain slices (25,26), and these channels are not activated during pressure-induced capillary constriction unlike upstream ensheathing pericytes and smooth muscle cells (22). However, VGCC are reportedly expressed by capillary pericytes (27,28), and the contribution of these channels to capillary pericyte Ca 2+ signaling and cortical hemodynamics in vivo remains unknown.…”

“…However, VGCC are reportedly expressed by capillary pericytes (27,28), and the contribution of these channels to capillary pericyte Ca 2+ signaling and cortical hemodynamics in vivo remains unknown. Other Ca 2+ channels also contribute to capillary pericyte Ca 2+ events, such as store operated Ca 2+ entry channels (26) and transient receptor potential (TRP) channels (25,26). Specifically, TRPC3 channels are expressed in brain pericytes as determined by transcriptomics (27), but their protein expression, contribution to Ca 2+ signaling in the pericyte subtypes, and influence on blood vessel hemodynamics have not been confirmed.…”