TRPC1‐ and TRPC3‐dependent Ca²⁺ signaling in mouse cortical astrocytes affects injury‐evoked astrogliosis in vivo

Abstract

Following brain injury astrocytes change into a reactive state, proliferate and grow into the site of lesion, a process called astrogliosis, initiated and regulated by changes in cytoplasmic Ca2+. Transient receptor potential canonical (TRPC) channels may contribute to Ca2+ influx but their presence and possible function in astrocytes is not known. By RT-PCR and RNA sequencing we identified transcripts of Trpc1, Trpc2, Trpc3 and Trpc4 in FACS-sorted glutamate aspartate transporter (GLAST)-positive cultured mouse cortical astrocytes and subcloned full-length Trpc1 and Trpc3 cDNAs from these cells. Ca2+ entry in cortical astrocytes depended on TRPC3 and was increased in the absence of Trpc1. After co-expression of Trpc1 and Trpc3 in HEK-293 cells both proteins co-immunoprecipitate and form functional heteromeric channels, with TRPC1 reducing TRPC3 activity. In vitro, lack of Trpc3 reduced astrocyte proliferation and migration whereas the TRPC3 gain-of-function moonwalker mutation and Trpc1 deficiency increased astrocyte migration. In vivo, astrogliosis and cortex edema following stab wound injury were reduced in Trpc3−/− but increased in Trpc1−/− mice. In summary, our results show a decisive contribution of TRPC3 to astrocyte Ca2+ signaling, which is even augmented in the absence of Trpc1, in particular following brain injury. Targeted therapies to reduce TRPC3 channel activity in astrocytes might therefore be beneficial in traumatic brain injury.

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“…As the primer sequences and lengths for TRPC1 and TRPC2 are similar (171 bp and 173 bp, respectively), we assumed that the downregulation of TPRC2 may lead to the growth of astrocytes. 65 However, further studies need to be conducted to show the other signal pathways that are involved in this mechanism.…”

The role of TRP channels in white matter function and ischaemia

“…In line with this, TRPC3 inhibitors reduce SOCE in spinal astrocytes [23]. However in another study, knockout of TRPC1 or TRPC3 and TRPC6 did not affect SOCE in cortical astrocytes [20], and TRPC appears to have no role in SOCE in primary microglia [32]. Thus, the mechanisms responsible for SOCE in glial cells may be more complicated than supposed, and may differ between cell types [28].…”

“…found to be expressed by astrocytes [17,[20][21][22][23], and TRPC3 [24] and TRPC6 [25] by microglia (reviewed by [26]). TRPC channels have also been found in the oligodendrocyte lineage, where TRPC1-6 are more highly expressed by OPCs and newly forming oligodendrocytes, and TRPC7 has greater expression in some mature oligodendrocyte populations [12].…”

“…Along with mediating SOCE in astrocytes, TRPC1 has the potential to regulate a huge variety of functions and to damage cells if they are excessively activated. Thus, astrocytes with TRPC1 genetically removed display increased migration in vitro and have increased injury-evoked astrogliosis in vivo [20]. Blocking TRPC1 with antibodies reduces glutamate release from cultured astrocytes [17].…”

“…However in Trpc2 -/-mice, both males and females mount other mice indiscriminately (reviewed by [33]). Only very low levels of TRPC2 have been found in astrocytes [20,22], microglia [32] and OPCs [12].…”