METH-induced, TAAR1-associated CREB signaling serves as a master regulator for astrocyte EAAT-2

Methamphetamine (METH) abuse accelerates the onset and severity of HIV-associated neurocognitive disorders (HAND) and astrocyte-mediated excitotoxicity. METH targets several receptors, particularly astrocyte trace amine associated receptor 1 (TAAR1), as we have previously reported. Molecular alterations of astrocyte TAAR1 correspond to changes in astrocyte excitatory amino acid transporter-2 (EAAT-2) levels and function; however, the signaling pathways downstream of METH-induced TAAR1 activation remain unclear. Astrocyte EAAT-2 is tightly regulated at the transcriptional and translational levels by cAMP and calcium, yet METH-mediated increases in these second messengers have not been shown to directly modulate astrocyte EAAT-2. Furthermore, HIV-1 relevant stimuli and IL-1b, increase TAAR1 and may exacerbate METH-mediated excitotoxicity via MAPK/ERK and NF-kB. We propose CREB activation serves as a master regulator of astrocyte EAAT-2. To investigate the temporal order of CREB activation we utilized genetically encoded calcium indicators, or GCaMPs, to visualize and quantify METH-induced calcium signaling. RNA interference targeting PKA and NF-kB subunit p65, in addition to PKA and MAPK/ERK specific inhibitors support their involvement in astrocyte EAAT-2 regulation. Furthermore, we investigated CREB phosphorylation at serine 133/142, the co-activator and co-repressor forms, respectively, following METH-induced activation. Overall, this work identifies critical signaling pathways and therapeutic targets for astrocyte EAAT-2 recovery.