Inhibiting EPAC1 improves brain edema and neurological outcomes by regulating AQP4 polarization after intracerebral hemorrhage

Intracerebral hemorrhage (ICH) is a severe cerebrovascular event associated with high mortality and disability, primarily due to perihematomal edema (PHE). Aquaporin-4 (AQP4) polarization plays a crucial role in PHE development; however, effective therapeutic strategies remain elusive. This study investigates the mechanisms regulating AQP4 polarization and brain edema in a murine ICH model, with autologous blood injected into the striatum of adult C57BL/6 J mice. Three approaches were employed: (1) a time-course analysis (0, 24, 48, and 72 h post-ICH) to assess brain edema and AQP4 localization via magnetic resonance imaging (MRI), electron microscopy, and immunofluorescence; (2) pharmacological modulation of EPAC1 using an inhibitor (ESI-09, 10 mg/kg, intraperitoneal) and an activator (8-CPT, 20 μM, intravenous) to evaluate its role in AQP4 polarization; (3) lentiviral knockdown of AQP4 to investigate its interaction with EPAC1. Results showed that AQP4 underwent partial depolarization, characterized by reduced AQP4-M23 expression and decreased colocalization with CD31, coinciding with significant brain edema at 72 h post-ICH. Proteomic and network analyses, utilizing AlphaFold for protein structure prediction, identified exchange protein directly activated by cyclic adenosine monophosphate 1 (EPAC1) as a key regulator of AQP4 polarization, with its interaction with AQP4 markedly reduced following ICH. Pharmacological inhibition of EPAC1 with ESI-09 mitigated AQP4 depolarization, whereas activation with 8-CPT exacerbated it. Moreover, AQP4 knockdown attenuated the protective effects of EPAC1 inhibition, leading to aggravated brain edema and worsened neurological deficits. These findings suggest that targeting EPAC1-mediated AQP4 polarization may provide a novel therapeutic strategy for ICH.