Selective Kv1.3 inhibition promotes M2 microglial polarization, accelerates hematoma resolution, and improves neurological recovery following collagenase-induced spontaneous intracerebral hemorrhage in mice.

Abstract

Background and aim: Spontaneous intracerebral hemorrhage (sICH) is a devastating subtype of stroke characterized by high mortality, severe neurological disability, and limited effective pharmacological treatment. Secondary brain injury following sICH is largely driven by excessive neuroinflammation, in which activated microglia predominantly adopt a pro-inflammatory (M1-like) phenotype that exacerbates neuronal damage. The voltage-gated potassium channel Kv1.3 is highly expressed in activated microglia and represents a promising therapeutic target for modulating neuroinflammatory responses. This study investigated whether selective Kv1.3 inhibition with 5-(4-phenoxybutoxy) psoralen (PAP-1) could promote immuno-modulatory microglial polarization, attenuate brain injury, and improve neurological recovery in a collagenase-induced mouse model of sICH.

Materials and methods: Adult male ICR mice with collagenase-induced sICH received daily intraperitoneal administration of PAP-1 (40 mg/kg) or vehicle for up to 7 days. Survival, body weight, hematoma volume, neuronal survival, Kv1.3 expression, and microglial polarization were evaluated using histological and immunohistochemical analyses. Neurological recovery was assessed using the modified neurological severity score, cylinder test, corner turn test, and open field test. Histological analyses were performed on post-injury days 1, 2, 3, and 7, whereas behavioral assessments and survival monitoring were conducted longitudinally throughout the experimental period.

Results: Selective Kv1.3 inhibition significantly improved neurological recovery following sICH. PAP-1 treatment increased 7-day survival from 40% to 70% and significantly reduced hematoma volume from day 2 onward, with reductions of approximately 19% on days 2 and 3 and 24% on day 7 compared with vehicle-treated mice. PAP-1 preserved perihematomal neurons, producing an approximately 2.9-fold increase in NeuN-positive neuronal density on day 7. Furthermore, PAP-1 suppressed Kv1.3 expression, reduced CD16/32-positive M1-like microglia, enhanced CD206-positive M2-like microglial polarization beginning on day 1, increased the M2/M1 ratio, and significantly improved sensorimotor and locomotor performance across all behavioral assessments. These findings demonstrate that selective Kv1.3 inhibition effectively attenuated neuroinflammation, accelerated hematoma resolution, and promoted functional recovery after experimental sICH.

Conclusion: Selective inhibition of Kv1.3 with PAP-1 promoted a favorable shift toward an immunomodulatory microglial phenotype, reduced hematoma burden, preserved perihematomal neurons, and accelerated neurological recovery following collagenase-induced sICH. The integration of histological, immunological, and functional outcomes provides strong evidence supporting Kv1.3 as a promising therapeutic target for early intervention after hemorrhagic stroke. Although the study was limited to male mice, was restricted to a 7-day observation period, and lacked detailed downstream mechanistic analyses, the findings establish a robust preclinical foundation for future investigations into long-term functional recovery, molecular signaling pathways, and translational evaluation of Kv1.3-targeted therapies in sICH.