Stroke remains a leading cause of disability in the United States. Despite recent advances, interventions to reduce damage and enhance recovery after
stroke are lacking. P2X4R, a receptor for
adenosine triphosphate (
ATP), regulates activation of myeloid immune cells (infiltrating monocytes/macrophages and brain-resident microglia) after
stroke injury. However, over-stimulation of P2X4Rs due to excessive
ATP release from dying or damaged neuronal cells can contribute to ischemic injury. Therefore, we pharmacologically inhibited P2X4R to limit the over-stimulated myeloid cell immune response and improve both acute and chronic
stroke recovery. We subjected 8-12-week-old male and female wild type mice to a 60 min right
middle cerebral artery occlusion (MCAo) followed by 3 or 30 days of reperfusion. We performed histological,
RNA sequencing, behavioral (sensorimotor, anxiety, and depressive), and biochemical (
Evans blue dye extravasation, western blot, quantitative PCR, and flow cytometry) analyses to determine the acute (3 days after MCAo) and chronic (30 days after MCAo) effects of P2X4R antagonist 5-BDBD (1 mg/kg P.O. daily x 3 days post 4 h of MCAo) treatment. 5-BDBD treatment significantly (p < .05) reduced
infarct volume, neurological deficit (ND) score, levels of
cytokine interleukin-1 beta (IL-1β) and blood brain barrier (BBB) permeability in the 3-day group. Chronically, 5-BDBD treatment also conferred progressive recovery (p < .05) of motor balance and coordination using a rotarod test, as well as reduced anxiety-like behavior over 30 days. Interestingly, depressive-type behavior was not observed in mice treated with 5-BDBD for 3 days. In addition, flow cytometric analysis revealed that 5-BDBD treatment decreased the total number of infiltrated leukocytes, and among those infiltrated leukocytes, pro-inflammatory cells of myeloid origin were specifically reduced. 5-BDBD treatment reduced the cell surface expression of P2X4R in flow cytometry-sorted monocytes and microglia without reducing the total P2X4R level in brain tissue. In summary, acute P2X4R inhibition protects against ischemic injury at both acute and chronic time-points after
stroke. Reduced numbers of infiltrating pro-inflammatory myeloid cells, decreased surface P2X4R expression, and reduced BBB disruption are likely its mechanism of neuroprotection and neuro-rehabilitation.