HDAC3 regulates microglial proliferation after ischemic stroke. Microglia, the brain's resident immune cells, become activated following stroke, contributing to neuroinflammation and neuronal loss. This study shows that HDAC3 deficiency in microglia reduces their proliferation, particularly of proinflammatory microglia, and prevents their transition to a proinflammatory state. HDAC3 knockout (HDAC3-miKO) mice showed improved long-term functional and histological outcomes after stroke. RNA-seq and ATAC-seq analyses revealed that HDAC3 deficiency affects mitosis and chromatin accessibility, with PU.1 acting as a downstream mediator of HDAC3's effects on microglial proliferation and polarization. Overexpression of PU.1 in microglia reversed the protective effects of HDAC3-miKO, indicating PU.1's role in HDAC3-mediated stroke outcomes. These findings suggest that the HDAC3/PU.1 axis regulates microglial proliferation and polarization, influencing the inflammatory state post-stroke and contributing to stroke pathophysiology. HDAC3 deficiency selectively inhibits the proliferation of proinflammatory microglia, while anti-inflammatory microglia remain unaffected. This selective regulation highlights the importance of microglial cell cycle control in stroke progression and opens new avenues for therapeutic interventions.HDAC3 regulates microglial proliferation after ischemic stroke. Microglia, the brain's resident immune cells, become activated following stroke, contributing to neuroinflammation and neuronal loss. This study shows that HDAC3 deficiency in microglia reduces their proliferation, particularly of proinflammatory microglia, and prevents their transition to a proinflammatory state. HDAC3 knockout (HDAC3-miKO) mice showed improved long-term functional and histological outcomes after stroke. RNA-seq and ATAC-seq analyses revealed that HDAC3 deficiency affects mitosis and chromatin accessibility, with PU.1 acting as a downstream mediator of HDAC3's effects on microglial proliferation and polarization. Overexpression of PU.1 in microglia reversed the protective effects of HDAC3-miKO, indicating PU.1's role in HDAC3-mediated stroke outcomes. These findings suggest that the HDAC3/PU.1 axis regulates microglial proliferation and polarization, influencing the inflammatory state post-stroke and contributing to stroke pathophysiology. HDAC3 deficiency selectively inhibits the proliferation of proinflammatory microglia, while anti-inflammatory microglia remain unaffected. This selective regulation highlights the importance of microglial cell cycle control in stroke progression and opens new avenues for therapeutic interventions.