We characterized inward rectifier (K IR ) currents in reactive astrocytes activated by CNS injury. We used primary cultures of reactive astrocytes obtained from gelatin sponge implants in adult rat brains, a system that yielded highly purified, homogeneous cultures with Ͼ95% of cells positive for GFAP, vimentin, and S-100. Ionic channels were studied in 1-21-day-old primary cultures using a nystatin-perforated patch clamp technique. Fast Na ϩ currents were identified in Ͻ2% of cells. Most cells exhibited outward currents positive to Ϫ50 mV, with one component being sensitive to charybdotoxin, iberiotoxin, and tetraethylammonium chloride, and another component being sensitive to 4-aminopyridine. Two populations of cells were distinguished, based on presence or absence of Ba 2ϩ -sensitive K IR current negative to the K ϩ reversal potential (E k ), with Ͼ80% of cells expressing K IR currents. In contrast to previous reports on mammalian astrocytes, the current-voltage curve showed no appreciable current between E k and Ϫ50 mV, reflecting strong rectification by K IR channels. The magnitude of K IR current at Ϫ130 mV (I Ϫ130 ) did not change significantly during 21 days in culture (123 cells), suggesting constitutive expression of K IR channels. The fraction of K IR -negative cells was not affected by serum-starvation for 16-24 h. In cells with I Ϫ130 Ն Ϫ30 pA, the membrane potential was invariably near E k and depolarized appreciably on addition of Ba 2ϩ , but in cells with I Ϫ130 Ͻ Ϫ30 pA, resting potentials ranged from Ϫ40 mV to Ϫ90 mV. We conclude that most adult reactive astrocytes constitutively express K IR channel(s) that exhibit strong rectification not previously observed in mammalian astrocytes.