Purpose: Inhibition of the BKCa channel attenuated the nitric oxide-induced increase in outflow facility and decrease in trabecular meshwork (TM) cell volume suggesting the involvement of the BKCa channel in TM cell function. This study examined the effects of activation of the BK Ca channel on outflow facility and TM cell volume and determined if the effects of NO and BKCa channel activation on TM cell volume were additive. Methods: Porcine eyes were used to measure outflow facility using the anterior segment organ culture perfusion system. Cell volume was measured using Calcein AM fluorescent dye, detected by confocal microscopy, and quantified using NIH ImageJ software. Results: NS1619 increased outflow facility 86% over baseline. Additionally, there was a concentration-dependent decrease in TM cell volume in response to NS1619, which was abolished by iberiotoxin (IBTX). While NS1619 alone and DETA-NO alone decreased TM cell volume, together their effects were not additive. The time course for NS1619-induced increases in outflow facility correlated with the time course for NS1619-induced decreases in cell volume. Conclusions: BKCa channel activation increases outflow facility and decreases cell volume suggesting that K+ efflux regulates TM cell function.