Tubuloglomerular feedback (TGF) is the mechanism by which the macula densa (MD) senses increases in luminal NaCl concentration and sends a signal to constrict the afferent arteriole (Af-Art). The kidney expresses constitutively heme oxygenase-2 (HO-2), and low levels of HO-1. HOs release carbon monoxide (CO), biliverdin and free iron. We hypothesized that renal HOs inhibit TGF via release of CO and biliverdin. Rabbit Af-Arts and attached MD were simultaneously microperfused in vitro. TGF response was determined by measuring Af-Art diameter before and after increasing NaCl in the MD perfusate. When HO activity was inhibited by adding stannous mesoporphyrin (SnMP) to the MD perfusate, TGF response increased from 2.1±0.2 µm to 4.1±0.4 µm (p = 0.003, control vs SnMP, n = 7). When a CO-releasing molecule, CORM-3 (50 µM) was added to the MD perfusate, TGF response decreased by 41%, from 3.6±0.3 µm to 2.1±0.2 µm (p< 0.001, control vs CORM-3, n = 12). When CORM-3 at 100 µM was added to the perfusate, it completely blocked TGF response, from 4.2±0.4 µm to -0.2±0.3 µm (p < 0.001, control vs CORM-3, n = 6). When biliverdin was added to the perfusate, TGF response decreased by 79%, from 3.4±0.3 µm to 0.7±0.4 µm (p = 0.001, control vs biliverdin, n = 6). The effects of SnMP and CORM-3 were not blocked by inhibition of nitric oxide synthase. We concluded that renal HO inhibits TGF probably via release of CO and biliverdin. HO regulation of TGF is a novel mechanism that could lead to a better understanding of the control of renal microcirculation and function.