National and global greenhouse gas (GHG) budgets are continually being refined as data become available. Primary sources of the potent GHG nitrous oxide (N₂O) include agricultural soil management and burning of fossil fuels, but comprehensive N₂O budgets also incorporate less prominent factors such as wetlands. Freshwater wetland GHG flux estimates, however, have high uncertainty, and wetlands have been identified as both sources and sinks. Here, we analyzed a regional database of >26,000 N₂O chamber flux measurements sampled across >150 wetlands from the Prairie Pothole Region (PPR) in the Great Plains of North America. Our goal was to identify important land use and hydrologic drivers of N₂O flux to help reduce uncertainty in N₂O models, and to incorporate these drivers into an upscaled estimate of wetland N₂O emissions from the U.S. portion of the PPR. Within individual wetlands, exposed soils with no standing water, such as along wetland edges, were hotspots that accounted for greater than 90% of wetland N₂O emissions. In contrast wet (i.e., ponded) areas had minimal or negative N₂O flux. N₂O flux from wetlands nested within croplands (16.3–17.3 μg N₂O m⁻² hr⁻¹) was, in some instances, nearly double that from wetlands within grasslands (9.2–14.4 μg N₂O m⁻² h⁻¹). We estimated that seasonal N₂O flux from PPR wetlands equated to roughly 0.2% (1.04 Tg CO₂ equivalents) of the U.S. N₂O budget (c. 2019). Overall, even though PPR wetlands are a small net source of N₂O to the atmosphere, their emissions are negligible relative to agricultural soil management. Policy and management to restore wetland hydrology and surrounding uplands from cropland to grasslands can reduce landscape N₂O fluxes. Future activities focused on wetland N₂O flux would benefit from inclusion of adjacent land use and hydrologic factors, as well as from incorporation of temporally dynamic ponded wetland areas.