River regulation is a key influence of river ecology, with dams, weirs and other impoundments altering natural hydrology, flow volume, velocity, seasonality, frequency and the duration and magnitude of flood events. Regulated flows can result in reduced water quality and habitat heterogeneity, depauperate macroinvertebrate assemblages and have a pivotal influence on periphyton primary producers and other components of river ecology. The release of environmental flows that mimic natural variability downstream of impoundments is often included as a management response aimed at mitigating impacts on river health due to flow regulation. Such flows are likely to be particularly important for rivers within or near dense urbanisation that would experience additional associated stressors compounded by potentially changing climatic conditions. The Coxs River, located in the Greater Blue Mountains Heritage Area, NSW, is one such river. Since the construction of Lake Lyell in 1980s flows on the upper Coxs River have been regulated. In 2011 following a decade of drought induced low flow, greater background flows coinciding with implementation of environmental flows have resulted in much greater flow variability. This has persisted for several years. The requisite ecological monitoring program possibly provides the longest (over 15 years) continuous dataset on the effect of river regulation on aquatic macroinvertebrates and periphyton in Australia. It demonstrates how several macroinvertebrate and periphyton taxa have responded to the return of natural conditions in the river following several low flow years. The availability of reference data from downstream of the influence of regulated flows provides a comparative measure of natural conditions that allow an assessment of the effect of the return of natural flow on river health. This study provides a new and valuable contribution to the understanding of the effect of river regulation in Australia.