The dynamics of interstitial assemblages, after a spate and during low discharge, was studied in a regulated channel (Miribel Canal) of the Upper Rhone River, France. Using a Bou-Rouch pump, three stations were samples: 1) Station IIA, a site fed by superficial water infiltrations, 2) Station IIC, a site fed by riparian phreatic water, and 3) Station IA, a site fed by both surface and phreatic waters. The spate greatly influenced the interstitial assemblages, their dynamics were different according to the hydrology of the given site. At Station IIA, the spate had a wash-out effect on the assemblages (reduction in abundance and diversity), whereas during low discharge the interstitial layer received a continuous influx of epigean organisms (benthic and limnophilous). At Stations IIC and IA, the spate introduced numerous limnophilous and benthic invertebrates into the interstices, which function as an organismic trap. However, these sites appear to be more isolated from surface waters during low discharge. Stygobites decreased or disappeared after the spate. They appear highly sensitive to hydrologic perturbations in the surface waters.