Elevated alkalinity values measured in sediment pore water samples from the Grand Calumet River–Indiana Harbor Canal, an International joint Commission Area of Concern (AOC), caused concern over the potential effects of alkalinity on cladoceran test responses. Forty-eight-hour tests with NaHCO3 and NaCl as reference toxicants indicated that NaHCO3 toxicity to Daphnia magna and Ceriodaphnia dubia was due to both Na+ and HCO3−, although greater HCO3− toxicity was observed to D. magna. Theoretical HCO3− concentrations in sediment pore waters from the AOC demonstrated sufficient HCO3− in several samples to produce acute toxicity, based on 48-h LC50s from the reference toxicant tests. X-ray microanalysis was conducted to qualitatively assess the effects on internal Cl− levels of D. magna after exposure to NaHCO3, to an AOC sediment pore water sample, and to NaSCN, a metabolic inhibitor of Cl− uptake. Based on these results the proposed mechanism of HCO3− toxicity to D. magna is the inhibition of the active uptake of Cl− from water. We suggest that pore water alkalinity should be considered when interpreting the results of sediment pore water and effluent toxicity tests with D. magna, other cladocerans, and, perhaps, other invertebrates and fish.