AbstractA biotesting method for the assessment of acute sublethal toxicity to charophyte cells ofNitellopsis obtusawas developed and applied for the study of Vilnius and Visaginas wastewaters (WW). This computer‐assisted testing procedure was based on the recording of the electrophysiological response of up to 16 single algal cells simultaneously. Membrane parameters of living cells were measured according to the K+‐anaesthesia method modified for multichannel recording with extracellular electrodes. Concentration‐dependent monotonical decrease of cell resting potential, induced by toxic WW action, was used to determine an effective concentration (EC) values. A multistep time‐saving procedure, when the cell group was exposed to subsequent treatment with increasing WW concentrations (30 min per treatment), proved to be adequate for biotesting. The ECs obtained by this procedure were in good agreement with those obtained alternatively by undertaking one‐step testing (i.e., 60 min exposure per concentration per cell group). The relative sensitivity of the cellular electrophysiological response, as a toxicity end point, was confirmed by comparing it with lethal concentration (LC) responses in 96‐ and 192‐h exposure tests conducted on the sameN. obtusacells. For Vilnius WW samples taken on different dates, the EC 50 sranged from 27 to 46% WW and corresponded to 96‐h LC 50 sof 28–50% WW. In contrast, Visaginas WWs sampled before and after chlorination showed no toxic effect either at the membrane or whole cell level.