Antitoxin is currently the only approved therapy for botulinum intoxications. The efficacy of antitoxin preparations is evaluated in animals. However, while in practice antitoxin is administered to patients only after symptom onset, in most animal studies, it is tested in relation to time post intoxication. This may be attributed to difficulties in quantitating early botulism symptoms in animals. In the current study, a novel system based on high-resolution monitoring of mouse activity on a running wheel was developed to allow evaluation of post-symptom antitoxin efficacy. The system enables automatic and remote monitoring of 48 mice simultaneously. Based on the nocturnal activity pattern of individual naïve mice, two criteria were defined as the onset of symptoms. Post-symptom treatment with a human-normalized dose of antitoxin was fully protective in mice exposed to 4 LD50 of BoNT/A and BoNT/B. Moreover, for the first time, a high protection rate was obtained in mice treated post-symptomatically, following a challenge with BoNT/E, the fastest acting BoNT. The running wheel system was further modified to develop a mouse model for the evaluation of next-generation therapeutics for progressive botulism at time points where antitoxin is not effective. Exposure of mice to 0.3 LD50 of BoNT/A resulted in long-lasting paralysis and a reduction in running activity for 16-18 days. Antitoxin treatment was no longer effective when administered 72 hr post intoxication, defining the time window to evaluate next-generation therapeutics. Altogether, the running wheel systems presented herein offer quantitative means to evaluate the efficacy of current and future anti-botulinum drugs.