The development of tolerance to the anticonvulsant effect of chronically administered phenobarbital was demonstrated in mice by the maximal electroshock (MES) test. Anticonvulsant activity decreased 50% over a 5 day period. Brain and plasma levels of phenobarbital measured 2 hr after a 25 mg/kg dose of phenobarbital were the same between days 1 and 5, indicating that distributional or pharmacokinetic parameters were not involved. The loss of anticonvulsant activity of phenobarbital is primarily related to the drug's effect to prevent the spread of MES-induced epileptic neuronal activity. In contrast, the anticonvulsant action of phenobarbital on seizure threshold, as measured by the minimal electroshock seizure threshold (EST) test, and on the spread of epileptic discharge induced by pentylenetetrazol (PTZ), as evaluated by the PTZ infusion test, remained unchanged.