The role of circulatory mouse thyroglobulin (MTg) level in activating mechanisms suppressive to induction of experimental autoimmune thyroiditis (EAT) was studied by two regimens to strengthen normal maintenance of self-tolerance in genetically susceptible mice. One was to administer graded doses of exogenous MTg either 7 days apart or daily for 10 days and then challenge the animals with MTg + LPS. The other was to infuse TSH via an osmotic pump for 7 days. The steady TSH infusion for 7 days resulted in an increase in MTg level peaking on Day 3. Such kinetics of MTg concentration in response to TSH coincided with enhanced resistance to EAT induction. After an initial rapid clearance rate of t1/2 of 3 hr, tolerogenic doses of exogenous MTg sustained similar levels for 2-3 days. In contrast, subtolerogenic doses declined to baseline levels in 2 days or less. Clearance can be best explained by a two-compartment model for distribution with an initial alpha phase (t1/2 about 3 hr), followed by a beta phase (t1/2 about 10 hr). We conclude that, for the prevention of EAT induction in the presence of potent adjuvants (CFA or LPS), a threshold, but above baseline, level of either exogenous or endogenous MTg, represented by the beta phase, is required for a critical period (greater than 2-3 days) to activate suppressor mechanisms over and above homeostatic regulation. Whether MTg concentration raised by TSH (TRH) administration activates suppressor T cells as observed after the injection of a tolerogenic dose of MTg remains to be determined.