Thyroid hormone insufficiency leads to impaired neurogenesis, behavioral alterations and cognitive deficits. Thyroid hormone receptors, expressed in brain regions involved in these behaviors, mediate the effects of thyroid hormone deficiency or excess. To determine the contribution of thyroid hormone receptor alpha (TRalpha) in these behaviors, we examined the behavior of euthyroid as well as hypo- and hyperthyroid mice lacking all isoforms of the TRalpha (TRalpha(o/o)). The hypothyroxinemic TRalpha(o/o) mice demonstrated behavioral inhibition, manifested in decreased activity and increased anxiety/fear in the open field test (OFT) and increased immobility in the forced swim test (FST) compared to C57BL/6J mice. TRalpha(o/o) mice also showed learning and recall impairments in the Morris water maze (MWM), which were exaggerated by hypothyroidism in TRalpha(o/o) mice. These impairments were concurrent with increased thigmotaxis, suggesting an increased anxiety-like state of the TRalpha(o/o) mice in the MWM. Expression of genes, known to be involved in processes modulating learning and memory, such as glucocorticoid receptor (GR), growth-associated protein 43 (GAP-43) and neurogranin (RC3), were significantly decreased in the hippocampus of TRalpha(o/o) mice. GR expression was also decreased in the frontal cortex and amygdala of TRalpha(o/o) mice, indicating that expression of GR is regulated, probably developmentally, by one or more isoforms of TRalpha in the mouse brain. Taken together these data demonstrate behavioral alterations in the TRalpha(o/o) mice, indicating the functional role of TRalpha, and a delicate interaction between TRalpha and TRbeta-regulated genes in these behaviors. Thyroid hormone-regulated genes potentially responsible for the learning deficit found in TRalpha(o/o) mice include GR, RC3 and GAP-43.