High-mobility group box-1 (HMGB1) is a nuclear protein with cytokine-type functions upon its extracellular release. HMGB1 activates inflammatory pathways by stimulating multiple receptors, chiefly toll-like receptor 4 (TLR4) and Receptor for Advanced Glycation End Products (RAGE). TLR4 and RAGE activation has been implicated in memory impairments, although the endogenous ligand subserving these effects is unknown. We examined whether HMGB1 induced memory deficits using novel object recognition test, and which of the two receptor pathways was involved in these effects. Non-spatial long-term memory was examined in wild type, TLR4 knockout, and RAGE knockout mice. Recombinant HMGB1 (10μg, intracerebroventricularly, i.c.v.) disrupted memory encoding equipotently in wild type, TLR4 knockout and RAGE knockout animals, but affected neither memory consolidation, nor retrieval. Neither TLR4 knockout nor RAGE knockout mice per se, exhibited memory deficits. Blockade of TLR4 in RAGE knockout mice using Rhodobacter sphaeroides lipopolysaccharide (LPS-Rs; 20 μg, i.c.v.) prevented the detrimental effect of HMGB1 on memory. These data show that elevated brain levels of HMGB1 induce memory abnormalities which may be mediated by either TLR4, or RAGE. This mechanism may contribute to memory deficits under various neurological and psychiatric conditions associated with the increased HMGB1 levels, such as epilepsy, Alzheimer's disease and stroke.