Gene knockout of insulin-regulated aminopeptidase: Loss of the specific binding site for angiotensin IV and age-related deficit in spatial memory

The AT₄ ligands, angiotensin IV and LVV-hemorphin 7, elicit robust effects on facilitating memory by binding to a specific site in the brain historically termed the angiotensin AT₄ receptor. The identification of the AT₄ receptor as insulin-regulated aminopeptidase (IRAP) is controversial, with other proteins speculated to be the target(s) of these peptides. In this study we have utilized IRAP knockout mice to investigate IRAP in the brain. We demonstrate that the high-affinity binding site for angiotensin IV is absent in IRAP knockout mice brain sections in parallel with the loss of IRAP immunostaining, providing irrefutable proof that IRAP is the specific high-affinity binding site for AT₄ ligands. However, our characterization of the behavioural phenotype of the IRAP knockout mice revealed a totally unexpected finding. In contrast to the acute effects of IRAP inhibitors in enhancing memory, deletion of the IRAP gene resulted in mice with an accelerated, age-related decline in spatial memory that was only detected in the Y maze paradigm. Moreover, no alterations in behaviour of the IRAP knockout mice were observed that could assist in elucidating the endogenous substrate(s). Our results highlight the importance of analysing the behavioural phenotype of knockout mice across different ages and in distinct memory paradigms.