Differential effect of amphetamine over the corticotropin-releasing factor CRF 2 receptor, the orexin OX 1 receptor and the CRF 2 -OX 1 heteroreceptor complex.

Stress is one of the factors underlying drug seeking behavior that often goes in parallel with loss of appetite. We here demonstrate that orexin 1 receptors (OX1 R) may form complexes with the corticotropin releasing factor CRF2 receptor. Two specific features of the heteromer were a cross-antagonism and a blockade by CRF2 of OX1 R signaling. In cells expressing one of the receptors, agonist-mediated signal transduction mechanisms were potentiated by amphetamine. Sigma 1 (σ1 ) and 2 (σ2 ) receptors are targets of drugs of abuse and, despite sharing a similar name, the two receptors are structurally unrelated and their physiological role is not known. We here show that σ1 receptors interact with CRF2 receptors and that σ2 receptors interact with OX1 R. Moreover, we show that amphetamine effect on CRF2 receptors was mediated by σ1 R whereas the effect on OX1 receptors was mediated by σ2 R. Amphetamine did potentiate the negative cross-talk occurring within the CRF2 -OX1 receptor heteromer context, likely by a macromolecular complex involving the two sigma receptors and the two GPCRs. Finally, in vivo microdialysis experiments showed that amphetamine potentiated orexin A-induced dopamine and glutamate release in the ventral tegmental area (VTA). Remarkably, the in vivo orexin A effects were blocked by a selective CRF2 R antagonist. These results show that amphetamine impacts on the OX1 R-, CRF2 R- and OX1 R/CRF2 R-mediated signaling and that cross-antagonism is instrumental for in vivo detection of GPCR heteromers.