Reply to Ahluwalia et al.: Contributions of melatonin receptors are tissue-dependent

In our recent publication (1), we advance the previous findings regarding melatonin type 1 (MT₁) receptor in brain mitochondria. We demonstrate that melatonin is synthesized in brain mitochondria and signals through the MT₁ receptor located in the outer mitochondrial membrane (MM). We further show that melatonin binds mitochondria MT₁, which in turn activates inhibitory Gi proteins in the intermembrane space and inhibits stress-mediated cytochrome c release (1, 2). We have exclusively worked with neurons or brain tissue, and have therefore not performed experiments in other cell types at this stage.

In the letter from Ahluwalia et al. (3), the authors propose that melatonin receptors, especially MT₂, are located in MMs of gastric endothelial cells (GEC). Their data expand the knowledge of melatonin receptors beyond brain and neuronal mitochondria. Furthermore, the authors find that melatonin increases both MT₁ and MT₂ protein levels in GEC cell mitochondria (3). In a previous study we demonstrated that melatonin administration inhibited the mutant huntingtin-induced reduction of MT₁ protein and mRNA, but not MT₂ protein and mRNA in the R6/2 murine brain (2), suggesting that dysfunctional melatonin signaling through MT₁ in the murine brain may be relevant to neurodegeneration and may be amenable to modulation by exposure to exogenous melatonin. In our more recent study (1), we found that luzindole, which inhibits MT₁ and MT₂, decreased melatonin’s ability to inhibit cytochrome c release while the MT₂ receptor-specific antagonist, 4P-PDOT, was unable to do so. These data indicate that luzindole antagonized melatonin’s inhibitory effect on cytochrome c release due to its inhibition of MT₁ and not MT₂ in the brain.

These findings by Ahluwalia et al. (3) and our studies (1, 2, 4) indicate that the contribution of melatonin receptors and its signaling pathways can presumably be cell- and tissue-dependent. However, it must be pointed out that caution needs to be taken when interpreting data obtained using MT₁ antibodies. Extensive MT₁ antibody validation data in our laboratory and by others (5) suggest that none of the currently commercially available MT₁ antibodies that we tested are specific to the MT₁ protein. Thus, work showing localization of endogenous melatonin receptors using antibody-based methods must be validated by including robust controls, such as tissues from knockout MT₁ and MT₂ mice to demonstrate antibody reliability.