Heteromeric MT1/MT2 melatonin receptors modulate the scotopic electroretinogram via PKCζ in mice

Melatonin plays an important role in the regulation of retinal functions, and previous studies have also reported that the action of melatonin on photoreceptors is mediated by melatonin receptor heterodimers. Furthermore, it has been reported that the melatonin-induced increase in the amplitude of the a- and b-wave is significantly blunted by inhibition of PKC. Previous work has also shown that PKCζ is present in the photoreceptors, thus suggesting that PCKζ may be implicated in the modulation of melatonin signaling in photoreceptors. To investigate the role PKCζ plays in the modulation of the melatonin effect on the scotopic ERG, mice were injected with melatonin and with specific inhibitors of different PKC isoforms. PKCζ knockout mice were also used in this study. PKCζ activation in photoreceptors following melatonin injection was also investigated with immunocytochemistry. Inhibition of PKCζ by PKCζ-pseudosubstrate inhibitor (20 μM) significantly reduced the melatonin-induced increase in the amplitude of the a- and b-wave. To further investigate the role of different PKCs in the modulation of the ERGs, we tested whether intra-vitreal injection of Enzastaurin (a potent inhibitor of PCKα, PKCβ, PKCγ, and PKCε) has any effect on the melatonin-induced increase in the a- and b-wave of the scotopic ERGs. Enzastaurin (100 nM) did not prevent the melatonin-induced increase in the amplitude of the a-wave, thus suggesting that PCKα, PKCβ, PKCγ, and PKCε are not involved in this phenomenon. Finally, our data indicated that, in mice lacking PKCζ, melatonin injection failed to increase the amplitude of the a- and b-waves of the scotopic ERGs. An increase in PKCζ phosphorylation in the photoreceptors was also observed by immunocytochemistry. Our data indicate that melatonin signaling does indeed use the PKCζ pathway to increase the amplitude of the a- and b-wave of the scotopic ERG.