Multiple cell targets for melatonin action in Xenopus laevis retina: Distribution of melatonin receptor immunoreactivity

Abstract

In the retina of the African clawed frog (Xenopus laevis), melatonin is synthesized by the photoreceptors at night, and binds to receptors that likely mediate paracrine responses. Melatonin appears to alter the sensitivity of the retinal cells to light, and may play a key role in regulating important circadian events that occur in the eye. A polyclonal antibody was raised against a 13 amino acid peptide corresponding to a region of the third cytoplasmic loop of the Xenopus laevis Mel_1c melatonin receptor. Western blot analysis revealed a major immunoreactive band of approximately 60 kD in neural retina and retinal pigment epithelium (RPE) membranes. Immunocytochemical labeling of sections of Xenopus eyes demonstrated intense melatonin receptor-like immunoreactivity in the inner plexiform layer (IPL). Immunolabeling with antibodies to glutamate decarboxylase (GAD) or tyrosine hydroxylase (TOH) appeared to co-localize with the melatonin receptor immunoreactivity in different sublaminas of the IPL. This suggests that both GABAergic and dopaminergic amacrine cells express melatonin receptor protein. There were also some melatonin receptor immunoreactive varicose fibers in the IPL that did not co-localize with either TOH or GAD, and may represent efferent fibers, since they could be followed into the optic nerve. Melatonin receptor immunoreactivity was also present on cell soma in the ganglion cell layer. Furthermore, a moderate level of melatonin receptor immunoreactivity was observed in the RPE and rod and cone photoreceptor cells. The presence of melatonin receptor immunoreactivity in these cells supports previous observations of melatonin receptor RNA expression in multiple cell types in the Xenopus retina. Expression of melatonin receptor protein in the photoreceptors suggests that melatonin may have a direct action on these cells.