N-methyl-D-aspartate receptor stimulation activates tyrosinase and promotes melanin synthesis in the Ink gland of the cuttlefish Sepia officinalis through the nitric oxide/cGMP signal transduction pathway -: A novel possible role for glutamate as physiologic activator of melanogenesis

The tyrosinase-catalyzed conversion of L-tyrosine to melanin represents the most distinctive biochemical pathway in the ink gland of the cuttlefish Sepia officinalis; however, the molecular mechanisms underlying its activation have remained so far largely uncharted. In this paper we demonstrate for the first time that L-glutamate can stimulate tyrosinase activity and promote melanin synthesis in Sepia ink gland via the N-methyl-D-aspartate (NMDA) receptor/NO/cGMP signal transduction pathway. Incubation of intact ink glands with either L-glutamate or NMDA resulted in an up to 18-fold increase of tyrosinase activity and a more than 6-fold elevation of cGMP levels. Comparable stimulation of tyrosinase was induced by an NO donor and by 8-bromo-cGMP. An NMDA receptor antagonist, NO synthase (NOS) inhibitors, and a guanylate cyclase blocker suppressed NMDA-induced effects. Immunohistochemical evidence indicated that enhanced cGMP production was localized largely in the mature part of the ink gland. Increased de novo synthesis of melanin was demonstrated in NMDA- and NO-stimulated ink glands by a combined microanalytical approach based on spectro-photometric determination of pigment levels and high performance liquid chromatography quantitation of pyrrole-2,3,5-tricarboxylic acid, a specific melanin marker, in melanosome containing fractions. These results fill a longstanding gap in the understanding of the complex biochemical mechanisms underlying activation of melanogenesis in the mature ink gland cells of S. officinalis and disclose a novel physiologic role of the excitatory neurotransmitter glutamate mediated by the NMDA receptor/NO/cGMP signaling pathway.