Dopamine in the ink defence system of Sepia officinalis: biosynthesis, vesicular compartmentation in mature ink gland cells, nitric oxide (NO)/cGMP-induced depletion and fate in secreted ink

Abstract

The biosynthesis, localization and fate of catecholamines in the ink gland of the cuttlefish Sepia officinalis were investigated by combined biochemical and immunohistocytochemical methodologies. HPLC analysis of crude ink gland extracts indicated the presence of dopa (2.18+/-0.82 nmol/mg of protein) and DA (dopamine, 0.06+/-0.02 nmol/mg of protein), but no detectable noradrenaline or adrenaline. DA was shown to derive from L-tyrosine, according to experiments performed by incubating intact ink glands with [L-14C]tyrosine. The biosynthetic process involves a tyrosine hydroxylase and a dopa decarboxylase pathway and is independent of tyrosinase. The tyrosine hydroxylase activity was detected under conditions of tyrosinase suppression in the cytosolic fraction, but not in the melanosomal fraction, of ink gland extracts, and the presence of the enzyme was confirmed by Western-blot analysis. Dopa and DA were found to be released from the ink glands by processes controlled through the NMDA-nitric oxide-cGMP (where NMDA stands for N -methyl-D-aspartate) signalling pathway, as apparent from incubation experiments performed with [L-14C]tyrosine in the presence of NMDA, diethylamine NONOate (diethylamine diazeniumdiolate), a nitric oxide donor, 8-bromo-cGMP or a guanylyl cyclase inhibitor. Immunohistochemical results coupled with electron microscopy indicated that DA was concentrated in vesicles specifically localized in the mature melanin-producing cells of the ink gland proximal to the lumen and separated from the melanin-containing melanosomes. NMDA receptor stimulation or exposure to an NO donor caused a marked loss of DA immunoreactivity in mature cells, consistent with a release process. In the lumen of the ink gland, where mature exhausted cells pour their contents, DA immunoreactivity was found to be associated with the melanin granules, due apparently to physical adsorption. Overall, these results point to DA as a marker of cell maturation in Sepia ink gland subject to release by the NO/cGMP signalling pathway, and disclose apparently overlooked DA-melanin interactions in secreted ink of possible relevance to the defence mechanism.

Ink From Longfin Inshore Squid, Doryteuthis pealeii, as a Chemical and Visual Defense Against Two Predatory Fishes, Summer Flounder, Paralichthys dentatus, and Sea Catfish, Ariopsis felis

“…This fraction includes but is not limited to melanin, raising the possibility that the defensive compounds could be either melanin itself or compounds adhering to or associated with melanin. For example, it is known that dopamine adsorbs onto melanin granules of ejected cuttlefish ink (Fiore et al, 2004). The bioactive fraction of squid ink also includes particulates besides melanin, and these could be the source of the chemical deterrents.…”

Étude préliminaire du traitement d’effluents contenant de l’encre de seiche par centrifugation et procédés à membranes

“…SHIRAI et al (1997) et LUCERO et al (1994 ont montré que l'encre est fortement concentrée en deux amines : la dopamine et la L dopa (LD) qui constituent un stimulus pour les neurones olfactifs et contient aussi un antioxydant non identifié qui prévient l'oxydation de ces deux amines à leur contact dans l'eau de mer. Divers enzymes mélanogéniques parmi lesquelles la tyrosinase, le dopachrome rearranging enzyme, la péroxydase et la P mel 17 (PALUMBO et al, 1994et PALUMBO 2003FOIRE et al, 2004).…”

The dynamic nitric oxide pattern in developing cuttlefish Sepia officinalis

“…In the cuttlefish Sepia officinalis, NO plays a key role in the defense system, regulating the metabolism of the ink gland and the activity of chromatophores Fiore et al, 2004;Mattiello et al, 2010). Moreover, NO functions as neurotransmitter in the central and peripheral nervous system, regulates blood pressure acting as vasodilatory mediator, modulates the statocyst activity and participates in manipulative behavior (Schipp and Gebauer, 1999;Palumbo et al, 1999;Tu and Budelmann, 2000;Di Cosmo et al, 2000;Halm et al, 2003;Di Cristo et al, 2007).…”