Immunocytochemical evidence for the presence of met‐enkephalin and leu‐enkephalin distinct neurons in the brain of the elasmobranch fish Scyliorhinus canicula

Abstract

Immunohistochemical methods have been used to investigate the distribution of various opioid peptides derived from mammalian proenkephalin in the central nervous system of Scyliorhinus canicula. The results indicate that both Leu- and Met-enkephalin-immunoreactive peptides are present in the dogfish brain. In contrast, enkephalin forms similar to Met-enkephalin-Arg-Phe or Met-enkephalin-Arg-Gly-Leu, and mammalian alpha-neo-endorphin, dynorphin A (1-8), dynorphin A (1-13), and dynorphin A (1-17) were not detected. Met- and Leu-enkephalin immunoreactivities were found in distinct neurons of the telencephalon and hypothalamus. In particular, cell bodies reacting only with the Met-enkephalin antiserum were localized in the preoptic nucleus and in the suprachiasmatic region of the hypothalamus. Conversely, cell bodies reacting only with the Leu-enkephalin antiserum were localized in the pallium and the nucleus lobi lateralis hypothalami. Several areas of the telencephalon and diencephalon exhibited both Met- and Leu-enkephalin-like immunoreactivity, but the two immunoreactive peptides were clearly contained in distinct perikarya. The overall distribution of Met-enkephalin-immunoreactive elements in the dogfish exhibited similarities to the distribution of proenkephalin-derived peptides previously reported for the brain of tetrapods. The fact that Met- and Leu-enkephalin-like peptides were detected in distinct neurons, together with the absence of dynorphin-related peptides, suggests the existence of a novel Leu-enkephalin-containing precursor in the dogfish brain.

Analyzing the evolution of the opioid/orphanin gene family

“…When these studies were begun in the 1980s, it was assumed that: (a) met-enkephalin and leu-enkephalin would be co-localized in neurons; (b) the molar ratio of metenkephalin to leu-enkephalin in CNS extracts would be approximately 4:1; and (c) both of these opioids would be encoded by the same precursor. Steady-state analyses of non-mammalian brain extracts (Kilpatrick et al, 1983;Lindberg & White, 1986), and immunocytochemical studies (Vallarino et al, 1994(Vallarino et al, , 1998 indicated that the evolution of the enkephalinergic system was more complicated than had been anticipated.…”

A Developmental Approach to Forebrain Organization in Elasmobranchs: New Perspectives on the Regionalization of the Telencephalon

“…Experimental studies have provided significant information on telencephalic connections in chondrichthyans [Smeets, 1982[Smeets, , 1983, and the Golgi method has shown a variety of cell types in the telencephalon [Manso and Anadón, 1993]. In addition, immunohistochemical techniques have provided a detailed picture of forebrain organization in the Lesser Spotted Dogfish, revealing neuronal systems that contain modulatory neuroactive substances, such as calbindin D28k [Rodriguez-Moldes et al, 1990], FMRFamide [Vallarino et al, 1991;Chiba et al, 1991], neuropeptide Y [Chiba and Honma, 1992], substance P [Rodríguez- Moldes et al, 1993], Met-and Leu-enkephalin [Vallarino et al, 1994], Thyrotropin-releasing hormone [Teijido et al, 2002] and choline acetyltransferase [Anadón et al, 2000].…”

Immunocytochemical localization of enkephalins in the brain of the African lungfish, Protopterus annectens, provides evidence for differential distribution of Met‐enkephalin and Leu‐enkephalin

“…The antisera to Met-enkephalin and Leu-enkephalin used in this study have been applied previously with success to localize Met-enkephalin and Leu-enkephalin in the brain of the cartilaginous fish S. canicula (Vallarino et al, 1994). Specificity controls showed that these antisera were highly specific.…”

“…In particular, Met-enkephalin and Leu-enkephalin have been localized in the brain of the elasmobranch fish Scyliorhinus canicula (Vallarino et al, 1994). The distribution of Leu-enkephalin has been described in the telencephalon of the spiny dogfish Squalus acanthias (Northcutt et al, 1988) and in the brainstem of the horn shark Heterodontus francisci (Stuesse et al, , 1992, in Hydrolagus collei Stuesse et al, 1992), and in S. acanthias and Raja binoculata (Stuesse et al, 1992).…”