Expression of melanotropin-related genes in goldfish brain, pituitary, and skin in response to background color

Poster presentado en el 17th International Congress of Comparative Endocrinology celebrado en Barcelona del 15 al 19 de julio de 2013In teleost fish, body color varies in response to changes in background color. The color is lighter in a white background than in a black background. Melanin- concentrating hormone (MCH) produced in hypothalamiand agouti signaling protein (ASP) in skins turn body color pale by aggregating pigments, while melanocyte-stimulating hormone (MSH) encoded on a proopiomelanocortin (POMC) gene disperses pigments. In the present study, we investigated the effects of a black or white background on expression levels of the genes for the hormonal peptides and corresponding receptors by real time RT-PCR in goldfish (Carassiusauratus).Peer Reviewe

Advances in comparative endocrinology : vol. VIII

Ponències presentades al 10th Congress of the Iberian Association of Comparative Endocrinology (AIEC), celebrat a la Universitat Jaume I, els dies 23 al 25 de setembre de 2015Les diverses comunicacions presentades al 10è Congrés de la Asociación Ibérica de Endocrinología Comparada (23-25 setembre 2016, Castelló) s'agrupen en aquest volum. Les intervencions han aportat els darrers avenços en àrees científiques com ara reproducció, metabolisme, estrés, resposta immune, creixement, mineralització i pigmentació...Las diversas comunicaciones presentadas en el 10º Congreso de la Asociación Ibérica de Endocrinología Comparada (23-25 septiembre 2016, Castellón) se agrupan en este volumen. Las intervenciones han aportado los últimos adelantos en áreas científicas como por ejemplo reproducción, metabolismo, estrés, respuesta inmune, crecimiento, mineralización y pigmentación...The present volume of Advances in Comparative Endocrinology collects the contributions of the participants at the 10th Congress of the Iberian Association of Comparative Endocrinology (AIEC). Eighteen years after the foundational meeting of our Association in Peñíscola, the return of this Congress to Castellón highlights the growing success of this initiative to foster the research and scientific development in the field of comparative endocrinology developed in the Iberian Peninsula. AIEC meetings have proven to be a way to keep in contact among research groups with common interests. Some of the participants in this last meeting were also present in the foundational one, others members came after and keep assisting every time. As one of the aims of AIEC has been to encourage students to participate, we are particulary proud of those young students and doctors from the first editions that have gained more permanent positions and continue participating in the AIEC meetings with new students

Hypothalamic Integration of Metabolic, Endocrine, and Circadian Signals in Fish: Involvement in the Control of Food Intake

The regulation of food intake in fish is a complex process carried out through several different mechanisms in the central nervous system (CNS) with hypothalamus being the main regulatory center. As in mammals, a complex hypothalamic circuit including two populations of neurons: one co-expressing neuropeptide Y (NPY) and Agouti-related peptide (AgRP) and the second one population co-expressing pro-opiomelanocortin (POMC) and cocaine- and amphetamine-regulated transcript (CART) is involved in the integration of information relating to food intake control. The production and release of these peptides control food intake, and the production results from the integration of information of different nature such as levels of nutrients and hormones as well as circadian signals. The present review summarizes the knowledge and recent findings about the presence and functioning of these mechanisms in fish and their differences vs. the known mammalian model

Transient Ectopic Overexpression of Agouti-Signalling

While flatfish in the wild exhibit a pronounced countershading of the dorso-ventral pigment pattern, malpigmentation is commonly observed in reared animals. In fish, the dorso-ventral pigment polarity is achieved because a melanization inhibition factor (MIF) inhibits melanoblast differentiation and encourages iridophore proliferation in the ventrum. A previous work of our group suggested that asip1 is the uncharacterized MIF concerned. In order to further support this hypothesis, we have characterized asip1 mRNAs in both turbot and sole and used deduced peptide alignments to analyze the evolutionary history of the agouti-family of peptides. The putative asip precursors have the characteristics of a secreted protein, displaying a putative hydrophobic signal. Processing of the potential signal peptide produces mature proteins that include an N-terminal region, a basic central domain with a high proportion of lysine residues as well as a proline-rich region that immediately precedes the C-terminal poly-cysteine domain. The expression of asip1 mRNA in the ventral area was significantly higher than in the dorsal region. Similarly, the expression of asip1 within the unpigmented patches in the dorsal skin of pseudoalbino fish was higher than in the pigmented dorsal regions but similar to those levels observed in the ventral skin. In addition, the injection/electroporation of asip1 capped mRNA in both species induced long term dorsal skin paling, suggesting the inhibition of the melanogenic pathways. The data suggest that fish asip1 is involved in the dorsalventral pigment patterning in adult fish, where it induces the regulatory asymmetry involved in precursor differentiation into mature chromatophore. Adult dorsal pseudoalbinism seems to be the consequence of the expression of normal developmental pathways in an inaccurate position that results in unbalanced asip1 production levels. This, in turn, generates a ventral-like differentiation environment in dorsal regions.Publicado

Pigment-dispersing activities and cortisol-releasing activities of melanocortins and their receptors in xanthophores and head kidneys of the goldfish Carassius auratus

The five subtypes of melanocortin receptors (MCRs) mediate the functions of α-melanocyte-stimulating hormone (α-MSH) and adrenocorticotropic hormone (ACTH). In fish, these hormones are involved in pigment dispersion and cortisol release, respectively. α-MSH-related peptides exhibit ACTH-like activity in certain fishes. We recently found that multiple Mcr transcripts are expressed in some cell types in the barfin flounder, which is related to regulation of α-MSH activities. Similar results were also observed for the cortisol-releasing activity of α-MSH-related peptides in the head kidney. The present study was undertaken to assess relationship between the expression of multiply expressed Mcrs and α-MSH activities using goldfish. We also determined if α-MSH-related peptides exhibit ACTH-like activity in goldfish. The transcripts of Mc1r, but not those of other subtypes, were observed in xanthophores. α-MSH, which has an acetyl group at the N-terminus, was found to disperse pigment in a dose-dependent manner in xanthophores. This potency was found to be slightly greater than that of desacetyl-α-MSH. These results support our findings that MCR has a higher affinity for α-MSH when single Mcr subtype is expressed. On the other hand, transcripts of Mc2r, but not those of other subtypes, were observed in the head kidney. ACTH1-24-stimulated cortisol release was observed in a dose-dependent manner, while α-MSH-related peptides showed no activity. It therefore appears that MC2R also acts as an ACTH-specific receptor in goldfish and that association of α-MSH-related peptides upon release of cortisol is uncommon in fishes. © 2011 Elsevier Inc.Peer Reviewe

Signal Transduction and Pathogenic Modifications at the Melanocortin-4 Receptor: A Structural Perspective

The melanocortin-4 receptor (MC4R) can be endogenously activated by binding of melanocyte-stimulating hormones (MSH), which mediates anorexigenic effects. In contrast, the agouti-related peptide (AgRP) acts as an endogenous inverse agonist and suppresses ligand-independent basal signaling activity (orexigenic effects). Binding of ligands to MC4R leads to the activation of different G-protein subtypes or arrestin and concomitant signaling pathways. This receptor is a key protein in the hypothalamic regulation of food intake and energy expenditure and naturally-occurring inactivating MC4R variants are the most frequent cause of monogenic obesity. In general, obesity is a growing problem on a global scale and is of social, medical, and economic relevance. A significant goal is to develop optimized pharmacological tools targeting MC4R without adverse effects. To date, this has not been achieved because of inter alia non-selective ligands across the five functionally different MCR subtypes (MC1-5R). This motivates further investigation of (i) the three-dimensional MC4R structure, (ii) binding mechanisms of various ligands, and (iii) the molecular transfer process of signal transduction, with the aim of understanding how structural features are linked with functional-physiological aspects. Unfortunately, experimentally elucidated structural information is not yet available for theMC receptors, a group of class A G-protein coupled receptors (GPCRs). We, therefore, generated MC4R homology models and complexes with interacting partners to describe approximate structural properties associated with signaling mechanisms. In addition, molecular insights from pathogenic mutations were incorporated to discriminate more precisely their individual malfunction of the signal transfer mechanism

Loss-of-function mutations in the melanocortin 1 receptor cause disruption of dorso-ventral countershading in teleost fish

13 pages, 8 figuresThe melanocortin 1 receptor (MC1R) is the central melanocortin receptor involved in vertebrate pigmentation. Mutations in this gene cause variations in coat coloration in amniotes. Additionally, in mammals MC1R is the main receptor for agouti‐signaling protein (ASIP), making it the critical receptor for the establishment of dorsal‐ventral countershading. In fish, Mc1r is also involved in pigmentation, but it has been almost exclusively studied in relation to melanosome dispersion activity and as a putative genetic factor involved in dark/light adaptation. However, its role as the crucial component for the Asip1‐dependent control of dorsal‐ventral pigmentation remains unexplored. Using CRISPR/Cas9, we created mc1r homozygous knockout zebrafish and found that loss‐of‐function of mc1r causes a reduction of countershading and a general paling of the animals. We find ectopic development of melanophores and xanthophores, accompanied by a decrease in iridophore numbers in the ventral region of mc1r mutants. We also reveal subtle differences in the role of mc1r in repressing pigment cell development between the skin and scale niches in ventral regionsThis work was funded by the Spanish Economy and Competitiveness Ministry projects AGL2011‐23581, AGL2014‐52473R, AGL2017‐89648P to JR. Partial funding was obtained from AGL2016‐74857‐C3‐3‐R to JMCR. L. Cal was supported by predoctoral fellowship FPI funded by Spanish Economy and Competitiveness Ministry (AGL2011‐23581) and by predoctoral fellowship of the Spanish Personnel Research Training Program funded by Spanish Economy and Competitiveness Ministry (EEBB‐C‐14‐00467). P Suarez‐Bregua was supported by AGL2014‐52473R and AGL2017‐89648P project contractPeer reviewe

Agouti overexpression in a transgenic model regulates integrity, permeability and electrogenic amino acid transport in zebrafish intestine

Overexpression of asip1 in transgenic zebrafish disrupts dorsoventral pigment pattern in addition to increasing food intake levels and linear growth. A higher feed intake is unnecessary in transgenic fish to enable larger and heavier growth. A plausible explanation may rely on the enhanced feeding efficiency mediated by improved nutrient absorption in transgenic animals. To test this hypothesis, wide scope transcriptomic techniques were used to elucidate the potential pathways involved in the enhanced nutrient absorption and intestinal epithelium permeability/integrity. In addition, the electrogenic capacity for amino acid transport was analysed. Transcriptomic analysis reveal that amino acid, monocarboxylates, ionic and vitamin transmembrane transporters were substantially modified. Enrichment analysis also revealed an inhibition of intestinal lipid metabolism and down-regulation of KEGG pathways related to membrane integrity suggesting augmented intestinal laxity that may enhance paracellular transport. Electrophysiological experiments carried out in Ussing chambers show that asip1 overexpression decrease membraned tissue resistance (Rt), indicating a modification of the intestinal barrier function in ASIP1 transgenic animals. Similarly, paracellular permeability was higher in transgenic zebrafish. Both the decrease in Rt and the increase in permeability point to an ASIP1-dependent decrease in the tissue barrier function. Electrogenic amino acid transport was also enhanced in transgenic animals providing strong indication that ASIP1 fish can extract more amino acids from their diet at similar feeding levels. Both transcriptomic and electrophysiological results suggest that asip1-overexpressing zebrafish display improved nutrient absorption and by extension a higher feed efficiency which explains enhanced growth in the absence of augmented food intake. The enhanced growth of ASIP1 zebrafish potentially mediated by improved nutrient uptake and feed efficiency suggests that the melanocortin system, specifically asip1 overexpression, is a potential target for the development of genetically engineered fish displaying improved performance and no differential lipid accumulation.info:eu-repo/semantics/publishedVersio

Role of the Melanocortin System in Gonadal Steroidogenesis of Zebrafish

In teleost, as in other vertebrates, stress affects reproduction. A key component of the stress response is the pituitary secretion of the adrenocorticotropic hormone (ACTH), which binds to the melanocortin 2 receptor (MC2R) in the adrenal glands and activates cortisol biosynthesis. In zebrafish, Mc2r was identified in male and female gonads, while ACTH has been shown to have a physiological role in modulating reproductive activity. In this study, the hypothesis that other melanocortins may also affect how the zebrafish gonadal function is explored, specifically steroid biosynthesis, given the presence of members of the melanocortin signaling system in zebrafish gonads. Using cell culture, expression analysis, and cellular localization of gene expression, our new observations demonstrated that melanocortin receptors, accessory proteins, antagonists, and agonists are expressed in both the ovary and testis of zebrafish (n = 4 each sex). Moreover, melanocortin peptides modulate both basal and gonadotropin-stimulated steroid release from zebrafish gonads (n = 15 for males and n = 50 for females). In situ hybridization in ovaries (n = 3) of zebrafish showed mc1r and mc4r in follicular cells and adjacent to cortical alveoli in the ooplasm of previtellogenic and vitellogenic oocytes. In zebrafish testes (n = 3), mc4r and mc1r were detected exclusively in germ cells, specifically in spermatogonia and spermatocytes. Our results suggest that melanocortins are, directly or indirectly, involved in the endocrine control of vitellogenesis in females, through modulation of estradiol synthesis via autocrine or paracrine actions in zebrafish ovaries. Adult zebrafish testes were sensitive to low doses of ACTH, eliciting testosterone production, which indicates a potential role of this peptide as a paracrine regulator of testicular function.publishedVersio

Unraveling the periprandial changes in brain serotonergic activity and its correlation with food intake-related neuropeptides in rainbow trout Oncorhynchus mykiss

This study explored changes in brain serotonin content and activity together with hypothalamic neuropeptide mRNA abundance around feeding time in rainbow trout, as well as the effect of one-day fasting. Groups of trout fed at two (ZT2) and six (ZT6) hours after lights on were sampled from 90 minutes before to 240 minutes after feeding, while additional groups of non-fed trout were also included in the study. Changes in brain amine and metabolite contents were measured in hindbrain, diencephalon and telencephalon, while in the diencephalon the mRNA abundance of tryptophan hydroxylase ( tph1 , tph2 ), serotonin receptors (5htr1a , 5htr1b and 5htr2c ) and several neuropeptides ( npy , agrp1 , cartpt , pomca1 , crfb ) involved in the control of food intake were also assessed. The results showed changes in the hypothalamic neuropeptides that were consistent with the expected role for each in the regulation of food intake in rainbow trout. Serotonergic activity increased rapidly at the time of food intake in the diencephalon and hindbrain and remained high for much of the postprandial period. This increase in serotonin abundance was concomitant with elevated levels of pomca1 mRNA in the diencephalon, suggesting that serotonin might act on brain neuropeptides to promote a satiety profile. Furthermore, serotonin synthesis and neuronal activity appear to increase already before the time of feeding, suggesting additional functions for this amine before and during food intake. Exploration of serotonin receptors in the diencephalon revealed only small changes for gene expression of 5htr1b and 5htr2c receptors during the postprandial phase. Therefore, the results suggest that serotonin may play a relevant role in the regulation of feeding behavior in rainbow trout during periprandial time, but a better understanding of its interaction with brain centers involved in receiving and processing food-related signals is still needed.Agencia Estatal de Investigación | Ref. PID2022-136288OB-C31Xunta de Galicia | Ref. ED431B 2019/37Agencia Estatal de Investigación | Ref. BES-2017-079708Xunta de Galicia | Ref. ED481B-2022-08