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

Sensing Glucose in the Central Melanocortin Circuits of Rainbow Trout: A Morphological Study

In mammals, glucosensing markers reside in brain areas known to play an important role in the control of food intake. The best characterized glucosensing mechanism is that dependent on glucokinase (GK) whose activation by increased levels of glucose leads in specific hypothalamic neurons to decreased or increased activity, ultimately leading to decreased food intake. In fish, evidence obtained in recent years suggested the presence of GK-like immunoreactive cells in different brain areas related to food intake control. However, it has not been established yet whether or not those neuronal populations having glucosensing capacity are the same that express the neuropeptides involved in the metabolic control of food intake. Therefore, we assessed through dual fluorescent in situ hybridization the possible expression of GK in the melanocortinergic neurons expressing proopiomelanocortin (POMC) or agouti-related protein (AGRP). POMC and AGRP expression localized exclusively in the rostral hypothalamus, in the ventral pole of the lateral tuberal nucleus, the homolog of the mammalian arcuate nucleus. Hypothalamic GK expression confined to the ependymal cells coating the ventral pole of the third ventricle but some expression level occurred in the AGRP neurons. GK expression seems to be absent in the hypothalamic POMC neurons. These results suggest that AGRP neurons might sense glucose directly through a mechanism involving GK. In contrast, POMC neurons would not directly respond to glucose through GK and would require presynaptic inputs to sense glucose. Ependymal cells could play a critical role relying glucose metabolic information to the central circuitry regulating food intake in fish, especially in POMC neurons

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

Expression of long-chain polyunsaturated fatty acid (LC-PUFA) biosynthesis genes during zebrafish Danio rerio early embryogenesis

Long-chain polyunsaturated fatty acids (LC-PUFAs) are essential in important physiological processes, many of which are particularly vital during embryonic development. This study investigated the expression of genes encoding enzymes involved in LC-PUFA biosynthesis, namely fatty acyl desaturase (Fad) and Elovl5- and Elovl2-like elongases, during early embryonic development of zebrafish. Firstly, zebrafish elovl2 cDNA was isolated and functionally characterised in yeast, showing high specificity towards C20 and C22 PUFAs, compared to C18 substrates. Secondly, spatial-temporal expression for elovl2 and the previously cloned fad and elovl5 were studied during zebrafish early embryonic development. Temporal expression shows that all three genes are expressed from the beginning of embryogenesis (zygote), suggesting maternal mRNA transfer to the embryo. However, a complete activation of the biosynthetic pathway seems to be delayed until 12 hpf, when noticeable increases of fad and elovl2 transcripts were observed, in parallel with high docosahexaenoic acid levels in the embryo. Spatial expression was studied by whole-mount in situ hybridization in 24 hpf embryos, showing that fad and elovl2 are highly expressed in the head area where neuronal tissues are developing. Interestingly, elovl5 shows specific expression in the pronephric ducts, suggesting an as yet unknown role in fatty acid metabolism during zebrafish early embryonic development. The yolk syncytial layer also expressed all three genes, suggesting an important role in remodelling of yolk fatty acids during zebrafish early embryogenesis. Tissue distribution in zebrafish adults demonstrates that the target genes are expressed in all tissues analysed, with liver, intestine and brain showing the highest expression

Transient ectopic overexpression of agouti-signalliprotein 1 (Asip1) induces pigment anomalies in flatfish

10 páginas, 9 figurasWhile 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 dorsal-ventral 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.This research was carried out with the financial support of the Xunta de Galicia Science Program INCITE (Incite09 402 193 PR to JR) and Science and Innovation Ministry (AGL2010-22247-C03-01 to JMC-R and ALG2011-23581 to JR). Additional funding was obtained from the “Generalitat Valenciana” (research grant PROMETEO 2010/006) to JMC-R. RMC was recipient of a JAE-postdoctoral fellowship from Consejo Superior de Investigaciones Científicas (CSIC)Peer reviewe

Distribution of two isoforms of tryptophan hydroxylase in the brain of rainbow trout (Oncorhynchus mykiss). An in situ hybridization study

© The Author(s) 2021.Serotonin (5-HT) is one of the principal neurotransmitters in the nervous system of vertebrates. It is initially synthesized by hydroxylation of tryptophan (Trp) by means of tryptophan hydroxylase or TPH which is the rate-limiting enzyme in the production of 5-HT. In most vertebrates, there are two isoforms of TPH present, TPH1 and TPH2, which exhibit different catalytic or substrate specificity as well as different expression domains. Studies carried out in mammals show that only tph2 is expressed in the brain whereas tph1-mRNA is primarily localized in the enterochromaffin cells and pineal gland. A large number of neurons are also considered to be serotonergic or “pseudo-serotonergic” as they accumulate and release 5-HT yet do not produce it as no amine-synthetic enzymes are expressed, yet a combination of 5-HT transporters is observed. Therefore, tph expression is considered to be the only specific marker of 5-HT-producing neurons that can discriminate true 5-HT from pseudo-serotonergic neurons. This work examined in situ hybridization to study the mRNA distribution of one paralogue for tph1 and tph2 in the central nervous system of rainbow trout. Results show a segregated expression for both paralogues that predominantly match previous immunocytochemical studies. This study thus adds valuable information to the scarce analyses focusing on the central distribution of the expression of serotonergic markers, particularly tphs, in the vertebrate brain thus characterizing the true serotonergic brain territories.This research was funded by the Spanish State Agency of Research (AEI), grant number AGL2016-74857-C3-3-R and PID2019-103969RB-C33 to JMCR and PID2019-103969RB-C31 to JMM. M.C. was recipient of a predoctoral fellowship (Program FPI) from Spanish Ministerio de Ciencia e Innovación (BES-2017-079708)

Transient overexpression of agouti-signalling protein 1 induces pigment anomalies in flatfish

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Fish melanocortin system

31 p. ReviewMelanocortin signalling is mediated by binding to a family of G protein-coupled receptors that positively couple to adenylyl cyclase. Tetrapod species have five melanocortin (MC1-MC5) receptors. The number of receptors varies in fish, zebrafish, for example, having six melanocortin receptors, with two copies of the melanocortin MC5 receptor, while pufferfish have 4 receptors with no melanocortin MC3 receptor and one copy of melanocortin MC5 receptor. Fish genomes also exhibit orthologue genes for agouti-signalling protein (ASP) and -related protein (AGRP). AGRP expression is confined to a small area in the hypothalamus but ASP is expressed in the skin. Fish melanocortin MC2 receptor is specific for ACTH and requires the cooperation of accessory proteins (MRAP) to reach functional expression. The four other melanocortin MC receptors distinctively bind MSHs. The interaction of α-MSH and melanocortin MC 1 receptor plays a key point in the control of the pigmentation and mutations of melanocortin MC1 receptor are responsible for reduced melanization. Both melanocortin MC4 and MC5 receptor are expressed in the hypothalamus, and central melanocortin MC4 receptor expression is thought to regulate the energy balance through the modulation of feeding behaviour. In addition, the peripheral melanocortin system also regulates lipid metabolism by acting at hepatic melanocortin MC2 and MC5 receptors. Both sea bass melanocortin MC1 and MC 4 receptors are constitutively expressed in vitro and both ASP and AGRP work as inverse agonists but only after inhibition of the phosphodiesterase system. Accordingly, the overexpression of AGRP and ASP transgenes promotes obesity and reduces melanization in zebrafish, respectively.This work was partially supported by grants from the Ministry of Science and Innovation (MICINN) AGL2007-65744-C03-02, CSD 2007-00002 and AGL2010-22247-C03-01 to JM C-R. MJA is recipient of a “Juan de la Cierva” research contract (2009) from the Spanish Science and Innovation Ministry.Peer reviewe

Molecular characterization of agouti-signaling protein 1 (asip1) in flatfish: an evolutionary scenario

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Pigment patterns in fish result from superimposition of two largely independent pigmentation mechanisms

Trabajo presentado en la 27th Conference of European Comparative Endocrinologists, celebrada en Rennes (Francia), del 25 al 29 de agosto de 2014Dorso-ventral pigment pattern differences are the most widespread pigmentary adaptations in vertebrates. In mammals, this pattern is controlled by regulating melanin chemistry in melanocytes using a protein, Asip. In fish, studies of pigment patterning have focused on stripe formation, identifying a core striping mechanism dependent upon interactions between different pigment cell types. In contrast, mechanisms driving the dorso-ventral countershading pattern have been overlooked. Here, we demonstrate that, in fact, zebrafish utilize two distinct adult pigment patterning mechanisms - an ancient dorsoventral patterning mechanism, and a more recent striping mechanism based on cell-cell interactions; remarkably, the dorso-ventral patterning mechanism also utilizes Asip. These two mechanisms function largely independently, with resultant patterns superimposed to give the full patternThis work was funded by the Spanish Science and Innovation Ministry project ALG2011-23581 and Xunta de Galicia INCITE-09 402 193 to JR. Partial funding was also obtained from Science and Innovation Ministry (AGL2010-22247-C03-01 to JMC-R)Peer reviewe