RESUMO
The dynamic changes in mRNA expression levels for vasotocin (AVT) and isotocin (IT) receptor gene levels were assessed in a time-course response study in immature male specimens of the gilthead sea bream (Sparus aurata) submitted to hyper- (55 salinity) and hypo-osmotic (5 salinity) challenges. Two different cDNAs for the AVT receptor and one for the IT receptor (V1a2-type and V2-type AVTR, and ITR, respectively) were cloned by screening an S. aurata brain cDNA library. Genes for these receptors were expressed differentially and is nearly ubiquitously in 26 of the examined tissues. In the gills, both environmental salinity challenges up-regulated AVTR V1a2-type gene expression concomitantly with mRNA expression protein activity of Na(+), K(+)-ATPase gene expression and protein, whereas the AVTR V2-type and cystic fibrosis transmembrane conductance regulator (CFTR) mRNA levels were associated with mRNAs environmental salinity, indicating a possible connection between AVTRs and these transporters. In kidney, AVTR V1a2-type gene expression peaked rapidly and lasted only a short time (12-24h) in response to both osmotic challenges. In contrast, AVTR V2-type mRNA levels were enhanced in specimens exposed to hyperosmotic conditions, whereas they decreased under hypoosmotic environments, suggesting an antidiuretic role related to the vasoconstriction function. In the hypothalamus, only the expression of the AVTR V2-type gene was enhanced at 7 and 14 days under both experimental conditions. In the liver, both AVTRs had increased mRNA levels, with the upregulation of their AVTR V2-type gene increasing faster than the V1a2-type. The ITR gene was not sensitive to variations of external salinity in any of the analyzed tissues. Our results demonstrate the involvement of the vasotocinergic, but not the isotocinergic, pathway as well as the hypothalamic function, in the adjustments of both osmoregulatory and metabolic processes after osmotic challenges.
Assuntos
Osmorregulação/genética , Ocitocina/análogos & derivados , Receptores de Vasopressinas/genética , Receptores de Vasopressinas/metabolismo , Dourada/genética , Vasotocina/metabolismo , Sequência de Aminoácidos , Animais , Sequência de Bases , DNA Complementar/genética , Regulação da Expressão Gênica/fisiologia , Brânquias/fisiologia , Hipotálamo/fisiologia , Fígado/fisiologia , Masculino , Dados de Sequência Molecular , Osmorregulação/fisiologia , Pressão Osmótica/fisiologia , Ocitocina/metabolismo , Filogenia , RNA Mensageiro/metabolismo , Salinidade , Dourada/metabolismoRESUMO
To investigate the physiological roles of arginine vasotocin (AVT) and isotocin (IT) in osmoregulatory process in gilthead sea bream (Sparus aurata), a time course study (0, 12h, and 1, 3, 7 and 14 days) has been performed in specimens submitted to hypoosmotic (from 40 salinity to 5 salinity) or hyperosmotic (from 40 salinity to 55 salinity) challenges. Plasma and liver osmoregulatory and metabolic parameters, as well as AVT and IT pituitary contents were determined concomitantly with hypothalamic pro-vasotocin (pro-VT) and pro-isotocin (pro-IT) mRNA expression levels. Previously, sequences coding for pro-VT and pro-IT cDNAs were cloned. Two osmoregulatory periods related to plasma osmolality and metabolic parameter variations could be distinguished: i) an adaptative period, from 12h to 3 days after transfer, and ii) a chronic regulatory period, starting at day 3 after transfer. Higher values in hypothalamic pro-VT and pro-IT mRNA expression as well as in pituitary AVT and IT storage levels in both hypo- and/or hyper-osmotic transfers have been distinguished. These increase correlated with changes in plasma cortisol levels, suggesting an interaction between this hormone and pro-VT expression. Furthermore, pro-IT expression enhancement also suggests a role of the isotocinergic system as a modulator in the acute stress response induced by hyper-osmotic challenge in S. aurata.