Cloning and functional characterization of the DA2 receptor gene in Chinese mitten crab (Eriocheir sinensis)

Abstract

Dopamine (DA) plays a modulatory role in numerous physiological processes such as light adaptation and food intake, and exerts these functions through DA receptors (DARs). This study presents, for the first time, isolation and characterization of the dopamine receptor 2(DA2 receptor) cDNA from the intestinal tissue of Eriocheir sinensis, an economically important freshwater aquaculture species in China. The DA2 receptor cDNA sequence, which was obtained by rapid amplification of cDNA ends, is 2369bp long, encode peptide of 589 amino acid, and is highly homologous to related sequences in crustaceans. Analysis of the deduced amino acid sequence and the structure of the DA2 indicated that this receptor is a member of the family of G protein-coupled receptors (GPCRs), as it contains seven transmembrane domains and other common signatures of GPCRs. RT-PCR showed that the expression of the DA2 receptor gene was distributed in various tissues, and high expression levels were observed in the cranial ganglia and the thoracic ganglia. Further study of the effect of photoperiod on DA2 expression showed that constant darkness induced a significant increase in DA2 expression in the cranial ganglia. Finally, analysis of DA2 receptor expression under different feeding statuses showed that there was significantly greater expression in the hepatopancreas and intestines after feeding than before feeding, but there were no differences in expression between the before feeding and during feeding periods in either tissue. Our results indicate that the DA2 receptor structurally belongs to the family of G protein-coupled receptors, and that the cranial ganglia are the main tissues in which the DA2 receptor participates in light adaptation during dark hours. In addition, the DA2 receptor in E. sinensis may be involved in the physiological regulation of the hepatopancreas and digestive tract after the ingestion of food. This study provides a foundation for further exploration of the light adaptation and digestive functions of the DA2 receptor in decapods.

5-HT2B, 5-HT7, and DA2 Receptors Mediate the Effects of 5-HT and DA on Agonistic Behavior of the Chinese Mitten Crab (Eriocheir sinensis)

“…The total RNA of the cerebral ganglion and thoracic ganglia was extracted using RNAiso Plus reagent . The mRNA levels of 5-HT2B, 5-HT7, and DA2 receptors were determined using an ABI 7500 real-time PCR system (Life Technology, USA).…”

“…The total RNA of the cerebral ganglion and thoracic ganglia was extracted using RNAiso Plus reagent. 33 The mRNA levels of 5-HT2B, 5-HT7, and DA2 receptors were determined using an ABI 7500 real-time PCR system (Life Technology, USA). The 5-HT and DA receptors were detected with gene-specific primers (Table 3), and 18S rRNA was used as the reference gene.…”

“…In our previous studies, the 5-HT7 and DA2 receptors in E. sinensis were cloned and characterized. 32,33 To the best of our knowledge, that was the first report on agonistic behavior in E. sinensis, and changes in the levels of 5-HT, DA, cAMP and PKA in the hemolymph were found after fighting. 29 In the current paper, we further investigated which receptors are involved in the regulation of 5-HT and DA in relation to agonistic behavior through downstream signaling pathways (cAMP pathway).…”

The transcriptome sequencing and functional analysis of eyestalk ganglions in Chinese mitten crab (Eriocheir sinensis) treated with different photoperiods

“…We used 18S ribosome RNA (18S) as a reference gene. The DEGs expression levels were calculated using the 2 -ΔΔCt method [23].…”

“…In our previous studies, we found that the eyestalks of E . sinensis express the DA2 receptor which participates in light adaptation during the dark hours [23]. However, there still lack information about the influence of photoperiods on the gene expression of eyestalk ganglion in E .…”