The spatiotemporal expression of multiple coho salmon ovarian connexin genes and their hormonal regulation in vitro during oogenesis

Abstract

BackgroundThroughout oogenesis, cell-cell communication via gap junctions (GJs) between oocytes and surrounding follicle cells (theca and granulosa cells), and/or amongst follicle cells is required for successful follicular development. To gain a fundamental understanding of ovarian GJs in teleosts, gene transcripts encoding GJ proteins, connexins (cx), were identified in the coho salmon, Oncorhynchus kisutch, ovary. The spatiotemporal expression of four ovarian cx transcripts was assessed, as well as their potential regulation by follicle-stimulating hormone (FSH), luteinizing hormone (LH) and insulin-like growth factor 1 (IGF1).MethodsSalmonid ovarian transcriptomes were mined for cx genes. Four gene transcripts designated cx30.9, cx34.3, cx43.2, and cx44.9 were identified. Changes in gene expression across major stages of oogenesis were determined with real-time, quantitative RT-PCR (qPCR) and cx transcripts were localized to specific ovary cell-types by in situ hybridization. Further, salmon ovarian follicles were cultured with various concentrations of FSH, LH and IGF1 and effects of each hormone on cx gene expression were determined by qPCR.ResultsTranscripts for cx30.9 and cx44.9 were highly expressed at the perinucleolus (PN)-stage and decreased thereafter. In contrast, transcripts for cx34.3 and cx43.2 were low at the PN-stage and increased during later stages of oogenesis, peaking at the mid vitellogenic (VIT)-stage and maturing (MAT)-stage, respectively. In situ hybridization revealed that transcripts for cx34.3 were only detected in granulosa cells, but other cx transcripts were detected in both oocytes and follicle cells. Transcripts for cx30.9 and cx44.9 were down-regulated by FSH and IGF1 at the lipid droplet (LD)-stage, whereas transcripts for cx34.3 were up-regulated by FSH and IGF1 at the LD-stage, and LH and IGF1 at the late VIT-stage. Transcripts for cx43.2 were down-regulated by IGF1 at the late VIT-stage and showed no response to gonadotropins.ConclusionOur findings demonstrate the presence and hormonal regulation of four different cx transcripts in the salmon ovary. Differences in the spatiotemporal expression profile and hormonal regulation of these cx transcripts likely relate to their different roles during ovarian follicle differentiation and development.

Maternal investment in fish oocytes and eggs: The molecular cargo and its contributions to fertility and early development

“…The increase in follicular gap junctions, composed of various forms of connexin proteins (York et al, 1993;Patiño and Kagawa, 1999;Bolamba et al, 2003), is a critical process in the acquisition of OMC (Yamamoto et al, 2008;Yamamoto and Yoshizaki, 2008;Yamamoto et al, 2011a). Both gonadotropins and Igf1 increase numbers of gap junctions (Yoshizaki et al, 1994;Chang et al, 1999Chang et al, , 2000Choi and Takashima, 2000;Choi and Takemura, 2001), and Lh, Fsh and Igf1 have connexin-specific and stage-specific effects on expression of connexins (Yamamoto et al, 2011a). The specific roles of gap junctions in development of OMC and resumption of meiosis of the oocyte remain to be elucidated.…”

Food-deprivation-induced suppression of pituitary–testicular-axis in the tilapia Oreochromis mossambicus

“…For example, reduction in feeding rate has been shown to inhibit gonadal maturation in the female Atlantic salmon Salmo salar (Thorpe et al 1990;Reimers et al 1993), whereas prolonged fasting caused reduction in the weights of the body and the ovary and increased the appearance of atretic follicles in immature female Coho salmon Oncorhynchus kisutch (Yamamoto et al 2011). Likewise, reduction in the plasma levels of estradiol and the size of eggs as well as the larvae were observed following 6 months of broodstock feeding at a half feed ration in the European seabass D. labrax (Cerda et al 1994).…”

Spatiotemporal Correlations between<b><i> amh</i></b> and <b><i>cyp19a1a</i></b> Transcript Expression and Apoptosis during Gonadal Sex Differentiation of Pejerrey, <b><i>Odontesthes bonariensis</i></b>

“…Dixogenin-11-UTP-labeled riboprobe was synthesized using T7 or SP6 RNA polymerase to generate sense or antisense probes. ISH was carried out using previously described protocols with some modifications [Yamamoto et al, 2011[Yamamoto et al, , 2014. Sections were initially permeabilized with 1 mg/ mL of proteinase K at 37 ° C for 12 min, acetylated, and incubated with 1 mg/mL RNA probe at 65 ° C for 16 h. After hybridization, sections were washed, and unbound probes were digested using 20 μg/mL of RNase A in order to reduce background signals.…”