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Expression and in vitro regulation of pituitary adenylate cyclase-activating polypeptide (pacap₃₈) and its type I receptor (pac₁-r) in the gonads of tilapia (Oreochromis mossambicus)

Department of Molecular Biotechnology, Da-Yeh University, Chang-Hua 515, Taiwan, ROC¹ Institute of Biotechnology, National Dong-Hwa University, Hualien 974, Taiwan, ROC

Correspondence should be addressed to W-T Huang; Email: kater{at}mail.dyu.edu.tw; C-F Weng; Email: cfweng{at}mail.ndhu.edu.tw)

Pituitary adenylate cyclase-activating polypeptide (PACAP), a pleiotropic neuropeptide, has diverse functions in mammals. However, studies of the expression and function of PACAP and its receptor in fish, particularly in the reproductive system, are still limited. In this report, semi-quantitative RT-PCR and immunohistochemical staining were performed to identify expression domains of commercially important tilapia (Oreochromis mossambicus). PACAP (tpacap₃₈) and its type I receptor (tpac₁-r). Transcripts were detected in the brain, gallbladder, gill, heart, intestine, kidney, muscles, pancreas, spleen, stomach, testes, and ovaries, but not in the liver. Expression of tpacap₃₈ and tpac₁-r mRNA in brain tissue was significantly higher in both sexes compared with other tissues. Addition of exogenous ovine PACAP₃₈ (0.25–5 nM), cAMP analog (dibutyryl-cAMP, 0.25–1.5 mM) or forskolin (adenylate cyclase activator, 1–10 µM) significantly upregulated tpacap₃₈ in the gonads via a dose- and time-dependent fashion. This effect reached a maximal level at 2 h after induction, and then decreased with prolonged culture for up to 4 or 8 h. Additionally, the expression levels of tpac₁-r were not significantly affected by ovine PACAP₃₈ or dibutyryl-cAMP in either sex. Forskolin had a slightly inductive effect and its function could be suppressed with the addition of protein kinase A (PKA) inhibitor, H89 (10 µM), indicating involvement of the cAMP-PKA signaling pathway in the regulation of tpacap₃₈. Expression of tpacap₃₈ and tpac₁-r in the gonads of tilapia suggests that PACAP may mediate gonadotropin action via paracrine/autocrine mechanisms in this bony fish.