Grass carp somatolactin: II. Pharmacological study on postreceptor signaling mechanisms for PACAP-induced somatolactin-α and -β gene expression

Abstract

Somatolactin (SL), the latest member of the growth hormone/prolactin family, is a novel pituitary hormone with diverse functions. However, the signal transduction mechanisms responsible for SL expression are still largely unknown. Using grass carp as an animal model, we examined the direct effects of pituitary adenylate cyclase-activating polypeptide (PACAP) on SL gene expression at the pituitary level. In primary cultures of grass carp pituitary cells, SL␣ and SL␤ mRNA levels could be elevated by PACAP via activation of PAC-I receptors. With the use of a pharmacological approach, the AC/cAMP/PKA and PLC/inositol 1,4,5-trisphosphate (IP 3)/PKC pathways and subsequent activation of the Ca 2ϩ /calmodulin (CaM)/CaMK-II cascades were shown to be involved in PACAP-induced SL␣ mRNA expression. Apparently, the downstream Ca 2ϩ /CaM-dependent cascades were triggered by extracellular Ca 2ϩ ([Ca 2ϩ ]e) entry via L-type voltagesensitive Ca 2ϩ channels (VSCC) and Ca 2ϩ release from IP3-sensitive intracellular Ca 2ϩ stores. In addition, the VSCC component could be activated by cAMP/PKA-and PLC/PKC-dependent mechanisms. Similar postreceptor signaling cascades were also observed for PACAP-induced SL␤ mRNA expression, except that [Ca 2ϩ ]e entry through VSCC, PKC coupling to PLC, and subsequent activation of CaMK-II were not involved. These findings, taken together, provide evidence for the first time that PACAP can induce SL␣ and SL␤ gene expression in fish model via PAC-I receptors through differential coupling to overlapping and yet distinct signaling pathways.pituitary adenylate cyclase-activating polypeptide; grass carp pituitary cells SOMATOLACTIN (SL), the latest member of the growth hormone (GH)/prolactin (PRL) family, can be identified only in fish but not in tetrapods (37). Phylogenetic analysis reveals that SL is derived from ancestral GH (16), and gene duplication in bony fish have given rise to two paralogous SL isoforms, namely SL␣ and SL␤ (47). Unlike GH and PRL, which are nonglycosylated and expressed mainly in the anterior pituitary, SL is a glycoprotein secreted by the periodic acid Schiff-positive cells located in the pars intermedia (15). Although the physiological functions of SL have not been fully characterized, its involvement in color presentation (7), body metabolism (8), osmoregulation (18), stress responses (30), and reproductive functions (31) has been proposed by various studies in fish models. Recently, it has been shown that swim bladder development can be inhibited in zebrafish by SL␤ but not SL␣ gene silencing, implying that the two SL isoforms may act differently during the process of embryo organogenesis (46). In rainbow trout, SL release can be suppressed by dopamine but stimulated by corticotropin-releasing hormone and gonadotropin-releasing hormone (GnRH; Ref. 13). In addition, SL mRNA expression can be upregulated by GnRH treatment (e.g., salmon; Ref. 26), which is in agreement with the previous reports (28) that GnRH receptors are expressed in fish pituitary cells with SL immunoreact...

Insulin-like growth factor as a novel stimulator for somatolactin secretion and synthesis in carp pituitary cells via activation of MAPK cascades

“…In rat pituitary GH4C1 cells, IGF-induced PRL secretion is mediated by the PI3K/Akt pathway (40), while ERK1/2 via subsequent activation of Ets-1 is involved in the gene expression response (7). Although our recent studies in grass carp have shown that cAMP/PKA and PLC/IP3/PKC pathways are involved in PACAP-induced SL␣ and SL␤ gene expression (19), the functional role of MAPK and PI3K/Akt cascades in SL release and synthesis have not been examined. In carp NIL cells, rapid phosphorylation of the signaling kinases, including MEK1/2, ERK1/2, MKK3/6, and p38 MAPK, were observed after IGF treatment.…”

“…Limited in vitro studies based on pituitary organ culture or pituitary cells suggest that SL release can be stimulated by leptin in sea bass (37), by gonadotropin-releasing hormone in rainbow trout (22), and by melanin-concentrating hormone in cichlid (5) and goldfish (45). Recently, pituitary adenylate cyclase-activiting polypeptide (PACAP), a potent GH-releasing factor in fish (50), was shown to stimulate SL gene expression in goldfish (31) and carp pituitary cells (20) via cAMP/PKA-and PLC/PKC-dependent pathways (3,19). These findings, as a whole, indicate that SL secretion and synthesis are regulated by both central and peripheral signals similar to that of other pituitary hormones.…”

Signal transduction mechanisms for autocrine/paracrine regulation of somatolactin-α secretion and synthesis in carp pituitary cells by somatolactin-α and -β

“…Grass carp pituitary cells were prepared by trypsin-DNase digestion method as described previously (15). After enzyme digestion, pituitary cells were seeded in 24-well clustered plates at a density of 2.5 ϫ 10 6 cells per well and cultured overnight at 28°C in carp MEM (MEM edium supplemented with 26 mM NaHCO3, 25 mM HEPES, 100 U/ml penicillin, 100 g/ml streptomycin, and 250 ng/ml fungizone, pH 7.7) with 5% FBS.…”

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)

“…It has been recently demonstrated that PACAP can induce somatolactin (SL) a and SLb gene expression in the pituitary cells of grass carp (Ctenopharyngodon idella) by first binding to PAC-I receptors. This activates the AC/cAMP/protein kinase A (PKA) and phospholipase C (PLC)/inositol 1, 4, 5-trisphosphate (IP 3 )/protein kinase C pathways and involved in the Ca 2C /calmodulin (CaM)/CaMK-II cascades (Jiang et al 2008a(Jiang et al , 2008b.…”