Effects of cysteamine hydrochloride (CSH)-a somatostatin-inhibiting agent-on serum growth hormone (GH) levels and growth in juvenile grass carp (Ctenopharyngodon idellus) were studied. Intraperitoneal (i.p.) injection and single or 10-day feeding of different doses of CSH significantly increased serum GH levels. CSH and luteinizing hormonereleasing hormone analog (LHRH-A, D-Ala6,Pro9-Net-LHRH), alone and in combination i.p. injection, and single or 10-day administration in diet resulted in an enhancement of serum GH contents; in addition, there was an additive, not synergistic effect of CSH and LHRH-A on elevation of serum GH levels. Ten day feeding of CSH, or CSH and LHRHA, alone and in combination caused a significant increase in muscle RNAyDNA ratio. These results provide evidence that CSH significantly increases serum GH levels and promotes short-term growth in juvenile grass carp.

A full-length cDNA encoding the neuropeptide Y (NPY) was cloned from the hypothalamus of orange spotted grouper (Epinepheluscoioides) by rapid amplification of cDNA ends approaches. The NPY cDNA sequence is 688 bp long and has an open reading frame of 300 bp encoding prepro-NPY with 99 amino acids. The deduced amino acid sequences contain a 28-amino-acids signal peptide followed by a 36- amino-acids mature NPY peptide. mRNA expression of NPY was determined using semi-quantitative RT-PCR followed by Southern blot analysis. NPY mRNA was expressed in olfactory bulb, telencephalon, pituitary, hypothalamus, optic tectum-thalamus, medulla oblongata, cerebellum and spinal cord. Low levels of NPY mRNA expression were found in retina, ovary and stomach, while much lower levels of expression were detected in liver, heart, gill, skin, anterior intestine, thymus and blood. No NPY mRNA expression was observed in unfertilized eggs, newly fertilized eggs, 16-cells stage and morula stage of the embryo and lower levels of expression were detected in the blastula, gastrula and neurula stages. It was highly expressed from lens formation stage to 52-day-old larval stage. NPY might be involved in the late embryonic and larval development of the orange spotted grouper.

The effects of [D-Ala6,Pro9-NEt]-LHRH (LHRH-A) alone and in combination with drugs which influence the actions of dopamine or the synthesis of catecholamines on gonadotropin (GtH) secretion and ovulation in the loach (Paramisgurnus dabryanus) were investigated. LHRH-A alone stimulated an increase in serum GtH levels in the loach, but was a relatively ineffective treatment for the induction of ovulation. Injection of the dopamine receptor antagonist pimozide caused a marked potentiation of the GtH-release response to LHRH-A, and combined injections of pimozide and LHRH-A were an effective treatment for the induction of ovulation. Reserpine, a drug which causes depletion of catecholamines from presynaptic terminals, also caused a marked potentiation of the GtH-release response to LHRH-A and combined treatment induced ovulation. Similarly, administration of alpha-methyl-para-tyrosine to block conversion of tyrosine to L-dopa, or carbidopa to block conversion of L-dopa to dopamine, potentiated the GtH-release response to LHRH-A and induced ovulation. In ontrast, the use of diethyldithiocarbamate, to block conversion of dopamine to norepinephrine, failed to augment the action of LHRH-A on GtH release and ovulation. The present results provide further vidence to suggest that dopamine functions as a gonadotropin release-inhibitory factor in teleosts, and demonstrate that the use of drugs which block either the synthesis or the actions of dopamine potentiates the action of LHRH-A in teleosts.

The effects of mammalian GnRH analog (D-Ala6, Pro9-NEt-LHRH), salmon GnRH, (Trp7, Leti8)-LHRH and its analog, (D-Arg6, Pro9-NEt)-sGnRH on growth hormone (GH) release were examined using a perifusion system for pituitary fragments of the common carp (Cyprinus carpio). Perifusion of 2 min pulses of different concentrations of LHRH-A, sGnRH or sGnRH-A stimulated a rapid and dose-dependent increase in GH release; sGnRH-A showed a significantly greater effect than LHRHA in stimulating GH release. Administration of LHRH-A in the diet ( 1 or 10pg g-t diet) for 5 weeks resulted in a highly significant increase in the growth rate of juvenile grass carp (Crenopharyngodon idellus). Implantation of oestradiol potentiated basal GH secretion in sexually-mature (pre-spawning) and sexually-regressed fish, whereas testosterone implantation did not affect the serum GH level. Both testosterone and oestradiol treatment potentiated the serum GH response to sGnRH-A; however, testosterone was significantly less potent than oestradiol in this respect. The serum GH response to sGnRH-A after testosterone and oestradiol implantation varied seasonally, with greater response in sexually-mature and sexually-regressed fish than those in early stages of ovarian development. These results suggested that sex steroids probably potentiate GH secretion via the influence on the action of GnRH on GH release.

Insulin-like growth factors I and II (IGF-I and IGF-II) are two highly homologous mitogenic peptides that are expressed ubiquitously and show diverse effects on development, growth, and metabolism. The cDNA encoding IGF-I of a teleost, the orange spotted grouper (Epinephelus coioides) was produced from liver by RT-PCR, and rapid amplification of cDNA ends, RACE. Typically, the deduced 186 amino acid protein contains a signal peptide, B, C, A, D and E domains. On the amino acid level, grouper IGF-I shares 97.3% similarity with black seabream (Sparus macrocephalus) with the differences focusing on the B and C domains. The analysis of the E domain showed that grouper IGF-I belonged to Ea-4 type. When mature amino acid sequence was compared with other vertebrates, it revealed higher similarity with black seabream and halibut, while lower similarity with human and mouse. The expression of IGF-I mRNA in adult tissues was studied using RT-PCR. IGF-I mRNA expression level in the liver was significantly higher than those in the brain and muscles. In other tissues, low amount of IGF-I mRNA expression was also detected. The coding region of IGF-I cDNA for mature IGF-I protein was subcloned into an expression plasmid pTRX and fused with E. coli thioredoxin (Trx). Moreover, we have successfully developed an expression system in E. coli to overproduce recombinant grouper IGF-I. Using western blotting, we found that the fusion protein could blot with antiserum to barramundi IGF-I further confirming the immunoactivity of the recombinant IGF-I.