Development of an enzyme-linked immunosorbent assay for the measurement of plasma growth hormone (GH) levels in channel catfish ( Ictalurus punctatus): assessment of environmental salinity and GH secretogogues on plasma GH levels

• Published in: General and comparative endocrinology Vol. 133; no. 3; pp. 314 - 322
• Main Authors: Drennon, Katherine, Moriyama, Shunsuke, Kawauchi, Hiroshi, Small, Brian, Silverstein, Jeffrey, Parhar, Ishwar, Shepherd, Brian
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01.10.2003
• Subjects: animal growth; Animals; bioassays; blood plasma; Brackish; catfish; Cattle; Channel catfish; dose response; Dose-Response Relationship, Drug; ELISA; enzyme-linked immunosorbent assay; Enzyme-Linked Immunosorbent Assay - methods; fish culture; Fresh Water - chemistry; Freshwater; GHRH; GHRP-2; Growth hormone; Growth Hormone - blood; Growth Hormone - drug effects; Growth Hormone - immunology; Growth Hormone-Releasing Hormone - agonists; Growth Hormone-Releasing Hormone - blood; Growth Hormone-Releasing Hormone - pharmacology; hormone secretion; hyperosmotic conditions; Ictaluridae - blood; Ictalurus punctatus; in vivo studies; Oligopeptides; osmoregulation; Pituitary Gland - cytology; Pituitary Gland - drug effects; Pituitary Gland - immunology; Prolactin - blood; Rabbits; Salinity; Seawater - chemistry; somatotropin; Teleostei; GHRP-2; Channel catfish; Growth hormone; GHRH; ELISA; Salinity
• ISSN: 0016-6480; 1095-6840; 1095-6840
• DOI: 10.1016/S0016-6480(03)00194-1

We report the development of a sensitive, and specific, competitive, antigen-capture enzyme-linked immunosorbent assay for the measurement of channel catfish ( Ictalurus punctatus) growth hormone (cfGH). The detection limit of the assay (90% binding) was 2.0 ng/ml and the ED 50 value (standard curve range 150–0.59 ng/ml) was 67.3 ng/ml. Recovery of cfGH-spiked plasma samples was determined to be 102%. Dose–response inhibition curves using serially diluted pituitary homogenates and plasma samples consistently showed parallelism with the standard curves using purified cfGH. The GH antibody (rabbit anti-catfish GH) specificity was demonstrated in competitive binding curves employing heterologous hormones and purified channel catfish prolactin (cfPRL). These studies show that there was no significant (0.006%) binding of cfPRL (competitive inhibition of cfGH binding), or heterologous hormones, within the working range of the assay. To physiologically validate the assay, catfish were injected (100 μg/g body weight, 3 injections every 5 days) with either bovine GHRH 1–29-amide or the synthetic hexapeptide GHRP-2 (KP-102: d-Ala- d-β-Nal-Ala-Trp- d-Phe-Lys-NH 2) suspended in corn oil. Following the last injection, half of the animals were sampled for plasma and the remaining transferred from fresh water (FW) to 12 ppt seawater (BW: brackish water). Twenty-four hours after transfer to BW, animals were again sampled for plasma. Plasma GH levels were significantly ( p<0.001) elevated in all the BW groups (control, KP-102, and bGHRH), compared with the FW (fresh water) groups. In addition, plasma GH levels were significantly ( p<0.001) elevated by treatment with either of the GH secretogogues, KP-102 or bGHRH. Our findings demonstrate that two regulatory mechanisms of GH elevation, one which is seen in euryhaline teleosts (salinity-induced GH levels) and another, which has been recently described in teleosts (GHRP-induced GH levels), are present in the stenohaline channel catfish.