Skip to main content
NCBI home page
Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1991 May 1;88(9):3540–3544. doi: 10.1073/pnas.88.9.3540

Isolation and characterization of two peptides with prolactin release-inhibiting activity from porcine hypothalami.

A V Schally 1, J G Guoth 1, T W Redding 1, K Groot 1, H Rodriguez 1, E Szonyi 1, J Stults 1, K Nikolics 1
PMCID: PMC51487  PMID: 2023899

Abstract

Two peptides with in vitro prolactin release-inhibiting activity were purified from stalk median eminence (SME) fragments of 20,000 pig hypothalami. Monolayer cultures of rat anterior pituitary cells were incubated with aliquots of chromatographic fractions and the inhibition of release of prolactin in vitro was measured by RIA in order to monitor the purification. The hypothalamic tissue extract was separated into 11 fractions by high-performance aqueous size-exclusion chromatography with one fraction showing a 4-fold increase in prolactin release-inhibiting factor (PIF) activity. This material was further purified by semipreparative reversed-phase (RP) HPLC. This process resulted in the separation of two distinct fractions that showed high PIF activity. These were further purified by semipreparative and analytical RP-HPLC to apparent homogeneity as judged by the UV absorbance profiles. Neither of the two peptides showed cross-reactivity with gonadotropin releasing hormone-associated peptide or with somatostatin-14 antibodies. Protein sequence analysis revealed that one of the PIF peptides was Trp-Cys-Leu-Glu-Ser-Ser-Gln-Cys-Gln-Asp-Leu-Ser-Thr-Glu-Ser-Asn-Leu-Leu- Ala-Cys - Ile-Arg-Ala-Cys-Lys-Pro, identical to residues 27-52 of the N-terminal region of the proopiomelanocortin (POMC) precursor (corresponding to amino acids 1-26 of the 16-kDa fragment). The sequence of the other PIF was Ala-Ser-Asp-Arg-Ser-Asn-Ala-Thr-Leu-Leu-Asp-Gly-Pro-Ser-Gly-Ala-Leu-Leu- Leu-Arg - Leu-Val-Gln-Leu-Ala-Gly-Ala-Pro-Glu-Pro-Ala-Glu-Pro-Ala-Gln-Pro-Gly-Val- Tyr, representing residues 109-147 of the vasopressin-neurophysin precursor. Synthetic peptides corresponding to the N-terminal region of POMC had significant PIF activity in vitro.

Full text

PDF
3540

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Acher R., Chauvet J. Structure, processing and evolution of the neurohypophysial hormone-neurophysin precursors. Biochimie. 1988 Sep;70(9):1197–1207. doi: 10.1016/0300-9084(88)90185-x. [DOI] [PubMed] [Google Scholar]
  2. Arimura A., Lundqvist G., Rothman J., Chang R., Fernandez-Durango R., Elde R., Coy D. H., Schally A. V., Meyers C. Radioimmunoassay of somatostatin. Metabolism. 1978 Sep;27(9 Suppl 1):1139–1144. doi: 10.1016/0026-0495(78)90032-x. [DOI] [PubMed] [Google Scholar]
  3. Bennett H. P. Isolation and characterization of the 1 to 49 amino-terminal sequence of pro-opiomelanocortin from bovine posterior pituitaries. Biochem Biophys Res Commun. 1984 Nov 30;125(1):229–236. doi: 10.1016/s0006-291x(84)80358-7. [DOI] [PubMed] [Google Scholar]
  4. Bernton E. W., Beach J. E., Holaday J. W., Smallridge R. C., Fein H. G. Release of multiple hormones by a direct action of interleukin-1 on pituitary cells. Science. 1987 Oct 23;238(4826):519–521. doi: 10.1126/science.2821620. [DOI] [PubMed] [Google Scholar]
  5. Enjalbert A., Moos F., Carbonell L., Priam M., Kordon C. Prolactin inhibiting activity of dopamine-free subcellular fractions from rat mediobasal hypothalamus. Neuroendocrinology. 1977;24(3-4):147–161. doi: 10.1159/000122759. [DOI] [PubMed] [Google Scholar]
  6. Esch F., Böhlen P., Ling N., Brazeau P., Guillemin R. Isolation and characterization of the bovine hypothalamic growth hormone releasing factor. Biochem Biophys Res Commun. 1983 Dec 28;117(3):772–779. doi: 10.1016/0006-291x(83)91664-9. [DOI] [PubMed] [Google Scholar]
  7. Greibrokk T., Hansen J., Knudsen R., Lam Y. K., Folkers K., Bowers C. Y. On the isolation of a prolactin inhibiting factor (hormone). Biochem Biophys Res Commun. 1975 Nov 3;67(1):338–344. doi: 10.1016/0006-291x(75)90321-6. [DOI] [PubMed] [Google Scholar]
  8. Henzel W. J., Rodriguez H., Watanabe C. Computer analysis of automated Edman degradation and amino acid analysis data. J Chromatogr. 1987 Aug 28;404(1):41–52. doi: 10.1016/s0021-9673(01)86835-7. [DOI] [PubMed] [Google Scholar]
  9. Hökfelt T. The possible ultrastructural identification of tubero-infundibular dopamine containing nerve endings in the median eminence of the rat. Brain Res. 1967 May;5(1):121–123. doi: 10.1016/0006-8993(67)90225-9. [DOI] [PubMed] [Google Scholar]
  10. McCann S. M. Physiology and pharmacology of LHRH and somatostatin. Annu Rev Pharmacol Toxicol. 1982;22:491–515. doi: 10.1146/annurev.pa.22.040182.002423. [DOI] [PubMed] [Google Scholar]
  11. Meites J., Clemens J. A. Hypothalamic control of prolactin secretion. Vitam Horm. 1972;30:165–221. doi: 10.1016/s0083-6729(08)60796-7. [DOI] [PubMed] [Google Scholar]
  12. Mizunuma H., Khorram O., McCann S. M. Purification of a non-dopaminergic and non-GABAergic prolactin release-inhibiting factor (PIF) in sheep stalk-median eminence. Proc Soc Exp Biol Med. 1985 Jan;178(1):114–120. doi: 10.3181/00379727-178-41991. [DOI] [PubMed] [Google Scholar]
  13. Nagy G., Mulchahey J. J., Smyth D. G., Neill J. D. The glycopeptide moiety of vasopressin-neurophysin precursor is neurohypophysial prolactin releasing factor. Biochem Biophys Res Commun. 1988 Feb 29;151(1):524–529. doi: 10.1016/0006-291x(88)90625-0. [DOI] [PubMed] [Google Scholar]
  14. Nikolics K., Mason A. J., Szönyi E., Ramachandran J., Seeburg P. H. A prolactin-inhibiting factor within the precursor for human gonadotropin-releasing hormone. Nature. 1985 Aug 8;316(6028):511–517. doi: 10.1038/316511a0. [DOI] [PubMed] [Google Scholar]
  15. PASTEELS J. L. [Administration of hypothalamic extracts to the rat pituitary in vitro, with a view to controlling the secretion of prolactin]. C R Hebd Seances Acad Sci. 1962 Apr 2;254:2664–2666. [PubMed] [Google Scholar]
  16. Plotsky P. M., Gibbs D. M., Neill J. D. Liquid chromatographic-electrochemical measurement of dopamine in hypophysial stalk blood of rats. Endocrinology. 1978 Jun;102(6):1887–1894. doi: 10.1210/endo-102-6-1887. [DOI] [PubMed] [Google Scholar]
  17. SCHALLY A. V., KUROSHIMA A., ISHIDA Y., REDDING T. W., BOWERS C. Y. THE PRESENCE OF PROLACTIN INHIBITING FACTOR (PIF) IN EXTRACTS OF BEEF, SHEEP AND PIG HYPOTHALAMI. Proc Soc Exp Biol Med. 1965 Feb;118:350–352. doi: 10.3181/00379727-118-29839. [DOI] [PubMed] [Google Scholar]
  18. Schally A. V., Coy D. H., Meyers C. A. Hypothalamic regulatory hormones. Annu Rev Biochem. 1978;47:89–128. doi: 10.1146/annurev.bi.47.070178.000513. [DOI] [PubMed] [Google Scholar]
  19. Schally A. V., Dupont A., Arimura A., Takahara J., Redding T. W., Clemens J., Shaar C. Purification of a catecholamine-rich fraction with prolactin release-inhibiting factor (PIF) activity from porcine hypothalami. Acta Endocrinol (Copenh) 1976 May;82(1):1–14. doi: 10.1530/acta.0.0820001. [DOI] [PubMed] [Google Scholar]
  20. Schally A. V., Redding T. W., Arimura A., Dupont A., Linthicum G. L. Isolation of gamma-amino butyric acid from pig hypothalami and demonstration of its prolactin release-inhibiting (PIF) activity in vivo and in vitro. Endocrinology. 1977 Mar;100(3):681–691. doi: 10.1210/endo-100-3-681. [DOI] [PubMed] [Google Scholar]
  21. Seger M. A., Bennett H. P. Structure and bioactivity of the amino-terminal fragment of pro-opiomelanocortin. J Steroid Biochem. 1986 Nov;25(5B):703–710. doi: 10.1016/0022-4731(86)90298-0. [DOI] [PubMed] [Google Scholar]
  22. Smyth D. G., Massey D. E. A new glycopeptide in pig, ox and sheep pituitary. Biochem Biophys Res Commun. 1979 Apr 27;87(4):1006–1010. doi: 10.1016/s0006-291x(79)80007-8. [DOI] [PubMed] [Google Scholar]
  23. TALWALKER P. K., RATNER A., MEITES J. IN VITRO INHIBITION OF PITUITARY PROLACTIN SYNTHESIS AND RELEASE BY HYPOTHALAMIC EXTRACT. Am J Physiol. 1963 Aug;205:213–218. doi: 10.1152/ajplegacy.1963.205.2.213. [DOI] [PubMed] [Google Scholar]
  24. Takahara J., Arimura A., Schally A. V. Suppression of prolactin release by a purified porcine PIF preparation and catecholamines infused into a rat hypophysial portal vessel. Endocrinology. 1974 Aug;95(2):462–465. doi: 10.1210/endo-95-2-462. [DOI] [PubMed] [Google Scholar]
  25. Vale W., Rivier C., Brazeau P., Guillemin R. Effects of somatostatin on the secretion of thyrotropin and prolactin. Endocrinology. 1974 Oct;95(4):968–977. doi: 10.1210/endo-95-4-968. [DOI] [PubMed] [Google Scholar]
  26. Wehrenberg W. B., Baird A., Ying S. Y., Rivier C., Ling N., Guillemin R. Multiple stimulation of the adenohypophysis by combinations of hypothalamic releasing factors. Endocrinology. 1984 Jun;114(6):1995–2001. doi: 10.1210/endo-114-6-1995. [DOI] [PubMed] [Google Scholar]
  27. Wormald P. J., Abrahamson M. J., Seeburg P. H., Nikolics K., Millar R. P. Prolactin-inhibiting activity of GnRH associated peptide in cultured human pituitary cells. Clin Endocrinol (Oxf) 1989 Feb;30(2):149–155. doi: 10.1111/j.1365-2265.1989.tb03736.x. [DOI] [PubMed] [Google Scholar]
  28. Yu W. H., Seeburg P. H., Nikolics K., McCann S. M. Gonadotropin-releasing hormone-associated peptide exerts a prolactin-inhibiting and weak gonadotropin-releasing activity in vivo. Endocrinology. 1988 Jul;123(1):390–395. doi: 10.1210/endo-123-1-390. [DOI] [PubMed] [Google Scholar]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

RESOURCES