Characterization of receptors using cyanine 3-labeled neuropeptides

Nigel W. Bunnett, Paul F. Dazin, Donald G. Payan, Eileen F. Grady

Research output: Contribution to journalArticlepeer-review

Abstract

We labeled substance P (SP), neurokinin A (NKA), and [Lys0]gastrin-releasing peptide-27 (GRP) with cyanine 3.18 (cy3). Cy3-peptides purified by HPLC were fully active, determined by [Ca2+]i mobilization. Binding was specific because it was abolished by unlabeled peptides and receptor antagonists. Transfected cells yielded a log-fold greater cy3 intensity than control cells by FACS. Confocal microscopy of transfected cells and cultured enteric neurons showed that cy3-SP bound to surface receptors and was internalized. Internalization was observed in living cells by capture of sequential images. Recovery of surface binding sites was monitored by flow cytometry using cy3-SP. Thus, cy3 neuropeptides can be used to isolate and study receptor-bearing cells.

Original languageEnglish (US)
Pages (from-to)733-740
Number of pages8
JournalPeptides
Volume16
Issue number4
DOIs
StatePublished - 1995

Keywords

  • Endocytosis
  • Flow analysis
  • Fluorescent peptides
  • Gastrin-releasing peptide
  • Neurokinin A
  • Substance P

ASJC Scopus subject areas

  • Biochemistry
  • Physiology
  • Endocrinology
  • Cellular and Molecular Neuroscience

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