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Quantifying Agonist Activity at G Protein-coupled Receptors
11:45

Quantifying Agonist Activity at G Protein-coupled Receptors

Published on: December 26, 2011

Hormone-defined cell system for studying G-protein coupled receptor agonist-activated growth modulation: delta-opioid

D Agarwal1, J A Glasel

  • 1Department of Biochemistry, University of Connecticut Health Center, Farmington 06030, USA.

Journal of Cellular Physiology
|April 1, 1997
PubMed

Insights

New NIH/3T3 fibroblast cell lines enable study of G-protein-coupled receptor (GPCR) agonist effects on cell growth. These cell lines, cultured in serum-free conditions, reveal distinct GPCR-specific growth regulation patterns.

Area of Science:

  • Cell Biology
  • Molecular Pharmacology
  • Signal Transduction

Background:

  • G-protein-coupled receptor (GPCR) agonists can alter fibroblast cell growth.
  • Existing cell lines express numerous endogenous GPCRs and are cultured in serum, complicating studies of specific GPCR-mediated growth regulation.
  • Understanding GPCR agonist-induced growth mechanisms requires a simplified experimental system.

Purpose of the Study:

  • To develop novel clonal NIH/3T3 fibroblast cell lines for studying specific GPCR agonist-induced growth regulation.
  • To investigate the effects of delta-opioid and serotonin-5HT2C receptor activation on cell proliferation in a controlled, serum-free environment.

Main Methods:

  • Developed new clonal NIH/3T3 cell lines (3T3DA) stably expressing either delta-opioid or serotonin-5HT2C receptors.
  • Cultured cells in serum-free, hormone-defined medium with insulin as the sole exogenous growth factor.
  • Administered specific GPCR agonists (morphine, serotonin) at micromolar concentrations for up to 6 days.
  • Assessed cell proliferation and cyclic AMP (cAMP) production.
  • Evaluated anchorage-dependent growth by colony formation in soft agar.

Main Results:

  • Distinct GPCR-specific, agonist-activated growth regulation patterns were observed in serum-free, but not serum-supplemented, cultures.
  • Morphine (>10 microM) inhibited delta-opioid receptor-expressing cell growth by 40%.
  • Serotonin (1 microM) stimulated serotonin-5HT2C receptor-expressing cell growth by approximately 100%.
  • Opioid agonist-induced growth inhibition and decreased cAMP production were pertussis toxin-sensitive, confirming classical opioid receptor signaling.
  • Neither untreated nor agonist-treated cells formed colonies in soft agar, indicating retained anchorage-dependent growth.

Conclusions:

  • The novel 3T3DA cell lines provide a robust system for dissecting GPCR-mediated growth control mechanisms.
  • Specific GPCRs can exert distinct, opposing effects on cell proliferation (inhibition by delta-opioid agonists, stimulation by serotonin-5HT2C agonists) under defined conditions.
  • These findings highlight the potential for targeted GPCR modulation in regulating cell growth.

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