Acute and chronic effects of opioids on delta and mu receptor activation of G proteins in NG108-15 and SK-N-SH cell membranes

J Neurochem. 1997 Apr;68(4):1462-72. doi: 10.1046/j.1471-4159.1997.68041462.x.

Abstract

To compare activation of G proteins by opioid receptors, opioid agonist-stimulated guanosine 5'-O-(3-[35S]thiotriphosphate) ([35S]GTP gamma S) binding in the presence of excess GDP was assayed in membranes from NG108-15 (delta) and SK-N-SH (primarily mu) cells. Basal [35S]GTP gamma S binding consisted of a single class of low-affinity sites (KD 400-500 nM). Addition of agonists produced a high-affinity site 100-300-fold higher in affinity than the basal site. The receptor/transducer amplification factor (ratio of activated G protein Bmax to opioid receptor Bmax) was 10-fold higher for SK-N-SH mu receptors than for NG108-15 delta receptors. Chronic delta agonist ([D-Ser2]-Leu-enkephalin-Thr; DSLET) treatment of NG108-15 cells resulted in an 80% loss of DSLET-stimulated [35S]-GTP gamma S binding within 1 h. Morphine treatment of SK-N-SH cells decreased mu agonist ([D-Ala2, N-Me-Phe4,Gly5-ol]-enkephalin; DAMGO)-stimulated [35S]GTP gamma S binding by 45% after 16 h, with no effect after 1 h. Loss of agonist response was due to a decrease in the Bmax of activated G proteins with no change in the KD. These results provide a quantitative description of G protein activation occurring on acute and chronic exposure to opioid agonists.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Analgesics / pharmacology
  • Animals
  • Enkephalin, Ala(2)-MePhe(4)-Gly(5)-
  • Enkephalin, Leucine / analogs & derivatives
  • Enkephalin, Leucine / pharmacology
  • Enkephalins / pharmacology
  • GTP-Binding Proteins / drug effects
  • GTP-Binding Proteins / metabolism*
  • Glioma
  • Guanosine 5'-O-(3-Thiotriphosphate) / metabolism
  • Humans
  • Hybrid Cells / chemistry
  • Hybrid Cells / drug effects
  • Hybrid Cells / metabolism
  • Membrane Proteins / drug effects
  • Membrane Proteins / metabolism
  • Mice
  • Naloxone / pharmacology
  • Naltrexone / analogs & derivatives
  • Naltrexone / pharmacology
  • Narcotic Antagonists / pharmacology
  • Narcotics / pharmacology*
  • Neuroblastoma
  • Protein Binding / drug effects
  • Rats
  • Receptors, Opioid, delta / agonists
  • Receptors, Opioid, delta / drug effects
  • Receptors, Opioid, delta / metabolism*
  • Receptors, Opioid, mu / agonists
  • Receptors, Opioid, mu / drug effects
  • Receptors, Opioid, mu / metabolism*
  • Signal Transduction / drug effects
  • Sulfur Radioisotopes
  • Time Factors

Substances

  • Analgesics
  • Enkephalins
  • Membrane Proteins
  • Narcotic Antagonists
  • Narcotics
  • Receptors, Opioid, delta
  • Receptors, Opioid, mu
  • Sulfur Radioisotopes
  • Enkephalin, Ala(2)-MePhe(4)-Gly(5)-
  • Naloxone
  • Guanosine 5'-O-(3-Thiotriphosphate)
  • Enkephalin, Leucine
  • Naltrexone
  • enkephalin, Ser(2), Leu(5), Thr(6)-
  • GTP-Binding Proteins
  • naltrindole