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Combinative ligand-receptor interactions: effects of cAMP, epinephrine, and met-enkephalin on RAW264 macrophage
1Department of Biological Sciences, Wayne State University, Detroit, Michigan 48202.
Abstract:
Cell surface ligand-receptor interactions play a central role in the regulation and expression of macrophage function. Included among these macrophage membrane receptors are the beta-adrenergic and opioid receptors. We studied the abilities of epinephrine, met-enkephalin, forskolin, and adenosine 3':5' cyclic monophosphate (cAMP) analogues to affect macrophage morphology, spreading, and adherence. Cell spreading was quantitated by measuring the perimeters of adherent cell images recorded by videomicroscopy. Epinephrine induced a dose-dependent decrease in macrophage spreading; at 10(-5) M epinephrine the mean perimeter was 10.4 +/- 0.3 microns in comparison to 15.0 +/- 1.0 microns for controls. The inhibition of spreading can be blocked by the antagonist propranolol. On the other hand, met-enkephalin induced a dose-dependent increase in macrophage spreading, with a perimeter of 18.5 +/- 1.0 microns at 10(-8) M. Since catecholamines and opioids are simultaneously released from chromaffin cells of the adrenal, we examined the combinative effects due to treatment with both ligands. When macrophages were exposed to 10(-5) M epinephrine and 10(-8) M met-enkephalin, cell morphology and spreading were indistinguishable from that due to 10(-5) M epinephrine alone. The epinephrine dose-response curve in the presence of 10(-8) M met-enkephalin was similar to that of epinephrine alone. The beta-adrenergic receptor is apparently capable of diminishing or abrogating the opioid receptor signal(s). These combinative and epinephrine-mediated effects may be at least partially accounted for by the action of cAMP. Forskolin and the cAMP analogues N6-2'-O-dibutyryladenosine 3':5' cyclic monophosphate (dbcAMP) and 8-bromoadenosine 3':5' cyclic monophosphate (Br-cAMP) affected cell morphology and spreading in the same fashion as epinephrine. These differences in morphology and spreading behavior were accompanied by changes in the distribution of F-actin, as judged by phalladicin staining and fluorescence microscopy. We suggest that cAMP and microfilaments play important roles in receptor-mediated neuroregulation of macrophage function.
Insights
Epinephrine decreases macrophage spreading via beta-adrenergic receptors, while met-enkephalin increases it through opioid receptors. Beta-adrenergic signaling can override opioid receptor effects, involving cyclic adenosine monophosphate (cAMP) and microfilaments in neuroregulation.
Area of Science:
- Immunology
- Cell Biology
- Neuroscience
Background:
- Macrophage function is regulated by cell surface ligand-receptor interactions.
- Beta-adrenergic and opioid receptors are present on macrophage membranes.
- Simultaneous release of catecholamines and opioids from adrenal chromaffin cells suggests combined signaling.
Purpose of the Study:
- To investigate the effects of epinephrine and met-enkephalin on macrophage morphology, spreading, and adherence.
- To examine the combinative effects of simultaneous beta-adrenergic and opioid receptor stimulation.
- To elucidate the role of cyclic adenosine monophosphate (cAMP) and microfilaments in these neuroregulatory processes.
Main Methods:
- Videomicroscopy was used to quantitate macrophage cell spreading by measuring cell perimeters.
- Ligands including epinephrine, met-enkephalin, forskolin, and cAMP analogues were applied to macrophages.
- Antagonist propranolol was used to block beta-adrenergic receptors.
- F-actin distribution was assessed using phalloidin staining and fluorescence microscopy.
Main Results:
- Epinephrine dose-dependently decreased macrophage spreading, an effect blocked by propranolol.
- Met-enkephalin dose-dependently increased macrophage spreading.
- Combined epinephrine and met-enkephalin treatment resulted in spreading similar to epinephrine alone, indicating beta-adrenergic receptor dominance.
- Forskolin and cAMP analogues mimicked epinephrine's effects on spreading and morphology.
- Changes in cell morphology and spreading correlated with alterations in F-actin distribution.
Conclusions:
- Beta-adrenergic receptor signaling can diminish or abrogate opioid receptor signaling in macrophages.
- Cyclic adenosine monophosphate (cAMP) and microfilaments are likely involved in the neuroregulation of macrophage function.
- Ligand-receptor interactions on the macrophage surface mediate neuroimmune responses.