Nicotine attenuates activation of tissue resident macrophages in the mouse stomach through the β2 nicotinic acetylcholine receptor.
BACKGROUND
The cholinergic anti-inflammatory pathway is an endogenous mechanism by which the autonomic nervous system attenuates macrophage activation via nicotinic acetylcholine receptors (nAChR). This concept has however not been demonstrated at a cellular level in intact tissue. To this end, we have studied the effect of nicotine on the activation of resident macrophages in a mouse stomach preparation by means of calcium imaging.


METHODS
Calcium transients ([Ca(2+)]i) in resident macrophages were recorded in a mouse stomach preparation containing myenteric plexus and muscle layers by Fluo-4. Activation of macrophages was achieved by focal puff administration of ATP. The effects of nicotine on activation of macrophages were evaluated and the nAChR involved was pharmacologically characterized. The proximity of cholinergic nerves to macrophages was quantified by confocal microscopy. Expression of β2 and α7 nAChR was evaluated by β2 immunohistochemistry and fluorophore-tagged α-bungarotoxin.


RESULTS
In 83% of macrophages cholinergic varicose nerve fibers were detected at distances <900 nm. The ATP induced [Ca(2+)]i increase was significantly inhibited in 65% or 55% of macrophages by 100 µM or 10 µM nicotine, respectively. This inhibitory effect was reversed by the β2 nAChR preferring antagonist dihydro-β-eryhtroidine but not by hexamethonium (non-selective nAChR-antagonist), mecamylamine (α3β4 nAChR-preferring antagonist), α-bungarotoxin or methyllycaconitine (both α7 nAChR-preferring antagonist). Macrophages in the stomach express β2 but not α7 nAChR at protein level, while those in the intestine express both receptor subunits.


CONCLUSIONS
This study is the first in situ demonstration of an inhibition of macrophage activation by nicotine suggesting functional signaling between cholinergic neurons and macrophages in the stomach. The data suggest that the β2 subunit of the nAChR is critically involved in the nicotine-induced inhibition of these resident macrophages.
Department of Clinical and Experimental Medicine, Translational Research Center for Gastrointestinal Disorders (TARGID), University of Leuven, Leuven, Belgium.
Andrea
A
Nemethova
Klaus
K
Michel
Pedro J
PJ
Gomez-Pinilla
Guy E
GE
Boeckxstaens
Michael
M
Schemann
D000255:Adenosine Triphosphate / Q000494:pharmacology; D000814:Aniline Compounds / Q000378:metabolism; D000818:Animals; D002118:Calcium / Q000378:metabolism; D017615:Enteric Nervous System / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D005731:Ganglionic Stimulants / Q000494:pharmacology; D005753:Gastric Mucosa / Q000378:metabolism; D007150:Immunohistochemistry; D008262:Macrophage Activation / Q000187:drug effects; D008264:Macrophages / Q000187:drug effects* / Q000378:metabolism; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D018613:Microscopy, Confocal; D009197:Myenteric Plexus / Q000187:drug effects / Q000378:metabolism; D009474:Neurons / Q000187:drug effects / Q000378:metabolism; D009538:Nicotine / Q000494:pharmacology*; D011978:Receptors, Nicotinic / Q000378:metabolism*; D013270:Stomach / Q000187:drug effects* / Q000294:innervation; D014966:Xanthenes / Q000378:metabolism
D000255:Adenosine Triphosphate; D000814:Aniline Compounds; D002118:Calcium; D005731:Ganglionic Stimulants; D009538:Nicotine; D011978:Receptors, Nicotinic; D014966:Xanthenes; C409648:Fluo 4