In this paper, we investigate the role of muscarinic acetylcholine
receptor (mAChR) activity in the regulation of inducible (i) nitric oxide
synthase (iNOS) expression and activity. The signaling pathway involved is
also examined. These experiments also provide a link between mAChR
activation and the nitric oxide (NO)-dependent regulation of retinal
vascular diameter. The diameter of the retinal vessels at a distance of 1
disc diameter from the center of the optic disc was measured in rats using
digital retinal photography, and both iNOS-mRNA gene expression and NOS
were specifically measured using RT-PCR and [U-14C]
citrulline assays, respectively. Stimulation of M1 and
M3 mAChR with carbachol caused an increase in vessel diameter,
in iNOS-mRNA levels and in NOS activity in the retina. Aminoguanidine, an
inhibitor of iNOS, attenuated all these effects. Inhibitors of
phospholipase C (PLC) and protein kinase C (PKC) but not
calcium/calmodulin (CaM) prevented the muscarinic-dependent increase
in iNOS-mRNA levels. The results obtained suggest that the activation of
mAChR increases retinal vessel diameters by increasing the production of
nitric oxide (NO) through iNOS activation and iNOS-mRNA gene expression.
The mechanism appears to occur secondarily to stimulation of PLC and PKC
enzymatic activity.