Visualization of gaseous monoxide reception by soluble guanylate cyclase in the rat retina

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Abstract
Immunohistochemistry using novel monoclonal antibodies (mAbs) allowed us to uncover tissue activities of soluble guanylate cyclase (sGC) fine tuned by NO and CO. Upon NO and CO applications in vitro, purified sGC increased the affinity to mAb3221 by 100- and 10-fold, respectively, but not to mAb28131. Immunohistochemistry for gas-generating enzymes revealed that NO occurred in amacrine, bipolar, and Müller's glia cells (MGCs), whereas CO was derived mostly from heme oxygenase (HO)-2 in MGCs. Basal sGC immunoreactivities in vivo to mAb3221 but not to mAb28131 were enhanced by injecting L-arginine and attenuated by blocking NO synthases, suggesting the ability of the former mAb to sense NO. Comparison of mAb-assisted immunohistochemistry suggested that sGC activities were enhanced by zinc protoporphyrin-IX, an HO inhibitor, and repressed completely by blocking NO. However, suggested roles of CO played in situ varied among different retinal layers. In inner plexiform and inner nuclear layers located in the proximity of the cellular NO sources, CO serves as a simple inhibitor of local sGC, while playing roles in housekeeping sGC activation in external limiting membrane standing far from them. These results suggest that CO generated in MGCs is a diffusible gas mediator regulating sGC in both autocrine and paracrine manners.
Funding Information
  • Japan Society for the Promotion of Science (13680850)