Localization and age-related changes of nitric oxide- and ANP-mediated cyclic-GMP synthesis in rat cervical spinal cord: an immunocytochemical study

An immunocytochemical technique was used to study the localization and deve lopmental aspects of cyclic GMP (cGMP)-synthesizing structures in the: cerv ical spinal cord of 2-week and 3-month-old Lewis rats in response to the ni tric oxide (NO) donor sodium nitroprusside (SNP) and/or atrial natriuretic peptide (ANP). By using cell-specific markers, the cell structures involved were investigated. To visualize cGMP, a combined technique of low- and hig h-power magnification, using a confocal laser scanning microscope was used. NOS-mediated cGMP synthesis was observed in the cervical spinal cord in la minae I, II and III in 14-day-old rats, which activity was mainly absent at the age of 3 months. The involvement of NO in the NMDA-mediated increase i n cGMP immunostaining (cGMP-IS) was demonstrated by the absence of cGMP-IS in slices incubated in the presence of NMDA together with the NOS inhibitor NG-nitro-L-arginine methyl ester (L-NAME). This NO-mediated effect of NMDA on cGMP-IS was completely absent in the 3-month-old rats. ANP-mediated cGM P synthesis resulted in an increase in cGMP in laminae I and II, which was generally similar at both ages. Astrocytes in both white and gray matter we re found to be cGMP-IS in the basal, NO- and ANP-stimulated conditions. Usi ng confocal laser microscopy, NO-mediated cGMP synthesis was observed in la rge cholinergic terminals nearby motor neurons in the ventral hem. An exten sive colocalization between NO-stimulated cGMP synthesis and parvalbumin-po sitive (GABAergic) neurons and fibers was observed in all laminae. In the A NP-stimulated condition, a colocalization with parvalbumin structures was f ound in laminae II and III. No NO- or ANP-mediated cGMP synthesis was found in fibers immunopositive for the presynaptic glutamate transporter, seroto nin, or tyrosine hydroxylase. (C) 2000 Elsevier Science B.V. All rights res erved.