Abstract
Abstract: :
Purpose: We have established that activation of the NO–cGMP signaling pathway stimulates presynaptic plasticity in cone photoreceptors but inhibits that in rod photoreceptors. In the present experiments, we investigated the expression of soluble guanylyl cyclase (sGC), the synthetic enzyme for cGMP, and the sensitivity of the enzyme to pharmacological manipulation. Methods: For detection of sGC, isolated adult salamander photoreceptors were incubated for 2 hours or 3 days followed by immunostaining for α– and ß–subunits of sGC with avidin–biotin complex technique. For the detection of cGMP production, retinal cells cultured for 1, 3 or 5 days were lysed with PCA and the cGMP concentration in the lysates was determined by radioimmunoassay. For the detection of NO production, nitrite in culture supernatant was measured with the Griess reaction. Results:The staining for both α– and ß–subunits of sGC could be observed in both cone and rod cells, and was primarily in the cell soma. The staining in 3–day–old photoreceptors was stronger than that in 2–hour–old photoreceptors. cGMP and NO were continuously produced in control cultures. 100µM SNAP, which elevated NO levels at day one, increased cGMP in retinal cultures which remained high for 5 days. 1 µM of sGC activator YC–1 significantly increased cGMP production by retinal cells cultured for 1 day, though this increase was not significant after 3 days. 50µM of sGC inhibitor ODQ, however, significantly decreased cGMP production in retinal cells cultured for 3 days. Conclusions: (1) sGC, present in cultured photoreceptors, appears to be upregulated with time, and is sensitive to stimulation by NO and YC–1 or inhibition by ODQ; (2) the continuous activity of the NO–cGMP signaling pathway coincides with the reported growth of cones while rod cell growth is not prevented at basal levels; (3) however, the pharmacological increase of cGMP levels correlates with observed stimulation of cone growth and inhibition of rod growth by sGC activators (Townes–Anderson and Zhang, Soc. For Neuroscience Abstract 2002) and confirms the importance of cGMP in photoreceptor synaptic plasticity.
Keywords: second messengers: pharmacology/physiology • plasticity • photoreceptors