Nitric oxide production and the expression of two nitric oxide synthases in the avian retina

Nitric oxide (NO) is known to exert multiple effects on the function of many retinal neurons and their synapses. Therefore, it is equally important to understand the potential sources of NO within the retina. To explore this, we employ a combination of 4-amino-5-methylamino-2',7'-difluorofluorescein diacetate (DAF-FM) based NO detection and immunohistochemistry for the NO synthetic enzymes, neuronal and endothelial nitric oxide synthase (nNOS and eNOS). We find DAF signals in photoreceptors, horizontal cells, amacrine cells, efferent synapses, Müller cells, and cells in the ganglion cell layer (GCL). nNOS immunoreactivity was consistent with the DAF signal with the exception that horizontal cells and Müller cells were not clearly labeled. eNOS-like immunoreactivity (eNOS-LI) was more widespread with photoreceptors, horizontal cells, occasional bipolar cells, amacrine cells, Müller cells, and cells in the GCL all showing labeling. Double labeling with antibodies raised against calretinin, syntaxin, and glutamine synthetase confirmed that horizontal cells, amacrine cells, and Müller cells (respectively) were expressing eNOS-LI. Although little or no nNOS labeling is observed in horizontal cells or Müller cells, the expression of eNOS-LI is consistent with the ability of these cells to produce NO. Together these results suggest that the capability to produce NO is widespread in the chicken retina. We propose that multiple forms of regulation for nNOS and eNOS play a role in the patterning of NO production in the chicken retina.