Neurochemical organization of the macaque retina: effect of TTX on levels and gene expression of cytochrome oxidase and nitric oxide synthase and on the immunoreactivity of Na+K+ATPase and NMDA receptor subunit I

Abstract

The present study examined the relationship between an important energy-generating enzyme (cytochrome oxidase; CO), a key energy-consuming enzyme (Na+ K+ ATPase) and neurochemicals associated with excitatory glutamatergic synapses (NMDAR1 and neuronal nitric oxide synthase, nNOS) in the adult macaque retina. Polyclonal antibodies against neuronal nitric oxide synthase and N-methyl-D-aspartate receptor subunit I were generated for immunohistochemical examination and labeled sites not previously reported were found. We have also isolated cDNAs for cytochrome oxidase subunits III (mitochondrial-encoded) and IV (nuclear-encoded), as well as for a fragment of neuronal nitric oxide synthase, from a human cDNA library. The distributions of mRNAs of these genes were analyzed by in situ hybridization. We found that three or more of the markers examined coexisted in a number of sites: (a) In the inner segments of photoreceptors, high energy demand for maintaining the dark current was placed by Na+ K+ ATPase. This was partially met by ATP-generating enzymes such as CO. Neuronal NOS was also present there for the synthesis of NO and the cascading event leading to the generation of cGMP and the gating of channels for visual transduction. (b) Both the outer and inner plexiform layers had detectable amounts of all four markers, although the levels varied among them. This was most likely due to the presence of depolarizing glutamatergic synapses arising from photoreceptors and bipolar cells and such synaptic events were energy-demanding. The involvement of NMDA receptors and nNOS in these synaptic layers is strongly implicated in the present study. (c) All four markers were present in the majority of retinal ganglion cells, with some inherent heterogeneity related to intensity and size. Retinal ganglion cells are known to receive excitatory synapses from glutamatergic bipolar cells and are themselves highly active. The presence of both NMDAR1 and nNOS in these cells were verified in the present study and the energy demands related to these synaptic activities were necessarily high. Thus, active ion transporting functions related to synaptic or non-synaptically induced repolarization from the basis for an interrelationship between the neurochemicals/enzymes studied. Finally, (d) all four markers and the gene expression of CO and nNOS in the macaque retina were regulated by neuronal activity.

Immunocytochemical Localization of the Postsynaptic Density Protein PSD-95 in the Mammalian Retina

“…NADPH diaphorase histochemistry, an indicator of nNOS-containing neurons, applied to the retina did not reveal a preferential staining of photoreceptor terminals (Sagar, 1986). Recently, macaque monkey retinal sections have been immunostained for nNOS (Wong-Riley et al, 1998), and strong immunoreactivity was observed in the OPL. Unfortunately, the resolution in the published micrographs is too low to detect the labeling of cone pedicles or rod spherules.…”

“…In the case of the N R2D subunit, Wenzel et al (1997a) reported extrasynaptic labeling of rod bipolar cells. N R1 immunoreactivity was observed in ganglion cells, in amacrine cells, and in both plexiform layers (Wong-Riley et al, 1998). Lo et al (1998) reported the presence of four different splice variants of NR1 and described the immunocytochemical labeling of rod bipolar and Müller cells.…”

“…The N R2A, N R2B, and N R2C subunits were expressed in subsets of amacrine cells; the N R2C subunit also was found at the location of bipolar cell perikarya . The NR1, the N R2A, and the N R2D subunits have been localized immunocytochemically Wenzel et al, 1997a;L o et al, 1998;Wong-Riley, 1998). Strong punctate NR2A immunofluorescence was observed in the I PL, and the application of electron microscopy showed that the N R2A subunit is aggregated at postsynaptic sites .…”

Glucose metabolism in mammalian photoreceptor inner and outer segments

“…22,38,60 Photoreceptors are the most metabolically active cells in the retina and account for the majority of the retinal energy consumption. [1][2][3] Active transport of ions against their concentration and electrical gradients is the largest energy consuming function in all neurons, including photoreceptors.…”

“…[1][2][3][4][5][6][7][8][9][10][11][12][13][14] The major photoreceptor energy substrate in vivo is glucose. [4][5][6][7][8][9][10][11][12][13][14][15][16] Glucose metabolism also performs many non-energetic roles in photoreceptors.…”