Inhibition of protein synthesis by activation of NMDA receptors in cultured retinal cells: a new mechanism for the regulation of nitric oxide production

Abstract

The synthesis of nitric oxide (NO) is limited by the intracellular availability of L-arginine. Here we show that stimulation of NMDA receptors promotes an increase of intracellular L-arginine which supports an increase in the production of NO In the last few years the biochemistry and metabolism of L-arginine (L-Arg) have been refocused due to the great importance attached to nitric oxide (NO) physiology (Wiesinger 2001). L-Arg is required for the synthesis of several molecules including polyamines, creatine, agmatine and NO and is a major constituent of proteins, and the metabolic overlapping of these reactions remains to be explored. In all cases the production of these compounds depends on the intracellular concentration of L-Arg. Two pathways have been described: (i) synthesis of L-Arg from different substrates and/or (ii) transport of L-Arg to intracellular compartments. The transport of cationic amino acids across cell membranes is mediated by a group of well-described transport systems (y + , y ) that take up L-Arg with different properties (sodium dependence, affinity and capacity). The transport is mediated predominantly by the ubiquitous y + system, which promotes high-affinity L-Arg uptake in a sodium-independent manner (MacLeod and

Consolidation and translation regulation: Figure 1.

“…Short physiological glutamatergic stimulus in primary cortical neurons induces activation of ERK and mTOR/S6K via calcium/CaM signaling controlled by voltage-dependent calcium channels and is NMDAR independent. Continuous glutamatergic stimulus triggers high-calcium influx, leading to a progressive increase in eEF2 phosphorylation and inhibition of translation as shown in cortical neurons, cultured retinal cells of chicks, and in hippocampal slices from rats (Marin et al 1997;Scheetz et al 2000;Belelovsky et al 2005;Lenz and Avruch 2005;Cossenza et al 2006;Maus et al 2006). The regulation of eEF2 by glutamate receptors is discussed in further detail below.…”

Synaptic Activity Controls Dendritic Spine Morphology by Modulating eEF2-Dependent BDNF Synthesis

“…Glutamate receptor activation modulates eEF2 phosphorylation (Marin et al, 1997;Scheetz et al, 2000;Cossenza et al, 2006;Kanhema et al, 2006;Sutton et al, 2007), and physical and functional interactions between eEF2K and mGluR1 have been documented (Park et al, 2008). We therefore investigated the glutamate receptor subtypes involved in the eEF2K activation (that sustains eEF2 phosphorylation) caused by long-term stimulation with bicuculline.…”

Studying Nitric Oxide in the Developing Retina: Neuromodulatory Functions and Signaling Mechanisms

“…More than 70% of intracellular taken up L-Arg was incorporated into proteins and when this incorporation was blocked by CHX, the free intracellular L-Arg pool increased significantly. Unexpectedly, when protein synthesis was inhibited, the intracellular L-Cit production increased and the extracellular content of L-Cit increased even more [23]. This particular phenomenon drew our attention to two facts: (1) High levels of L-Cit found in the extracellular environment could be related with some releasing system working in concert with intercellular recycling pathways.…”

“…Dopamine-coupled cAMP production was studied during chick development and also in monolayer cultures [20,21]. Adenosine and different types of adenosine receptors, modulating adenylyl cyclase activity, were also detected in the developing chick retina [22,23]. Adenosine-elicited cAMP accumulation was detected in the chick retina since embryonic day 14, attaining a maximum effect at day 17 and decreasing in the post-hatching period [22].…”

“…Since L-Arg is an important amino acid present in proteins, we decided to use the protein synthesis inhibitor cyclohexymide (CHX) during uptake assays to avoid the protein synthesisdependent component in L-Arg uptake [23]. Interestingly, we demonstrated that protein synthesis represented an important driving force for L-Arg uptake [23]. More than 70% of intracellular taken up L-Arg was incorporated into proteins and when this incorporation was blocked by CHX, the free intracellular L-Arg pool increased significantly.…”