Glutamate receptors modulate sodium‐dependent and calcium‐independent vitamin C bidirectional transport in cultured avian retinal cells

Abstract

Vitamin C is transported in the brain by sodium vitamin C co‐transporter 2 (SVCT‐2) for ascorbate and glucose transporters for dehydroascorbate. Here we have studied the expression of SVCT‐2 and the uptake and release of [14C] ascorbate in chick retinal cells. SVCT‐2 immunoreactivity was detected in rat and chick retina, specially in amacrine cells and in cells in the ganglion cell layer. Accordingly, SVCT‐2 was expressed in cultured retinal neurons, but not in glial cells. [14C] ascorbate uptake was saturable and inhibited by sulfinpyrazone or sodium‐free medium, but not by treatments that inhibit dehydroascorbate transport. Glutamate‐stimulated vitamin C release was not inhibited by the glutamate transport inhibitor l‐β‐threo‐benzylaspartate, indicating that vitamin C release was not mediated by glutamate uptake. Also, ascorbate had no effect on [3H] d‐aspartate release, ruling out a glutamate/ascorbate exchange mechanism. 2‐Carboxy‐3‐carboxymethyl‐4‐isopropenylpyrrolidine (Kainate) or NMDA stimulated the release, effects blocked by their respective antagonists 6,7‐initroquinoxaline‐2,3‐dione (DNQX) or (5R,2S)‐(1)‐5‐methyl‐10,11‐dihydro‐5H‐dibenzo[a,d]cyclohepten‐5,10‐imine hydrogen maleate (MK‐801). However, DNQX, but not MK‐801 or 2‐amino‐5‐phosphonopentanoic acid (APV), blocked the stimulation by glutamate. Interestingly, DNQX prevented the stimulation by NMDA, suggesting that the effect of NMDA was mediated by glutamate release and stimulation of non‐NMDA receptors. The effect of glutamate was neither dependent on external calcium nor inhibited by 1,2‐bis (2‐aminophenoxy) ethane‐N′,N′,N′,N′,‐tetraacetic acid tetrakis (acetoxy‐methyl ester) (BAPTA‐AM), an internal calcium chelator, but was inhibited by sulfinpyrazone or by the absence of sodium. In conclusion, retinal cells take up and release vitamin C, probably through SVCT‐2, and the release can be stimulated by NMDA or non‐NMDA glutamate receptors.

Nitric Oxide Modulates Sodium Vitamin C Transporter 2 (SVCT-2) Protein Expression via Protein Kinase G (PKG) and Nuclear Factor-κB (NF-κB)

“…Previous studies have demonstrated the presence of SVCT-2 transcripts in the inner nuclear layer of the rat retina (3). Moreover, presence of the SVCT-2 protein has been described in the inner nuclear and ganglion cell layers of the chicken retina (13). In cultured chicken retinal cells it has been demonstrated that ascorbate release is modulated by ionotropic glutamate receptors and proposed that ion flux through AMPA/kainate receptors promotes SVCT-2 reversal (13).…”

Caveolin-1–mediated internalization of the vitamin C transporter SVCT2 in microglia triggers an inflammatory phenotype

“…This assay was done exactly as previous described (86). N9 cells previously treated with LPS (1 µg/ml; 24 hours) were rinsed twice with HBSS, containing 140 mM NaCl, 5 mM KCl, 20 mM Hepes, 4 mM glucose, 1 mM MgCl2, and 2 mM CaCl2 (pH 7.4) and then incubated with [ 14 C]ascorbate (0.1 µCi/ml) for 40 min.…”

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

“…In line with this, high-affinity proteins called sodium vitamin C co-transporters (SVCTs) stereospecifically transport ascorbate in a Na + -dependent manner using two Na + ions for each transported ascorbate molecule [106,107]. Previous studies showed the presence of SVCT-2 transcripts in the inner nuclear layer of the rat retina [107] and our group demonstrated the presence of SVCT-2 in both cultured chick retinal cells and post-hatched chick retina [90,108].…”