an extracellular alkalization induced by 1 glutamatergic activation of isolated retinal horizontal cells

35 36 37 Extracellular acidification induced by retinal horizontal cells has been 38 hypothesized to underlie lateral feedback inhibition onto vertebrate 39 photoreceptors. To test this hypothesis, the H-sensitive fluorophore HAF (540 hexadecanoylaminofluorescein) was used to measure changes in H from 41 horizontal cells isolated from the retina of the catfish. HAF staining conditions 42 were modified to minimize intracellular accumulation of HAF and maximize 43 membrane-associated staining, and ratiometric fluorescent imaging of cells 44 displaying primarily membrane-associated HAF fluorescence was conducted. 45 Challenge of such HAF-labeled cells with glutamate or the ionotropic glutamate 46 receptor agonist kainate produced an increase in the fluorescence ratio, 47 consistent with an alkalinization response of +0.12 pH units and +0.23 pH units, 48 respectively. This alkalinization was blocked by the AMPA receptor antagonist 49 CNQX, the L-type calcium channel blocker nifedipine, and lanthanum. The 50 alkalinization reported by HAF was consistent with extracellular alkalinizations 51 detected in previous studies using self-referencing H-selective microelectrodes. 52 The spatial distribution of the kainate-induced changes in extracellular H was 53 also examined. An overall global alkalinization around the cell was observed, 54 with no obvious signs of discrete centers of acidification. Taken together these 55 data argue against the hypothesis that glutamatergic-induced efflux of protons 56 from horizontal cells mediates lateral feedback inhibition in the outer retina. 57 58 59 60 61 62