| Budget Amount *help |
¥3,710,000 (Direct Cost: ¥3,500,000、Indirect Cost: ¥210,000)
Fiscal Year 2007: ¥910,000 (Direct Cost: ¥700,000、Indirect Cost: ¥210,000)
Fiscal Year 2006: ¥2,800,000 (Direct Cost: ¥2,800,000)
|
|---|
| Research Abstract |
Feedback from horizontal cells (HCs) to cone photoreceptors plays a key role in the center-surround-receptive field organization of retinal neurons. Recordings from cone photoreceptors in newt retinal slices were obtained by the whole-cell patch-clamp technique, using a superfusate containing a GABA antagonist (100 μM picrotoxin). Surround illumination of the receptive field increased the voltage-dependent calcium current (I_<Ca>) in the cones, and shifted the activation voltage of I_<Ca> to negative voltages. External alkalinization also increased cone I_<Ca> and shifted its activation voltage toward negative voltages. Enrichment of the pH buffering capacity of the extracellular solution increased cone I_<Ca>, and blocked any additional increase in cone I_<Ca> by surround illumination. Hyperpolarization of the HCs by a glutamate receptor antagonist augmented cone I_<Ca>, whereas depolarization of the HCs by kainate suppressed cone I_<Ca>.
From these results, we propose that pH changes
… More
in the synaptic clefts, which are intimately related to the membrane voltage of the HCs, mediate the feedback from the HCs to cone photoreceptors. To verify the idea that depolarized HCs released protons, we measured, by a fluorescence ratio imaging technique, the pH of the immediate external surface (pH_a) of HCs isolated from the carp or goldfish retina. When HCs stained by 5-hexadecanoylaminofluorescein, a pH-sensitive lipophilic dye, were depolarized by bath-application of kainate or high-K^+ medium, pHa was lowered. The amount of pH_s change was monotonically dependent on the amount of depolarization, as much as 0.21±0.05 pH unit by 100 mV depolarization (induced by 100 mM K^+). Acidification was suppressed by 400 nM bafilomycin Al, a specific inhibitor of vacuolar type H^+ pump (V-ATPase), suggesting that protons were released from HCs by the voltage-sensitive H^+ pump by depolarization. Immunocytochemical analysis, using an and V-ATPase antibody, revealed the existence of the V-ATPase on the plasma membrane of dissociated HCs. These results confirm that protons are released from HCs and support the hypothesis that the feedback from HCs to cones could be proton-mediated. Less
|
