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J Physiol Volume 585, Number 2, 401-412, December 1, 2007 DOI: 10.1113/jphysiol.2007.142646
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NEUROSCIENCE

Depolarization of isolated horizontal cells of fish acidifies their immediate surrounding by activating V-ATPase

 Hiroshi Jouhou1,2, Kazunori Yamamoto1, Akinori Homma1, Masayuki Hara1, Akimichi Kaneko3 and Masahiro Yamada1

1 Department of Biological Sciences, Tokyo Metropolitan University, Hachioji, Tokyo, Japan
2 Astellas Pharmaceutical Inc., Yodogawa, Osaka, Japan
3 School of Rehabilitation, Seijoh University, Aichi, and School of Health Science, Kio University, Koryo-cho, Nara, Japan

In order to interpret the formation of receptive field surrounds in retinal neurons, a proton-mediated mechanism was proposed to mediate feedback from horizontal cells (HCs) to cone photoreceptors. To verify the idea that depolarized HCs release protons, we measured, by a fluorescence ratiometric technique, the pH of the immediate external surface (pHs) 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, pHs was lowered. The amount of pHs change was monotonically dependent on the degree of depolarization, as much as 0.21 ± 0.05 pH units by 100 mV depolarization (induced by 100 mM K+). Acidification was suppressed by 400 nM bafilomycin A1, a specific inhibitor of the vacuolar type H+ pump (V-ATPase), suggesting that depolarization released protons from HCs via the voltage-sensitive H+ pump. Immunocytochemical analysis, using an anti-V-ATPase antibody, revealed the existence of V-ATPase in dissociated HCs. These results support the hypothesis that the feedback from HCs to cones could be proton mediated.

(Received 7 August 2007; accepted after revision 3 October 2007; first published online 11 October 2007)
Corresponding author M. Yamada: Department of Biological Sciences, Tokyo Metropolitan University, Minami-ohsawa, Hachioji, Tokyo 192-0397, Japan. Email: myamada{at}tmu.ac.jp

H. Jouhou and K. Yamamoto contributed equally to this work.






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