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Ji Hyun Jeon, Sooyeon Sim, Sun-Sook Paik, Myung-Hoon Chun, In-Beom Kim; TMEM16A/ANO1, A Ca2+-Dependent Chloride Channel is Expressed in ON Bipolar Cells of the Goldfish Retina. Invest. Ophthalmol. Vis. Sci. 2011;52(14):1161. doi: https://doi.org/.
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A chloride conductance activated by intracellular Ca2+ was first reported in salamander rod photoreceptors. It has subsequently been observed in cones and ON bipolar cells isolated from goldfish retina. However, the molecular identity of the channel is unknown. Recently, a transmembrane protein, TMEM16A, also known as anoctamin 1 (ANO1), has been identified as a Ca2+-dependent chloride channel (CaCC) and it is expressed in nervous tissue such as retina and dorsal root ganglia. Thus, we tested whether TMEM16A/ANO1 is responsible for the Ca2+-dependent chloride current in goldfish bipolar cells.
Bipolar cells were isolated from the retina of goldfish (Carassius auratus) and membrane currents were recorded in the whole-cell recording configuration. Single- and double-label experiments with antibodies against TMEM16A/ANO1 and PKC, the ON bipolar cell marker, were also performed.
In the soma and axon-terminal of ON bipolar cells, depolarization evoked an inward calcium current and a slowly declining tail current. To examine whether the tail current was due to chloride flux, the reversal potential for Cl- was measured and a Cl- channel blocker was applied. The measured reversal potential was almost same as the expected ECl value and the extracellular application of 2 mM SITS to cells blocked the tail current but not the calcium current. TMEM16A/ANO1 immunoreactivity was found in many bipolar cells with somas in the inner nuclear layer (INL). Both plexiform layers, the outer plexiform layer (OPL) and the inner plexiform layer (IPL) contained immunoreactive puncta. In addition, the TMEM16A/ANO1-immunoreactive bipolar cells were also labeled for PKC.
Our electrophysiological results confirm that ON bipolar cells have a Ca2+-dependent chloride tail current and our immunofluorescent results suggest that TMEM16A/ANO1 might be a strong candidate for the CaCC which generates the tail current in ON bipolar cells of the goldfish retina.
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