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A. Bosco, A. Wurm, G.P. Nicchia, A. Frigeri, A. Rendon, A. Reichenbach, D.C. Spray; Aquaporin–4 Regulates Horizontal Cell Volume in Developing Mouse Retina . Invest. Ophthalmol. Vis. Sci. 2006;47(13):4198.
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The recent discovery of aquaporin–4 (AQP4) and inwardly rectifying K+ (Kir4.1) channels in the embryonic and perinatal mouse retina suggests that its ionic and osmotic homeostasis may be already regulated during retinal development, well before vision starts. Differentiating horizontal cells specifically coexpress AQP4 and Kir4.1 channels from embryonic day 18.5th until eye opening (postnatal day 15, P15) when both channels become confined to Müller glial cell perivascular membranes [Bosco et al., 2005]. In the present study we have investigated whether horizontal cell AQP4 channels are functional in postnatal mice, and therefore competent to control the developing retina's extracellular space balance.
Neural retinal flat mounts from P5 mice were loaded with a solution containing 200 nM AQP4 or rat AQP1siRNA (silencing control) and 3% Alexa Fluor 594 nm–conjugated dextran (MW 10,000 kDa) by scrape–loading method [Cusato et al., 2003], cultured overnight, and sliced to image the dextran–loaded horizontal cell somata by confocal microscopy. Horizontal cell volume regulation (swelling) was tested in whole retina cultures after overnight AQP4 mRNA silencing, as well as in the silencing control retinas. For each horizontal cell, the soma area was measured during perfusion with isotonic solution, and thereafter, under hypotonic stress, with or without blockage of K+ channels by Ba2+ [Pannicke et al., 2004]. AQP4 and neurofilament 150 kDa were immunodetected and AQP4–fluorescence levels quantified.
Control horizontal cells from postnatal retina did not swell upon hypotonic stress, unless their K+ channels were blocked. AQP4 expression knock down prevented the hypotonic horizontal cell swelling evoked by K+ channel blockage.
Like Müller cells, young horizontal cells effectively regulate their cell volume with a transmembrane water flux coupled to K+ currents. Their AQP4 channels provide a rapid mechanism to free these differentiating neurons of surplus osmolarity via water flux.
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