In Vitro Phosphorylation in Isolated Horizontal Cells of the Fish Retina: Effects of the State of Light Adaptation

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Abstract
Horizontal cells, which are second‐order neurons of the vertebrate retina, exhibit synaptic plasticity governed by light and dark adaptation. We have investigated the alterations in the protein phosphorylation patterns of isolated carp (Cyprinus carpio) horizontal cells in relation to their state of light adaptation by using an in vitro phosphorylation assay and compared the resulting data with protein synthesis patterns of the whole retina. Phosphoproteins and [35S]methionine‐labelled proteins were analysed by one‐ and two‐dimensional gel electrophoresis followed by autoradiography. The state of light adaptation significantly affected the in vitro phosphorylation of horizontal cell proteins with molecular weights of 68, 56/58, 47, 28 and 15 kDa, but had no effect on the protein synthesis of retinal proteins. In the light the most prominent increase of 32P incorporation was observed in the 47 kDa protein. The biochemical properties of this protein closely resembled those of the growth‐associated GAP‐48, found in the fish retina. In addition, the phosphorylation of horizontal cell homogenates in the presence of protein kinase activators such as cyclic AMP, calcium, calmodulin and phospholipids revealed that horizontal cells of the fish retina contain cyclic AMP‐, calcium/calmodulin‐ and calcium/phospholipid‐dependent protein kinase activity resulting in the phosphorylation of several horizontal cell proteins, including the phosphoproteins which were affected by the state of light adaptation.