Dataset Information

Immunocytochemical localization of synaptic proteins to photoreceptor synapses of Drosophila melanogaster.

ABSTRACT: The location of proteins that contribute to synaptic function has been widely studied in vertebrate synapses, far more than at model synapses of the genetically manipulable fruit fly, Drosophila melanogaster. Drosophila photoreceptor terminals have been extensively exploited to characterize the actions of synaptic genes, and their distinct and repetitive synaptic ultrastructure is anatomically well suited for such studies. Synaptic release sites include a bipartite T-bar ribbon, comprising a platform surmounting a pedestal. So far, little is known about the composition and precise location of proteins at either the T-bar ribbon or its associated synaptic organelles, knowledge of which is required to understand many details of synaptic function. We studied the localization of candidate proteins to pre- or postsynaptic organelles, by using immuno-electron microscopy with the pre-embedding method, after first validating immunolabeling by confocal microscopy. We used monoclonal antibodies against Bruchpilot, epidermal growth factor receptor pathway substrate clone 15 (EPS-15), and cysteine string protein (CSP), all raised against a fly head homogenate, as well as sea urchin kinesin (antibody SUK4) and Discs large (DLG). All these antibodies labeled distinct synaptic structures in photoreceptor terminals in the first optic neuropil, the lamina, as did rabbit anti-DPAK (Drosophila p21 activated kinase) and anti-Dynamin. Validating reports from light microscopy, immunoreactivity to Bruchpilot localized to the edge of the platform, and immunoreactivity to SUK4 localized to the pedestal of the T-bar ribbon. Anti-DLG recognized the photoreceptor head of capitate projections, invaginating organelles from surrounding glia. For synaptic vesicles, immunoreactivity to EPS-15 localized to sites of endocytosis, and anti-CSP labeled vesicles lying close to the T-bar ribbon. These results provide markers for synaptic sites, and a basis for further functional studies.

SUBMITTER: Hamanaka Y

PROVIDER: S-EPMC4029604 | biostudies-literature | 2010 Apr

REPOSITORIES: biostudies-literature

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Immunocytochemical localization of synaptic proteins to photoreceptor synapses of Drosophila melanogaster.

Hamanaka Yoshitaka Y Meinertzhagen Ian A IA

The Journal of comparative neurology 20100401 7

The location of proteins that contribute to synaptic function has been widely studied in vertebrate synapses, far more than at model synapses of the genetically manipulable fruit fly, Drosophila melanogaster. Drosophila photoreceptor terminals have been extensively exploited to characterize the actions of synaptic genes, and their distinct and repetitive synaptic ultrastructure is anatomically well suited for such studies. Synaptic release sites include a bipartite T-bar ribbon, comprising a pla ...[more]

PMID: 20127822

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Project description:Synaptic signaling complexes are held together by scaffold proteins, each of which is selectively capable of interacting with a number of other proteins. In previous studies of rabbit retina, we found Synapse-Associated Protein-102 (SAP102) and Channel Associated Protein of Synapse-110 (Chapsyn110) selectively localized in the tips of horizontal cell processes at contacts with rod and cone photoreceptors, along with several interacting ion channels. We have examined the equivalent suites of proteins in mouse retina and found similarities and differences. In the mouse retina we identified Chapsyn110 as the scaffold selectively localized in the tips of horizontal cells contacting photoreceptors, with Sap102 more diffusely present. As in rabbit, the inward rectifier potassium channel Kir2.1 was present with Chapsyn110 on the tips of horizontal cell dendrites within photoreceptor invaginations, where it could provide a hyperpolarization-activated current that could contribute to ephaptic signaling in the photoreceptor synapses. Pannexin 1 and Pannexin 2, thought to play a role in ephaptic and/or pH mediated signaling, were present in the outer plexiform layer, but likely not in the horizontal cells. Polyamines regulate many ion channels and control the degree of rectification of Kir2.1 by imposing a voltage-dependent block. During the day polyamine immunolabeling was unexpectedly high in photoreceptor terminals compared to other areas of the retina. This content was significantly lower at night, when polyamine content was predominantly in Müller glia, indicating daily rhythms of polyamine content. Both rod and cone terminals displayed the same rhythm. While polyamine content was not prominent in horizontal cells, if polyamines are released, they may regulate the activity of Kir2.1 channels located in the tips of HCs. The rhythmic change in polyamine content of photoreceptor terminals suggests that a daily rhythm tunes the behavior of suites of ion channels within the photoreceptor synapses.

Dataset Information

Immunocytochemical localization of synaptic proteins to photoreceptor synapses of Drosophila melanogaster.

Publications

Immunocytochemical localization of synaptic proteins to photoreceptor synapses of Drosophila melanogaster.

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