Determination of basal phosphodiesterase activity in mouse rod photoreceptors with cGMP clamp

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Abstract

Light regulates cGMP concentration in the photoreceptor cytoplasm by activating phosphodiesterase (PDE) molecules through a G-protein signalling cascade. Spontaneous PDE activity is present in rod outer segments even in darkness. This basal PDE activity (βdark) has not been determined in wild type mammalian photoreceptor cells although it plays a key role in setting the sensitivity and recovery kinetics of rod responses. We present a novel method for determination of βdark using local electroretinography (LERG) from isolated mouse retinas. The method is based on the ability of PDE inhibitors to decrease βdark, which can be counterbalanced by increasing PDE activity with light. This procedure clamps cytoplasmic cGMP to its dark value. βdark can be calculated based on the amount of light needed for the “cGMP clamp” and information extracted from the registered rod photoresponses. Here we apply this method to determine βdark values for the first time in the mammalian rods and obtain the following estimates for different mouse models: 3.9 s−1 for wild type, 4.5 s−1 for guanylate cyclase activating proteins (GCAPs) knockout, and 4.4 s−1 for GCAPs and recoverin double knockout mice. Our results suggest that depletion of GCAPs or recoverin do not affect βdark.

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Original languageEnglish
Article number1183
Pages (from-to)1-14
Number of pages14
JournalScientific Reports
Volume9
Issue number1
Publication statusPublished - 4 Feb 2019
MoE publication typeA1 Journal article-refereed

    Research areas

  • SINGLE-PHOTON RESPONSES, LIGHT-SENSITIVE CURRENT, RHODOPSIN PHOSPHORYLATION, RETINAL RODS, ACTIVATED RHODOPSIN, PHOTOTRANSDUCTION, CONE, PHOTORESPONSES, ADAPTATION, DYNAMICS

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