Forskolin modulates photoresponses in mouse rods but not as strongly as in amphibian photoreceptors

Abstract

Purpose: Intracellular [cAMP] is regulated by circadian rhythm. The level of cAMP peaks at night and decreases during the daytime (Chaurasia et al., J. Neurochem., 2006). The increase in [cAMP] can be mimicked by introducing the adenylate cyclase activator, forskolin, which accelerates the synthesis of cAMP. Photoreceptor sensitivity and kinetics have recently been shown to be regulated by cAMP via multiple targets in the phototransduction cascade in amphibian rods (Astakhova et al., J. Gen. Physiol., 2012). Single cell recordings on Rana ridibunda rods suggest that forskolin increases intracellular Ca²⁺ and decreases basal PDE activity. This is seen as delayed deactivation kinetics and over twofold increase in rod sensitivity. The object of this study was to find out whether similar kinds of effects by forskolin are present also in mice rods.

Methods: Dark-adapted ERG responses to green LED light flashes were recorded from WT and GCAP^-/- mice (C57BL/6J). Isolated retina was perfused with modified Ringer’s solution at 37±1°C containing BaCl₂, DL-AP4 and/or aspartate to isolate rod photoresponses. 5 or 10 µM forskolin dissolved in DMSO was introduced in the perfusion solution for 20 minutes before solution was changed back to normal. Saturated and small stimulus responses were followed throughout the experiment. ERG responses were simultaneously recorded transretinally (TERG) and across the outer segment layer with microelectrodes (LERG).

Results: Forskolin increased the saturated photoresponse amplitude by 40-50 % in CGAP^-/- mice, while only a minor increase was observed in WT mice. The maximal effect was achieved within 10 minutes after introducing forskolin. With TERG fractional small stimulus responses of WT and CGAP^-/- rods grew 30 % and 50 % respectively, while 10 % and 20 % increases were seen in LERG recordings. Also in CGAP^-/- mice the time constant of small response recovery (τ_rec) as well as the time-to-peak increased by 10-20 % in LERG recordings.

Conclusions: Forskolin caused notable effects in CGAP^-/- mouse rod sensitivity and time-to-peak which seemed to be a result from a delayed deactivation of responses. The effects were considerably smaller than in amphibian photoreceptors. The growth in the saturation amplitude observed in CGAP^-/- mice rods can be due to the lack of Ca²⁺ feedback to guanylate cyclase activity.

Keywords: 648 photoreceptors • 510 electroretinography: non-clinical • 711 second messengers: pharmacology/physiology